MFPL Research Groups 2014

A joint venture of

2014

RESEARCH GROUPS

M A X F . P E R U T Z L A B O R A T O R I E S

History of the Max F. Perutz Laboratories

2005 2006 2007 2008 2009 2010 2011 2012 2013 2014 2015 2016

Foundation of the Max F. Perutz Laboratories GmbH

more than 148 successfully completed PhD degrees

14 group leaders have joined the MFPL

Foundation of the VBC wide scientific core facility CSF with a broad range of new technologies

Symposium “Crossing Frontiers in Life Sciences” on the occasion of the 100th birthday of Max F. Perutz

as of December 2014

6 ERC-Starting Grants 5 START Awards3 Human Frontier Science Program (HFSP) Grants

MFPL group leaders have won many prestigious research awards:The Max F. Perutz Laboratories (MFPL) are a research and training center of the University of Vienna and the Medical University of Vienna in the field of Molecular Biology at the Vienna Biocenter.

Max Perutz was born in 1914 in Vienna to a family of textile manufacturers who made their fortune during the industrial revolution in the 19th century, through the introduction of mechanical spinning and weaving. He attended the Theresianum, where a perceptive teacher awakened his interest in chemistry.

In 1932 he entered the University of Vienna, but because of the poor prospects for a scientific career in Austria in 1936 he decided to move to the Cavendish Laboratory in Cambridge. After Hitler´s invasion

of Austria, the family business was expropriated, his parents became refugees and his natural choice was to continue his career in Cambridge.

Perutz and his co-workers managed to solve the structure of hemoglobin in 1959. The work was published in Nature in February 1960, and Perutz was awarded the Nobel Prize in Chemistry in 1962 together with John Kendrew, who had solved the structure of myoglobin.

In addition to his studies, Perutz pioneered the new research field of Molecular Biology and was instrumental in founding the Laboratory of Molecular Biology (LMB) in Cambridge, UK. He was also involved in establishing the European Molecular Biology Organization (EMBO) in Heidelberg, Germany.

Max F. Perutz died in February 2002 in Cambridge.

„In science, truth always wins.“ M A X F . P E R U T Z

To honor an extraordinary teacher and scientist, the Max F. Perutz Laboratories were named after Max Ferdinand Perutz, who, together with John C. Kendrew, was awarded the 1962 Nobel Prize in Chemistry for his studies on the structure of globular proteins.

M F P L - 2 0 1 4 R E S E A R C H G R O U P S

2014 Research Groups

Content

Research Groups 4

Publications 62


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Research Groups

Gustav Ammerer Signal transduction and transcriptional regulation in yeast

Manuela Baccarini Deciphering the MAPK pathway in vivo

Andreas Bachmair Protein modifiers in plants and retrotransposon biology

Andrea Barta Post-transcriptional regulation of gene expression in plants

Dieter Blaas Early interactions of viruses with host cells

Udo Bläsi Post-transcriptional regulation in bacteria and archaea

Alexander Dammermann Centriole assembly and function

Thomas Decker Host responses and innate immunity to bacteria

Kristina Djinović-Carugo Structural biology of the cytoskeleton

Gang Dong Structural studies of ciliogenesis

Silke Dorner The regulation of gene expression by small ncRNAs

Roland Foisner Lamins in nuclear organization and human disease

Peter Fuchs Stress response in simple epithelia

Boris Görke Signal transduction and post-transcriptional regulation in model bacteria

Angela Hancock Molecular basis of adaptive evolution

Andreas Hartig Origin and biogenesis of peroxisomes

Marcela Hermann LDL-R gene family, apolipoproteins and lipid transfer

Joachim Hermisson Theoretical population genetics

Reinhold Hofbauer Consequences of carnitine deficiency and CSF-1 inhibition

N.-Erwin Ivessa Protein biogenesis and degradation from the ER

Michael Jantsch Transcriptome diversification through RNA editing

Verena Jantsch Meiosis in Caenorhabditis elegans

Franz Klein Chromosome structure and meiotic recombination

Alwin Köhler Gene expression and chromosome dynamics

Gottfried Köhler Biomolecular optical spectroscopy

Robert Konrat Computational biology and biomolecular NMR spectroscopy

Pavel Kovarik Signaling and gene expression in inflammation

Heinrich Kowalski Molecular and structural biology of picornaviruses

Claudine Kraft Regulation and signaling in autophagy

Karl Kuchler Host-pathogen interactions & mechanisms of fungal virulence

Thomas Leonard Structural biology of lipid-activated signal transduction

Josef Loidl Meiotic chromosome pairing and recombination

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Research at the MFPL is curiosity-driven and spans the field of Molecular and Cell Biology. Most groups investigate basic research questions but a significant number are also active in more applied fields of biology. In 2014, more than 470 people from 44 nations worked at the MFPL.Detailed information about all MFPL research groups, their research focus, list of publications and team can be found on the MFPL website: www.mfpl.ac.at/groups

http://www.mfpl.ac.at/groups


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Sascha Martens Molecular mechanisms of autophagy

Isabella Moll Bacterial stress response and ribosome heterogeneity

Ernst MüllnerErythrocyte (patho)physiology

Johannes Nimpf ApoER2 and VLDL receptor

Egon Ogris PP2A enzyme biogenesis and monoclonal antibodies

Friedrich Propst The neuronal cytoskeleton in axon guidance

Florian Raible Hormonal control of animal energy expenditure

Johann Rotheneder Cell cycle regulation and DNA damage response

Matthias Schäfer RNA modifications: their impact on gene expression and innate immunity

Peter Schlögelhofer Meiotic recombination

Renée Schroeder RNA aptamers that regulate the transcriptome

Christian Seiser Chromatin modifiers in development and disease

Tim Skern Interactions between viruses and cells

Dea Slade DNA damage response

Kristin Tessmar-Raible Lunar periodicity and inner brain photoreceptors

Alipasha Vaziri Dynamics of coupled biological systems: methods and phenomena

Gijs Versteeg Ubiquitin-mediated regulation of immune signaling

Arndt von Haeseler Bioinformatics

Christina Waldsich Exploring RNA folding: from structure to function

Graham Warren Golgi biogenesis

Georg Weitzer Somatic stem cells of the heart

Gerhard Wiche Cytolinker proteins in signaling and disease

Angela Witte ɸCh1, a model for gene regulation in haloalkaliphilic archaea

Ivan Yudushkin Functional imaging of signaling networks

Bojan Žagrović Biophysics of macromolecular interactions

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G U S T A V A M M E R E R

Signal transduction and transcriptional regulation in yeast

In this field, some of the unresolved questions concern the dynamic interactions between different signaling factors and their effectors - e.g. in what cellular context they might happen, how they are controlled by phosphorylation events and how these interactions change during cellular signaling events.

To approach these questions in budding yeast we have established and optimized an enzyme driven protein proximity assay. This assay is based on a mammalian histone methyl-transferase and its highly specific substrate, the N-terminal fragment of histone 3. Apart from successfully characterizing known protein interactions in well studied signal systems such as the high osmolarity response, the mating pathway and the induction of autophagy, we have also generally used this approach for validating protein interactions that have been suggested by mass spectrometry analysis or by genetic data.

Another project deals with the problem of how cell cycle dependent signals coordinate the transcriptional regulation of genes. In this case we have focused on the regulation of the mitotic cyclin gene CLB2. This gene is repressed in the G1-phase of the cell cycle, it is de-repressed at the START of S-phase and fully induced by a positive feedback mechanism during G2-phase and mitosis. Moreover, it has been found that its expression is modulated by checkpoint pathways such as in-duced by genotoxic stresses or cell wall damage. We have addressed questions of how phosphory-lation events affect stability and function of im-portant cell cycle specific transcription factors, and how their specific modifications can be correlated with changes in the underlying chromatin structure and chromatin modification patterns.

Gustav Ammerer

TEAMJillian AugustineJessica FerrariIsabella Hansmann David HollensteinAleksandra JovanovicWolfgang ReiterJiri Veis

One of our major aims is to understand the cogs and wheels of phosphorylation-modulated signal transduction machineries in the yeast S. cerevisiae.

SELECTED PUBLICATIONSYelamanchi SK, Veis J, Anrather D, Klug H, Ammerer G. Genotoxic Stress Prevents Ndd1-Dependent Transcriptional Activation of G2/M-Specific Genes in Saccharomyces cerevisiae. Mol. Cell. Biol. 2014;34(4):711-24. PMID: 24324010 Reiter W, Klopf E, De Wever V, Anrather D, Petryshyn A, Roetzer A, Niederacher G, Roitinger E, Dohnal I, Görner W, Mechtler K, Brocard A, Schüller C, Ammerer G. Yeast protein phosphatase 2A-Cdc55 regulates the transcriptional response to hyperosmolarity stress by regulating Msn2 and Msn4 chromatin recruit-ment. Mol. Cell. Biol. 2013;33(5):1057-72. PMID: 23275436 Reiter W, Anrather D, Dohnal I, Pichler P, Veis J, Grøtli M, Posas F, Ammerer G. Validation of regulated protein phosphorylation events in yeast by quantitative mass spectrometry analysis of purified proteins. Proteomics 2012;12(19-20):3030-43. PMID: 22890988

RNA FISH of the CTT1 gene before and after stress. DNA blue, RNA green. Notice the nuclear RNA signal in the middle panel documenting active transcription of the CTT1 gene.

http://www.ncbi.nlm.nih.gov/pubmed/24324010




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M A N U E L A B A C C A R I N I

Deciphering the MAPK pathway in vivo

Focusing specifically on the two best known Raf kinases, B-Raf and C-Raf, we have discovered that B-Raf is the essential ERK activator in all instances in which its ablation results in a phenotype. In contrast, C-Raf mediates pathway cross-talk in-dependently of its kinase activity, by binding to, and inhibiting, the cytoskeleton-based Rok-alpha, which controls cell shape, motility, and in some cell types differentiation.

These non-redundant functions of Raf are best illustrated in a Ras-driven skin carcinogenesis model, in which B-Raf drives MEK/ERK activation and proliferation. In contrast, C-Raf works as an endogenous Rok inhibitor essential for blocking differentiation. Elimination of B-Raf and C-Raf from the epidermis enforces abrupt tumor regression, combining delayed proliferation and increased dif-ferentiation. These results emphasize that therapies targeting both Raf kinase-dependent and -indepen-dent pathways may potentially be more effective and less prone to inducing adverse effects and/or resistance.

Through its interaction with Rok-alpha, C-Raf also regulates the cohesion and the collective migration of endothelial cells, and ultimately sprouting and tumor-induced angiogenesis. Down-stream of the small GTPase Rap1, C-Raf is recruited to VE-cadherin containing adherens junctions, where it is required to anchor Rok-alpha and modu-late junctional actomyosin during adherens junction maturation.

We have also established MEK1 as the critical negative regulator of both MEK2/ERK and PIP3 signaling. MEK1 mediates the regulation of MEK2 in the context of a MEK1:MEK2 heterodimer that is

negatively regulated by ERK-mediated phosphoryla-tion of MEK1. The same ERK-mediated phosphorylation also regulates the formation of a trimeric complex among MEK1, the adaptor protein MAGI1, and the lipid and protein phosphatase PTEN. If this complex is not formed, PTEN is not recruited to the membrane, leading to the accumulation of PIP3 and to the activation of the parallel Akt pathway.

By showing that their essential in vivo functions are fundamentally different, these results have changed the way we look at Raf and MEK kinases, beginning to reveal how the pathway is wired in vivo.

Manuela Baccarini

TEAMBertram AschenbrennerChristian BaumgartnerClemens BognerTania BrandstötterStefan BuchleitnerAnna Lina CavalloBotond CsehEnrico DesideriKarin EhrenreiterInes JericSanya Eduarda KuzetSilvia Munico MartinezJoanna NowackaCristiana PelorossoJosipa RaguzElena RodionovaStefanie ToiflAndrea Varga

Happy together? A trimeric complex consisting of MEK1, the adaptor protein MAGI1 and the lipid and protein phosphatase PTEN is necessary for the regulation of PIP3 signaling (see text for details).

SELECTED PUBLICATIONSWimmer R, Cseh B, Maier B, Scherrer K, Baccarini M. Angiogenic sprouting requires the fine tuning of endothelial cell cohesion by the Raf-1/Rok-α complex. Dev Cell. 2012;22(1):158-71. PMID: 22209329 Catalanotti F, Reyes G, Jesenberger V, Galabova-Kovacs G, de Matos Simoes R, Carugo O, Baccarini M. A Mek1-Mek2 heterodimer determines the strength and duration of the Erk signal. Nat Struct Mol Biol. 2009;16(3):294-303. PMID: 19219045 Zmajkovicova K, Jesenberger V, Catalanotti F, Baumgartner C, Reyes G, Baccarini M. MEK1 is required for PTEN membrane recruitment, AKT regulation, and the maintenance of peripheral tolerance. Mol Cell. 2013;50(1):43-55. PMID: 23453810

The Raf kinases A-, B-, and C-Raf (Raf-1) and their target MEK are attractive therapeutic targets. Yet, surprisingly little is known about their essential functions in the context of the whole organism.

http://www.ncbi.nlm.nih.gov/pubmed/?term=Angiogenic+sprouting+requires+the+fine+tuning+of+endothelial+cell+cohesion+by+the+Raf-1%2FRok-%CE%B1+complex

http://www.ncbi.nlm.nih.gov/pubmed/?term=19219045



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A N D R E A S B A C H M A I R

Protein modifiers in plants and retrotransposon biology

Covalent attachment of ubiquitin to substrate proteins is essential for many processes. Best known is its role in protein degradation. Several ubiquitin moieties, linked as a chain to the substrate protein, can serve as a signal for rapid proteolytic destruction of the sub-strate. However, ubiquitylation also has non-proteolytic functions. One ubiquitylation complex of interest to us operates at the cell membrane. The ubiquitin chains formed by this complex have a predominantly regulatory role, and seem to influence the activity state of certain membrane proteins. This process is important for correct establishment of the plant architecture.

Recently we discovered enzymes that extend single SUMO residues into chains. Together with previously known ubiquitin ligases that bind to SUMO chains, to attach a ubiquitin chain, these enzymes establish a pathway that leads from (mono)sumoylation of substrates via SUMO chain formation to ubiquitin-dependent proteolysis. Mu-tants in this pathway can better cope with salt stress, but less well with osmotic stress, suggesting that the pathway is important for reaction to environmental inputs.

Another project is dealing with retrotrans-posons, We want to convert retrotransposon Tto1 into a novel tool for plant investigation and im-

provement. On the way to this goal, we are learning a lot about the life cycle of this retrotransposon. Transposition of ret-rotransposons is usually induced by stress, and therefore not easy to control. We have modified Tto1 such that its transpositional activity can be induced by the chemical substances Estradiol or Dexamethasone. This novel regulatory circuitry is first tested in the model plant Arabidopsis thaliana and shall then be used in other plants including crops, which currently offer only limited options for genetic manipulation. For instance, insertion of Tto1 into a crop plant´s genome may generate mutations for gene analysis, and genetic variability for breeding programs.

Andreas Bachmair

TEAMJolanta Ambroz-Kumorowski Katarzyna Hanczaryk Lilian Nehlin Konstantin Tomanov Ionida Ziba

Many proteins are modified after their synthesis. We are interested in how the small modifier proteins ubiquitin and SUMO are attached to substrate proteins in plants, and how these processes change substrate properties.

SELECTED PUBLICATIONSTomanov K, Zeschmann A, Hermkes R, Eifler K, Ziba I, Grieco M, Novatchkova M, Hofmann K, Hesse H, Bachmair A. Arabidopsis PIAL1 and 2 promote SUMO chain formation as E4 type SUMO ligases, and are involved in stress responses and sulfur metabolism. Plant Cell 2014; tpc.114.131300. [Epub ahead of print]. PMID: 25415977 Gibbs DJ, Bacardit J, Bachmair A, Holdsworth MJ. The eukaryotic N-end rule: conser-ved mechanisms and diverse functions. Trends Cell Biol. 2014; 24(10):603-11. PMID: 24874449 Gibbs DJ, Md IN, Movahedi M, Lozano-Juste J, Mendiondo GM, Berckhan S, Marin-de la Rosa N, Conde JV, Sousa Correia C, Pearce SP, Bassel GW, Hamali B, Talloji P, Tomé DF, Coego A, Beynon J, Alabadi D, Bachmair A, León J, Gray JE, Theodoulou FL, Holdsworth MJ. Nitric oxide sensing in plants is mediated by proteolytic control of Group VII ERF transcription factors. Mol. Cell 2014; 53(3): 369-79. PMID: 24462115

Arabidopsis thaliana plants with impaired SUMO conjugation (left pot) are smaller than siblings with intact sumoylation system (right pot, plants supported by wood sticks).





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A N D R E A B A R T A

Post-transcriptional regulation of gene expression in plants

Alternative splicing is one of the posttranscrip-tional events to expand the repertoire of gene prod-ucts and it has been exploited for various differ-entiation processes. In plants, the significance of alternative splicing was long underestimated, but we and others have shown that it greatly impacts on development and responses to the environment. As alternative splicing in Arabidopsis is not well characterized, we are using RNAseq to define the rules and targets of alternative splicing.

SR (Ser/Arg) proteins are important splicing factors and to date we have isolated and partially characterized several Arabidopsis SR proteins, which are important for splice site selection and spliceosome assembly. In addition, we have isolated several regulatory proteins which seem to be essential to drive the splicing process, like SRPK

kinases, helicases and cyclophilins. To elucidate their mechanisms of action some of the plant SR pro-teins and cyclophilins are currently characterized in greater detail in terms of their RNA targets, inter-acting proteins and their impact on flowering and UV-stress response. Interestingly, some of these fac-tors seem to connect splicing to transcription and are therefore currently investigated in greater detail. In addition, the influence of DNA methylation on alternative splicing is evaluated on a genome wide basis. As some of the alternative splicing events are regulated by light by signals from the chloro-plast, screens to establish the signaling pathways are employed. Dynamic RNAseq analysis performed during day and night will allow building a “diurnal map” and establish a network of alternative splicing events impacting the circadian clock.

Andrea Barta

TEAMZahra AyatollahiNicola CavallariArmin FuchsMariya KalynaYamile MarquezManali MishraEzequiel PetrilloNicola Wiskocil

Model of exitron evolution. Exitrons are alternatively spliced internal regions of protein-coding exons. Evolution of exitrons involved loss of introns and retroposition. Vestigial exonic splicing regulatory elements facilitate evolution of core splicing signals and the re-establishment of an AS event.

SELECTED PUBLICATIONSPetrillo E, Godoy Herz MA, Fuchs A, Reifer D, Fuller J, Yanovsky MJ, Simpson C, Brown JW, Barta A, Kalyna M, Kornblihtt AR. A chloroplast retrograde signal regulates nuclear alternative splicing. Science. 2014;344(6182):427-30. PMID: 24763593 Göhring A, Jacak J, Barta A. Imaging of endogenous messenger RNA splice variants in living cells reveals nuclear retention of transcripts inaccessible to nonsense-mediated decay in Arabidopsis. Plant Cell. 2014;26(2):754-64. PMID: 24532591 Marquez Y, Brown JW, Simpson C, Barta A, Kalyna M. Transcriptome survey reveals increased complexity of the alternative splicing landscape in Arabidopsis. Genome Res. 2012;22(6):1184-95. PMID: 22391557

What determines the complexity of higher organisms? No correlation has been found to DNA content and gene number and therefore studies in the field are now focusing on post-transcriptional processes and the impact of the dynamic transcriptome on the complexity of gene expression.

Consequences of exitron splicing: Exitrons are alternatively spliced internal regions of protein-coding exons



http://www.ncbi.nlm.nih.gov/pubmed/?term=Transcriptome+survey+reveals+increased+complexity+of+the+alternative+splicing+landscape+in+Arabidopsis


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D I E T E R B L A A S

Early interactions of viruses with host cells

Most of these pathways converge in vesicular structures, the endosomes. Depending on whether the virus is covered with a lipid membrane (enveloped) or lacking such a membrane (naked), its genome is then being released into the cytoplasm by different mechanisms. Enveloped viruses usually fuse with cellular membranes, which results in the nucleocapsid arriving in the cytosol. Non- enveloped viruses either disrupt the endosomal membrane, with subviral particles being transferred into the cytoplasm, or the genomic nucleic acids are threaded through a pore and the remaining empty capsids are further shuttled to lysosomes for

degradation. For some viruses there is experimental support for RNA transfer between endosomal and cytoplasmic compartments which, however, has so far not been demonstrated explicitly and the pu-tative membrane pore has not been visualized.

Working with human rhino-viruses (HRVs) that lack a lipid membrane and are the predomi-nant cause of common colds, we aim at identifying so far unknown viral receptors, the different

mechanisms underlying viral uptake, the process of genome release and the structural basis of the transfer of the viral genome through lipid membranes. We address these questions by using biochemical, molecular biological, biophysical, and structural biology techniques, such as selection and characterization of viral mutants, expression library screening, fluorescence correlation spectros-copy, capillary electrophoresis, and cryo-electron microscopy.

In the last few years we have identified heparan sulphate as an additional receptor for some rhinovirus types and characterized the uptake path-way by this proteoglycan and by the intercellular adhesion molecule 1, the receptor of about 90 different HRV types. We developed a liposomal in vitro system mimicking the transfer of the viral RNA through the endosomal membrane that is currently being used for structural analysis. Within the frame of several international collaborations, solving the 3-D structure of subviral particles is underway.

Dieter Blaas

TEAMRick ConzemiusIrene GöslerAlexander Otahal

To infect a host cell, viruses usually recognize particular structures on the cell surface. Following binding to these “viral receptors”, the virons are taken up into the cell along different entry routes.

SELECTED PUBLICATIONSGarriga D, Pickl-Herk A, Luque D, Wruss J, Castón JR, Blaas D, Verdaguer N. Insights into minor group rhinovirus uncoating: the X-ray structure of the HRV2 empty capsid. PLoS Pathog. 2012;8(1):e1002473. PMID: 22241997 Fuchs R, Blaas D. Productive entry pathways of human rhinoviruses. Adv Virol. 2012; 2012:826301. PMID: 23227049 Weiss VU, Subirats X, Pickl-Herk A, Bilek G, Winkler W, Kumar M, Allmaier G, Blaas D, Kenndler E. Characterization of rhinovirus subviral A particles via capillary electrophoresis, electron microscopy and gas-phase electrophoretic mobility molecular analysis: Part I. Electrophoresis 2012; 33(12):1833-41. PMID: 22740471

Native and subviral com-mon cold virion particles reconstructed from electron microscopic images similar to the one in the background. Some are cut open to allow a view onto the viral genome that became compacted to a rod-like structure upon crosslinking and induction of uncoating.

Cryo-electron micrograph of virus particles at-tached to liposomal membranes via a derivative of the low-density lipoprotein receptor mimick-ing viral binding to the host cell.





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U D O B L Ä S I

Post-transcriptional regulation in bacteria and archaea

The RNA chaperone Hfq has been recognized as the principle post-transcriptional regulator of catabolite repression in the human pathogen Pseudomonas aeruginosa. Hfq was shown to act as a translational repressor that prevents ribosome loading through binding to A-rich sequences within the ribosome binding site of several mRNAs encod-ing catabolic enzymes. Furthermore, the non-coding RNA CrcZ was shown to bind to and to sequester Hfq, which in turn abrogates Hfq-mediated trans-lational repression. This novel mechanistic twist on Hfq function not only highlighted the central role of RNA based regulation in PAO1 carbon metab-olism but also broadened the view of Hfq-medi-ated post-transcriptional mechanisms. In addition,

CrcZ-mediated regulation of Hfq was shown to im-pact on virulence traits such as biofilm formation and antibiotic susceptibility. The underlying molec-ular events are currently being studied.

Another research focus is directed towards a better understanding of post-transcriptional regulatory mechanisms in the model crenarchaeon Sulfolobus solfataricus (Sso). Previous studies revealed the sequence of events in archaeal transla-tion initiation as well as unprecedented function(s) of archaeal translation initiation factors. Ongoing studies concentrate on the elucidation of the func-tion of archaeal Sm proteins in RNA turnover and on molecular mechanisms underlying non-coding RNA mediated regulation in Sso.

Udo Bläsi

TEAMIsabel Del Pino Gomez Dorothea Heitzinger Diana Claudia Humer Birgit Märtens Diego Oxilia Speratti Petra Pusic Armin Resch Marlena Rozner Wolfgang Sattler Elisabeth Sonnleitner Muralidhar Tata

Increased biofilm formation in a P. aeruginosa crcZ deletion mutant was visualized by confocal scanning laser microscopy. The life/dead stain permits to differentiate between live (green) and dead cells (red). Insets: Biofilm formation in plastic tubes.

SELECTED PUBLICATIONSSonnleitner E, Bläsi U. Regulation of Hfq by the RNA CrcZ in Pseudomonas aeruginosa carbon catabolite repression. PLoS Genet. 2014;10(6):e1004440. PMID : 24945892 Märtens B, Manoharadas S, Hasenöhrl D, Zeichen L, Bläsi U. Back to translation: removal of aIF2 from the 5´-end of mRNAs by translation recovery factor in the crenarchaeon Sulfolobus solfataricus. Nucl. Acids Res.2014;42(4):2505-2511. PMID: 24271401 Märtens B, Manoharadas S, Hasenöhrl D, Manica A, Bläsi U. Antisense regulation by transposon derived RNAs in the hyperthermophilic archaeon Sulfolobus solfataricus. Embo Rep. 2013;14(6):527-33. PMID: 23579342

Prokaryotes in general and microbial human pathogens in particular are constantly challenged by changing environmental conditions. They employ a number of post-transcriptional control mechanisms including proteinaceous translational regulators, small regulatory RNAs (sRNAs) as well as features inherent to mRNA structure, which permit a fast adaptation to new environments such as the host.

X-ray structure of the homo-heptameric Sm2 protein of S. solfataricus. The protomers are color coded. The structure was elucidated in collaboration with the group of K. Djinović-Carugo at the MFPL.

http://www.ncbi.nlm.nih.gov/pubmed/?term=Regulation+of+Hfq+by+the+RNA+CrcZ+in+Pseudomonas+aeruginosa+carbon+catabolite+repression.

http://www.ncbi.nlm.nih.gov/pubmed/?term=Back+to+translation%3A+removal+of+aIF2+from+the+5%C2%B4-end+of+mRNAs+by+translation+recovery+factor+in+the+crenarchaeon+Sulfolobus+solfataricus

http://www.ncbi.nlm.nih.gov/pubmed/?term=Antisense+regulation+by+transposon+derived+RNAs+in+the+hyperthermophilic+archaeon+Sulfolobus+solfataricus


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A L E X A N D E R D A M M E R M A N N

Centriole assembly and function

Centrioles perform two distinct functions in eukaryotic cells: 1) they recruit pericentriolar material to form centrosomes that organize the microtubule cytoskeleton and position the mi-totic spindle, and 2) they template cilia, cellular projections that perform a variety of critical sen-sory and motile functions. Centrosome and cilia abnormalities have been linked to aneuploidy and tumorigenesis as well as developmental disorders including ciliopathies and microcephaly. Despite their importance to human physiology and pathol-ogy, centrioles have remained poorly understood at the molecular level, largely due to the technical challenges posed by the small size of this organelle.

In our lab we are using a combination of bio-chemical, cell biological and genetic approaches in the nematode C. elegans and more recently also the fruit fly Drosophila melanogaster to investi-gate the fundamental and conserved molecular mechanisms underlying centriole assembly and function. In previous work we have taken advan-tage of the availability of data from genome-wide RNAi-based screens to define the molecular

requirements for centriole assembly. The six- protein molecular pathway we identified has since been found to be conserved from ciliates to ver-tebrates, and is thought to form the core of the centriole assembly machinery in all eukaryotes. We further identified the hydrolethalus syndrome protein HYLS-1 as a core centriolar protein that is incorporated into centrioles during their assembly to confer on them the ability to initiate cilia. The single amino acid missense mutation associated with hydrolethalus syndrome impairs HYLS-1 func-tion in ciliogenesis, identifying this disorder as a severe (perinatal lethal) ciliopathy.

Current research builds on this foundation, seeking to answer three main questions: 1) How do centrioles assemble, in particular what are the specific mechanistic contributions of each of the six proteins in the centriole assembly pathway; 2) how do centrioles recruit pericentriolar material to form centrosomes and what is the molecular nature of this material; and 3) how do centrioles form cilia, focusing on the events immediately downstream of HYLS-1.

Alexander Dammermann

TEAMGabriela CabralJeroen DobbelaereBalazs ErdiTriin LaosMadalina MireaMax RoesslerCornelia Rumpf-KienzlDaniel Serwas

Centrioles are small cylindrical organelles whose distinguishing feature is an outer wall composed of a nine-fold symmetric array of stabilized microtubules.

SELECTED PUBLICATIONSCabral G, Sans SS, Cowan CR, Dammermann A. Multiple mechanisms contribute to centriole separation in C. elegans. Curr Biol. 2013;23(14):1380-7. PMID: 23885867 Qiao A, Cabral G, Lettman, Molly M, Dammermann A, Dong G. SAS-6 coiled-coil structure and inter-action with SAS-5 suggest a regulatory mechanism in C. elegans centriole assembly. EMBO J 2012;31(22):4334-47. PMID: 23064147 Dammermann A, Pemble H, Mitchell BJ, McLeod I, Yates JR, Kintner C, Desai AB, Oegema K. The hydrolethalus syndrome protein HYLS-1 links core centriole structure to cilia formation. Genes Dev. 2009;23(17):2046-59.PMID: 19656802

(A) Centriole assembly pathway as delineated in C. elegans. (B) C. elegans early embryo, stained for SAS-4 (centrioles,yellow), γ-tubulin (pericentriolar material, blue), Aurora-A (peripheral pericentriolar material and astral microtubules, red) and microtubules (black). (C) Depletion of HYLS-1 in Xenopus embryo results in failure of cilia assembly (acetylated tubulin, green). Basal bodies (γ-tubulin, blue) are disorganized.





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T H O M A S D E C K E R

Host responses and innate immunity to bacteria

To increase protection cells participating in the innate response initiate an adaptive immune re-sponse. Protection and immunoregulation by the innate immune system requires that a microbe is detected and physical contact is translated into al-tered gene expression of the infected cell.

Antimicrobial gene products provide protective effector mechanisms. Moreover, secreted cytokines fulfill the task of communicating between cells in-volved in the antimicrobial response to maximize the common antimicrobial effort. One important group of cytokines is formed by the interferons (IFN), subdivided into three distinct classes (IFN-I, II, III). Collectively IFN play an indispensable role in the im-mune system. To reprogram gene expression in tar-get cells, IFN employ Jak-Stat signal transduction.

Our research aims at understanding how the syn-thesis of IFN-I is regulated when cells or animals are infected with intracellular bacteria and how they give rise to changes in the cellular transcrip-tome. To this end we infect normal cells and mice and compare them with infected mice that cannot

synthesize IFN-I or that cannot respond to them. In addition, mice with reduced or absent responses to IFN are used to study the impact of the cytokines in a mouse model of acute intestinal inflamma-tion. In this situation we test the hypothesis that IFN contribute to inflammation, thus worsening the outcome. These efforts are coordinated with collab-orators at the University of Vienna that determine corresponding changes in the composition of the intestinal microbiota. More recently we included an analysis of type III interferons (or IFN-lambda) in our analysis.

Infected cells activate numerous signaling pathways to coordinate transcriptional responses to pathogens. A large set of genes requires tran-scriptional cooperativity of the type I IFN and NFκB pathways. Using combined microarray and ChIP-Seq analysis we performed genome-wide analysis of type I IFN/NFκB-regulated genes in macrophages infected with Listeria monocytogenes and deter-mined the mechanisms underlying cooperative control of transcriptional initiation and elongation.

Thomas Decker

TEAMChristophe CapelleDuygu DemirözFabian KallingerAndrea MajorosEkaterini Platanitis Birgit RappFelix RosebrockUrsula StixDaniel SzappanosFotima TouraevaRoland TschismarovSebastian Wienerroither

SELECTED PUBLICATIONSJamieson AM, Pasman L, Yu S, Gamradt P, Homer RJ, Decker T, Medzhitov R. Role of Tissue Protection in Lethal Respiratory Viral-Bacterial Coinfection. Science 2013;340(6137):1230-4. PMID: 23618765 Kernbauer E, Maier V, Stoiber D, Strobl B, Schneckenleithner C, Sexl V, Reichart U, Reizis B, Kalinke U, Jamieson A, Müller M, Decker T. Conditional Stat1 ablation reveals the importance of interferon signa-ling for immunity to Listeria monocytogenes infection. PLoS Pathog. 2012;8(6):e1002763. PMID: 22719255 Farlik M, Reutterer B, Schindler C, Greten F, Vogl C, Müller M, Decker T. Nonconventional initiation complex assembly by STAT and NF-kappaB transcription factors regulates nitric oxide synthase expression. Immunity. 2010;33(1):25-34. PMID: 20637660

The first line of defense is set by the innate immune system which rapidly acts to limit host colonization and the dissemination of microbes.

Gene activation in cells infected with Listeria monocytogenes by cooperation of the type I IFN (blue) and NFκB (red) pathways. The type I interferon receptor (IFNAR) activates Janus kinases (JAKs) to phosphorylate transcription factor ISGF3 whereas pattern recognition receptors (PRR) activate the IκB kinase complex (IKK) and NFκB. ISGF3 contacts the initiation and mediator complexes to recruit RNA polymerase II (pol II). NFκB stimulates promoter binding of histone modifying enzymes and it contacts mediator to recruit both pol II and its activating kinases. Together these events render antimicrobial genes competent for transcriptional initiation and elongation.

http://www.ncbi.nlm.nih.gov/pubmed/?term=Role+of+Tissue+Protection+in+Lethal+Respiratory+Viral-Bacterial+Coinfection

http://www.ncbi.nlm.nih.gov/pubmed/?term=Conditional+Stat1+ablation+reveals+the+importance+of+interferon+signaling+for+immunity+to+Listeria+monocytogenes+infection.



12

K R I S T I N A D J I N O V I Ć - C A R U G O

Structural biology of the cytoskeleton

A striking feature of muscle proteins, and particularly of the specialized striated muscle sarcomere compartment Z-disk, is the high frequency of multiple protein-protein interactions. We aim to generate detailed structural information on the protein-protein interaction network in the Z-disk, from its basic components - alpha-actinin and filamin C – to macromolecular complexes and the adaptor and regulatory Z-disk proteins centred on them.

Several of the components required for the generation of Z-disk complexes are already available in quantities and quality required for structural studies: alpha-actinin, several segments of filamin C, myotilin, FATZ, myopodin and ZASP. We are proceeding with generation of binary and higher complexes and their biochemical, biophysical and structural characterization combining high resolution studies (X-ray diffraction, NMR) with lower resolution approaches that can either yield molecular envelopes (SAXS, SANS, EM) or specific distance information (mass-spectrometry, NMR). These activities are complemented by the development of bioinformatics tools for results and fine tuning of the protein constructs to be structurally analyzed. New bioinformatics

strategies are being designed to extend our prediction capabilities.

With other groups and faculties we are working on structural studies of the RNA chaperone Hfq and its interactions with RNA (Bläsi), yeast magnesium channel Mrs2 (Schweyen), and try-pasonomal cytoskeletal protein MORN1 (Warren). In collaboration with M. Wagner (Faculty for Life Sciences, Univ. Vienna) and C. Obinger (University of Natural Resources and Life Sciences, Vienna) we are studying the family of bacterial chlorite dismutases.

In order to overcome the major bottlenecks in structural and functional studies of proteins, which are availability of milligram amounts of active, chemically and conformationally pure protein and crystallization, an FFG funded Laura Bassi Center for Optimized Structural Studies (COSS) was established. The COSS center is a partnership between the MFPL, the University of Vienna and the Campus Science Support Facilities GmbH as scientific partners on the one side, and the Research Institute for Molecular Pathology (IMP) and BIOMIN Holding GmbH as industrial partners on the other side.

Kristina Djinović-Carugo

TEAMGustavo Arruda BezerraOliviero CarugoAlexej CharnagalovEuripedes De Almeida RibeiroIrina GrishkovskayaAndreas HagmüllerStefan HofbauerAnne-Sophie HummJulius KostanMichael KrutzlerGeorg MlynekAriadna Rodriguez ChamorroSara SajkoEduardo Henrique Salviano BezerraMartina SchneiderClaudia SchreinerAntonio SpongaValeria StefaniaJulia SteinerKarolina Zielinska

We are interested in the molecular mechanisms underlying the actin-based cytoskeleton of the striated muscle.

SELECTED PUBLICATIONSRibeiro Ede A Jr, Pinotsis N, Ghisleni A, Salmazo A, Konarev PV, Kostan J, Sjöblom B, Schreiner C, Polyansky AA, Gkougkoulia EA, Holt MR, Aachmann FL, Zagrović B, Bordignon E, Pirker KF, Svergun DI, Gautel M, Djinović-Carugo K. The Structure and Regulation of Human Muscle α-Actinin. Cell. 2014;159(6):1447-60. PMID: 25433700 Kostan J, Salzer U, Orlova A, Toeroe I, Hodnik V, Senju Y, Zou J, Schreiner C, Steiner J, Meriläinen J, Nikki M, Virtanen I, Carugo O, Rappsilber J, Lappalainen P, Lehto VP, Anderluh G, Egelman EH, Djinovič-Carugo K. Direct Interaction of Actin Filaments with F-BAR Protein Pacsin2. EMBO Rep. 2014;15(11):1154-62. PMID: 25216944 Pavšič M, Gunčar G, Djinović-Carugo K, Lenarčič B. Crystal structure and its bearing towards an understanding of key biological functions of EpCAM. Nat Commun. 2014;5:4764. PMID: 25163760

Model of E. coli Hfq RNA chaperone function.The sRNA displayed by the full-atom model is shown bound to the proximal face of Hfq (green solvent accessible surface). The mRNA is bound on the distal side represented by polyA9 orange flat cartoon representation. The model of a hypothetical mRNA chain is displayed in orange oval cartoon. Hfq acts by restructuring the mRNA, which may be accomplished by the conformationally flexible C-termini. The structural variability of both RNAs in a transient ternary 1:1:1 complex would allow to sample large spaces and Hfq would not only act as a platform for binding and by increasing the local concentration of both ligands, but also serve to promote their flexibility, and consequently successful annealing in a stochastic manner.




13

G A N G D O N G

Structural studies of ciliogenesis

Gang Dong

TEAMJohannes LesigangEkaterina ShimanovskayaKeni Vidilaseris

Eukaryotic cilia and flagella are specialized organelles that are highly conserved from protists to mammals.

SELECTED PUBLICATIONSShimanovskaya E, Viscardi V, Lesigang J, Lettman MM, Qiao R, Svergun DI, Round A, Oegema K, Dong G. Structure of the C. elegans ZYG-1 Cryptic Polo Box Suggests a Conserved Mechanism for Centriolar Docking of Plk4 Kinases. Structure 2014; 22(8):1090-104. PMID: 24980795 Vidilaseris K, Shimanovskaya E, Esson HJ, Morriswood B, Dong G. Assembly Mechanism of Trypanosoma brucei BILBO1, a Multidomain Cytoskeletal Protein. J Biol Chem 2014;289(34):23870-81. PMID: 25031322 Qiao R, Cabral G, Lettman MM, Dammermann A, Dong G. SAS-6 coiled-coil structure and interaction with SAS-5 suggest a regulatory mechanism in C. elegans centriole assembly. EMBO J 2012;31(22):4334-47. PMID: 23064147

As the name ‘flagella’ suggests, these are whip-like cellular appendages that can project either as immotile antennae, or beat vigorously in the extracellular media. There are also many immotile cilia with essential functions in cell signaling. These organelles consist of a membrane-sheathed axoneme, which is an extension of the mother centriole-derived basal body, and hundreds of associated proteins. Cilia and flagella have attracted much attention in recent years because of their roles in the transduction of extracellular signals and their association with an expanding number of human disorders, including respiratory distress syndrome, male sterility, polycystic kidney disease, retinal de-generation, Bardet-Biedl syndrome, and many more, which are collectively called ciliopathies.

Cilia and flagella are assembled and maintained through intraflagellar transport (IFT), following the docking and fusion of the mother centriole to the apical membrane of the cell. IFT is carried out by two distinct protein complexes, IFT-A and B, which transport ciliary cargos within cilia and flagella using the microtubule-associated motor proteins kinesin-II and dynein.

Our laboratory has been focusing on a number of aspects of ciliogenesis, including centriole/basal body duplication, vesicle targeting to the ciliary base, flagellar pocket biogenesis, and intraflagel-lar transport. We are committed to the structural studies of these processes, complemented by func-tional studies carried out with our network of col-laborators. Our principle tool is X-ray crystallogra-phy, but we are also proficient with NMR and EM. Our structural studies are further complemented by site-directed mutagenesis, in vitro biochemical experiments, and in vivo assays to test our mecha-nistic hypotheses. The available new structures will enhance our understanding of how these protein complexes function in the cell and provide hints as to how their malfunction leads to human diseases.

Our long-term goal is to provide an assembly map for centrioles and cilia by systematically character-izing a number of multi-subunit protein complexes that are essential for the biogenesis of these two critical eukaryotic organelles – centrioles and cilia.





14

S I L K E D O R N E R

The regulation of gene expression by small ncRNAs

In the past decade the finding of small non-cod-ing RNAs has entirely revolutionized the way we think about the regulation of gene expression. The major focus of our research is the RNA-mediated gene silencing by siRNAs (small interfering RNAs) and miRNAs (micro RNAs) in Drosophila. miRNAs are small non-coding RNAs that have been well established as key regulators of gene expression, which result in translation repression and/or mRNA destabilization upon binding to their target mRNAs. Key factors for miRNA-mediated mRNA degradation are the components of the miRNA effector complex (AGO1 and GW182) and the gen-eral mRNA degradation machinery (deadenylation and decapping enzymes). An important question we address in our projects is how the mRNA de-

capping machinery gets recruited to mRNAs after targeting by miRNAs. The decapping step in mRNA degradation is of particular interest because it is an irreversible step and decapped mRNAs are exposed to exonucleolytic digestion. Interestingly, we iden-tified an interaction of GW182 and HPat, a general decapping activator. Thus the decapping step is not only a consequence of deadenylation but decap-ping activators get recruited to the miRNA effector complex. Furthermore, the recruitment of HPat to the miRNA effector complex provides a mechanism to commit mRNAs for degradation. Currently, we investigate the timing and order of recruitment of additional mRNA degradation factors to the miRNA effector complex.

Silke Dorner

TEAMSanja AnticStefanie HosinerDaniela Teichmann

Post-transcriptional processes such as mRNA splicing, mRNA degradation, mRNA surveillance, RNA editing, translational repression, and RNA-mediated gene silencing, play crucial roles in the regulation of eukaryotic gene expression.

SELECTED PUBLICATIONSBarisic-Jäger E, Krecioch I, Hosiner S, Antic S, Dorner S. HPat a Decapping Activator Interacting with the miRNA Effector Complex. PLoS One. 2013;8(8):e71860. PMID: 23977167 Jäger E, Dorner S. The decapping activator HPat a novel factor co-purifying with GW182 from Drosophila cells. RNA Biol. 2010;7(3):381-5. PMID: 20458171 Dorner S, Lum L, Kim M, Paro R, Beachy PA, Green R. A genomewide screen for components of the RNAi pathway in Drosophila cultured cells. Proc Natl Acad Sci USA. 2006;103(32):11880-5. PMID: 16882716

We established an inducible expression system for Drosophila cell cultures that allows the measurement of mRNA turnover rates. Left: Northern blot analysis of mRNA levels after a transcriptional pulse. Right: Quantitative analysis of mRNA decay based on the Northern blot experiments shown.


http://www.ncbi.nlm.nih.gov/pubmed/?term=The+decapping+activator+HPat+a+novel+factor+co-purifying+with+GW182+from+Drosophila+cells.



15

R O L A N D F O I S N E R

Lamins in chromatin organization and human disease

We study the dynamics and regulation of the assembly and interactions of lamins and their func-tions during cell differentiation, as well as the effect of disease causing mutations in lamins. In particu-lar, we investigate the specific role of a group of lamin-binding proteins, defined by the presence of a structural motif – the LEM (LAP-Emerin-MAN1) domain – which mediates association of these pro-teins with chromatin. Regulation and functions of nucleoplasmic lamins in health and disease

While most LEM proteins are integral proteins of the inner nuclear membrane, Lamina-Associated Polypeptide 2α (LAP2α) localizes throughout the nucleus. Binding of LAP2α to lamins causes relo-calization of a pool of lamins from the peripheral lamina to the nuclear interior. Knock-down of LAP2α in mice leads to loss of the nucleoplasmic lamin pool (Fig.1) and causes hyperproliferation of early progenitor cells in regenerating tissues. The mo-lecular mechanisms of these functions are poorly understood. We find that nucleoplasmic lamis and LAP2α bind to chromatin and occupy large regions on chromosomes (Fig. 2) thereby contributing to chromatin organization and gene regulation. In the

lamin-linked premature ageing disease Hutchison Gilford Progeria (HGPS) LAP2α and the nucleop-lasmic pool of lamins are lost. We propose that lamin-LAP2α complexes regulate gene expression in adult stem cells during tissue homeostasis and an impairment of these functions in lamin-linked diseases may contribute to the tissue pathologies.Ankle 1, a nuclease potentially involved in DNA damage repair

Ankyrin and LEM domain-containing protein 1 (Ankle1) is a novel LEM domain protein conserved from C. elegans to man. Ankle1 shuttles between cytoplasm and nucleus, and ectopically expressed Ankle1 in the nucleus induces DNA double strand breaks and DNA damage signaling. Ankle1 may be a new component of specific DNA repair pathways.

Roland Foisner

TEAMJuliane BraunAna Catarina CarraoThomas DechatSimona FerraioliKevin GessonJosef GotzmannBernhard MoserDragana MustaficNana NaetarSelma Osmanagic-MyersUrsula PilatMichael Peter SkoruppaMaciej SzafraniecSandra Vidak Elisabeth ZierLivija Zlopasa

Higher order chromatin organization is an important mechanism to regulate gene expression during development and differentiation. The nuclear lamina, a scaffold structure at the nuclear envelope, which consists of lamins and several lamin-binding proteins, is involved in “functional chromatin organization” in metazoan nuclei. Mutations in lamins cause a variety of human diseases including muscular dystrophy, lipodystrophy and accelerated aging syndromes.

SELECTED PUBLICATIONSGesson K, Vidak S, Foisner R. Lamina-associated polypeptide (LAP)2α and nucleoplasmic lamins in adult stem cell regulation and disease. Semin Cell Dev Biol. 2014;29:116-24.PMID: 24374133 Brachner A, Braun J, Ghodgaonkar M, Castor D, Zlopasa L, Ehrlich V, Jiricny J, Gotz-mann J, Knasmüller S, Foisner R. The endonuclease Ankle1 requires its LEM and GIY-YIG motifs for DNA cleavage in vivo. J Cell Sci. 2012;1-10(125):1048-57. PMID: 22399800 Naetar N, Korbei B, Kozlov S, Kerenyi MA, Dorner D, Kral R, Gotic I, Fuchs P, Cohen TV, Bittner R, Stewart CL, Foisner R. Loss of nucleoplasmic LAP2alpha-lamin A complexes causes erythroid and epidermal progenitor hyperproliferation. Nat Cell Biol. 2008;10(11):1341-8. PMID: 18849980

Fig. 1: Immunofluorescence analyses of wild-type (+/+) and LAP2α-deficient (-/-) fibroblasts showing nuclear localization of LAP2α (green) and lamins A/C (red). Bar, 10μm.

Fig. 2: Genome browser view showing LAP2α and lamin A/C chromatin-immu-noprecipitation/deep sequencing profiles on chromosome 1 (ln CHIP/input) and chromatin binding domains obtained using SICER and EDD algorithm.





16

P E T E R F U C H S

Stress response in simple epithelia

There is increasing evidence for the involvement of keratin-associated proteins with the modulation of these functions. One of these proteins is epiplakin, a member of the plakin protein family. Compared to the other protein family members epiplakin has an unusual structure comprising solely 16 (mouse) or 13 (human) plakin repeat domains. Its expression is restricted to epithelial tissues and it binds to intermediate filaments, mainly to keratins, which are the only binding partners identified so far. Epiplakin-deficient mice generated in our laboratory are viable and show no obvious phenotype. These findings are in clear contrast to other proteins belonging to the plakin protein family like plectin, desmoplakin, and BPAG1, which play an important role in mechanically strengthening the skin as shown by phenotypes

of knock-out mice. Subsequent experiments using primary keratinocytes from epiplakin-deficient mice showed that the biological role of epiplakin seems to be different from these plakins and to be con-nected with cellular stress response

rather than with maintenance and regulation of cytoskeletal architecture. This protective function appears to be more prominent in simple epithelial tissues as shown by the knock-down of epiplakin in HeLa cells which led to the dis-ruption of intermediate filament networks, contrasting the situation in keratinocytes.

However, a comprehensive analysis of the in vivo function of epiplakin in simple epithelia using defined animal models is still missing to date. In order to further elucidate the biological function of epiplakin in simple epithelia, we are performing a combination of experiments using mouse injury models and experiments based on cell culture, biochemistry and video microscopy. In the mouse, we use several stress models for simple epithelia in different organ systems which are complemented by experiments with primary cells. In addition we use biochemical and cell culture based methods to investigate epiplakin interaction with simple epithelial keratins in more detail and to reveal epiplakin functions in keratin network recovery after stress.

Peter Fuchs

TEAMSandra SzaboKarl Wögenstein

A major role of the keratin intermediate filaments in simple epithelia is to protect cells from mechanical and non-mechanical stresses.

SELECTED PUBLICATIONSWögenstein KL, Szabo S, Lunova M, Wiche G, Haybaeck J, Strnad P, Boor P, Wagner M, Fuchs P. Epiplakin deficiency aggravates mu-rine caerulein-induced acute pancreatitis and favors the formation of acinar keratin granules. PLoS One. 2014;9(9):e108323. PMID: 25232867 Gross K, Fuchs P, Wiche G. Stress-induced recruitment of epiplakin to keratin networks increases their resistance to hyperphosphorylation-induced disruption. J Cell Sci. 2008;121(Pt 6):825-33. PMID: 18285451 Spazierer D, Fuchs P, Reipert S, Fischer I, Schmuth M, Lassmann H, Wiche G. Epiplakin is dispensable for skin barrier function and for integrity of keratin network cytoarchitecture in simple and stratified epithelia. Mol Cell Biol. 2006;26(2):559-68. PMID: 16382147

Immunolocalization of epiplakin in various mouse tissues. Frozen sections prepared from tissues of adult mice, as indicated, were processed for immunolabeling using anti-epiplakin antibodies.

Co-localization and subcellular co-distribution of epiplakin with keratin aggregates after okadaic acid (OA)-induced filament disruption in wild-type keratinocytes. Primary mouse keratinocytes, treated with OA for 2, 4 and 6 hours were immunolabeled using epiplakin (red) and pan-keratin (green) antibodies.





17

B O R I S G Ö R K E

Signal transduction and post-transcriptional regulation in model bacteria

Our research focuses on the regulatory circuits underlying signal perception and transduction and cellular regulation in Escherichia coli and Bacillus subtilis. Mechanistically, we focus on the roles of small regulatory RNAs (sRNAs) and the functions of protein phosphorylation and protein-protein inter-action for bacterial signal transduction and cellular regulation.

In particular, we are investigating regulation of the cell wall biosynthesis pathway in E. coli, which involves a network of two sRNAs, a novel RNA-binding protein and a two-component system. This network controls synthesis of the GlmS enzyme that catalyzes formation of glucosamine-6-phos-phate (GlcN6P), a key metabolite required for cell wall biosynthesis. Both sRNAs are highly similar. However, only sRNA GlmZ is a direct activator that base-pairs with the glmS mRNA. The sRNA GlmY, however, activates glmS indirectly by protecting GlmZ from degradation. The novel RNA binding pro-tein RapZ binds GlmZ and targets it to cleavage by RNase E. GlmY counteracts this process by acting as specific decoy for RapZ. GlmY accumulates when the concentration of GlcN6P decreases in the cell. As a result, GlmS is adjusted to the level of its enzymatic product, thereby mediating GlcN6P homeostasis in the cell.

Secondly, we investigate two-component sys-tems (TCSs), which are the primary sensory systems of bacteria. A TCS consists of a sensor histidine

kinase, which perceives a dedicated signal in the environment and transduces this information via a response regulator into the cell leading to changes in gene expression or of other cellular activities. We recently identified the multi-protein phospho-relay system PTSNtr as a modulator of the activities of two central histidine kinases, the K+ sensing kinase KdpD and the phosphate sensing kinase PhoR. KdpD and PhoR regulate genes required for uptake of potassium and phosphate sources, respectively. Notably, the non-phosphorylated form of protein IIANtr, the output domain of PTSNtr, is the regulatory agent that stimulates the activities of the sensor kinases through protein-protein interaction. By this mechanism, the PTSNtr controls K+ and phosphate homeostasis.

Boris Görke

TEAMPrajakta BajadSvetlana DuricaYvonne GöpelMuna Ayesha KhanMarkus Mörk-Mörkenstein

In order to survive and propagate, bacteria must be able to sense the environmental changes and to respond to them quickly and adequately.

SELECTED PUBLICATIONSGöpel Y, Khan M, Görke B. Ménage à trois: Post-transcriptional control of the key enzyme for cell envelope synthesis by a base-pairing small RNA, an RNase adaptor protein, and a small RNA mimic. RNA Biol. 2014;11(5):433-42. PMID: 24667238 Göpel Y, Papenfort K, Reichenbach B, Vogel J, Görke B. Targeted decay of a regulatory small RNA by an adaptor protein for RNase E and counteraction by an anti-adaptor RNA. Genes Dev. 2013;27(5):552-64. PMID: 23475961 Lüttmann D, Göpel Y, Görke B. The phosphotransferase protein EIIA(Ntr) modulates the phosphate starvation response through interaction with histidine kinase PhoR in Escherichia coli. Mol Microbiol. 2012;86(1):96-110. PMID: 22812494

Regulation of glmS expression by sRNAs GlmY and GlmZ and the RNase adaptor protein RapZ (Göpel et al., RNA Biol., 2014).





18

A N G E L A H A N C O C K

Molecular basis of adaptive evolution

Our research integrates population genet-ics, bioinformatics, quantitative genetics and controlled experiments (in Arabidopsis) to clarify how species respond to environmental selection pressures.Identifying variation that underlies adaptation to the environment

We conduct population genetic analyses on spa-tially-explicit genomic data sets to understand how adaptation progressed in natural systems. We are particularly interested in clarifying what types of molecular variants underlie adaptation, what mode of selection drove adaptive differentiation and why these factors sometimes differ among species.

Reconstructing evolutionary histories in island Arabidopsis

By comprehensively characterizing the evolu-tionary process in particular cases, we can uncover general principles of adaptation and evolutionary change. However, in complex organisms from nat-ural populations, this is a daunting task because it requires knowledge of the natural environment, the important adaptive traits, the genetic basis of phenotypic variation, and evidence that differences in this genetic basis equate to fitness differentials in the natural population. Ecologically interesting populations of well-studied model organisms can provide the background knowledge and tools nec-essary to overcome this challenge.

Islands represent powerful systems for unrav-eling evolutionary histories because in these sys-tems complexity is reduced relative to mainland populations and natural processes can be studied in relative isolation. To this end, we are using pop-ulations of the model plant Arabidopsis thaliana from Macaronesian archipelagos to dissect pheno-typic variation and reconstruct adaptive histories. Ongoing and planned projects focus on identifying functional genetic variants, modeling their evolu-tionary histories and testing for fitness differen-tials in simulated and natural environments using a combination of population genetic analysis, trait mapping, genome editing and field work.

Angela Hancock

TEAMMia Goessinger

Adaptation to different local environments can result in large-scale phenotypic diversity across a species’ range. Determining how this variation is produced and maintained is a central goal of evolutionary biology.

SELECTED PUBLICATIONSHorton MW, Hancock AM, Huang YS, Toomajian C, Atwell A, Auton A, Muliyati NW, Platt A, Sperone FG, Wilhjálmsson BJ, Nordborg M, Borevitz JO, Bergelson J. Genome-wide patterns of genetic variation in worldwide Arabidopsis thaliana accessions from the RegMap panel. Nat. Genet. 2012;44:212-216. PMID: 22231484 Hancock AM, Brachi B, Faure N, Horton MW, Jarymowycz LB, F. Sperone G, Toomajian C, Roux F, Joy Bergelson J. Adaptation to Climate Across the Arabidopsis thaliana Genome. Science 2011;334(6052):83-86. PMID: 21980108 Hancock AM, Witonsky DB, Ehler E, Alkorta-Aranburu G, Beall C, Gebremedhin A, Sukernik R, Utermann G, Pritchard J, Coop G, Di Rienzo A. Colloquium paper: human adaptations to diet, subsistence, and ecoregion are due to subtle shifts in allele frequency. Proc. Natl. Acad. Sci. USA 2010;107 Suppl 2:8924-30. PMID: 20445095

Arabidopsis growing in Cape Verde Islands.

http://www.ncbi.nlm.nih.gov/pubmed/?term=Genome-wide+patterns+of+genetic+variation+in+worldwide+Arabidopsis+thaliana+accessions+from+the+RegMap+panel.


http://www.ncbi.nlm.nih.gov/pubmed/?term=Colloquium+paper%3A+human+adaptations+to+diet%2C+subsistence%2C+and+ecoregion+are+due+to+subtle+shifts+in+allele+frequency


19

A N D R E A S H A R T I G

Origin and biogenesis of peroxisomes

Peroxisomes are highly versatile organelles and essential for life. They participate in many metabolic processes, most notably the degradation of fatty acids and the glyoxylate cycle. Synthesis of organelles and their degradation has to be tightly regulated in agreement with the metabolic status of the cell. Accordingly, peroxisomes need to be maintained in sufficient number to ensure metabolic homeostasis. A network of interacting proteins guar-antees the biogenesis of functional peroxisomes, the transport of peroxisomal matrix proteins across the organellar membrane, and the control of size, shape and number of these compartments. Dispens-able peroxisomes are degraded in a process called pexophagy. Employing yeast as model system we aim to elucidate the molecular mechanisms leading to new peroxisomes either through proliferation of already existing ones or via a de novo biogenesis pathway through fission from the ER.

Currently, our main interest is focused on the mechanism of the de novo biogenesis initiated at the ER. Proteins exclusively involved in the biogene-

sis of peroxisomes are called peroxins (Pex-proteins). Among these the Pex11 protein is a membrane elongation factor, and in yeast, we showed that this protein acts only on already existing peroxisomes leading to proliferation. Two distantly related yeast proteins, Pex25p and Pex27p, play similar roles at the peroxisomal membrane and, in addition, participate in the de novo biogenesis. Together with Pex3p the Pex25 protein is essential for the initiation of peroxisome generation at the ER. Distinct vesicles emanating from the ER may slowly mature into peroxisomes or may fuse with each other or already existing peroxisomes to form mature organelles. The priming event at the ER, the proteins involved and the molecular mechanism are so far unknown, and will be the focus of our future work.

Andreas Hartig

Eukaryotic cells contain organelles separating metabolic pathways. This spatial separation ensures optimal flux of metabolic intermediates and increases the efficiency of the metabolism.

SELECTED PUBLICATIONSWeber C, Hartig A, Hartmann RK, Rossmanith W. Playing RNase P evolution: swapping the RNA catalyst for a protein reveals functional uniformity of highly divergent enzyme forms. PLoS Genet. 2014;10(8):e1004506. PMID: 25101763 Kunze M, Hartig A. Permeability of the peroxisomal membrane: lessons from the glyoxylate cycle. Front Physiol.2013;4(204). PMID: 23966945 Huber A, Koch J, Kragler F, Brocard C, Hartig A. A Subtle Interplay between Three Pex11 Proteins Shapes de novo Formation and Fission of Peroxisomes. Traffic 2012;13 (1):157-167. PMID: 21951626

Peroxisomes are either transported to the newly born bud or are generated de novo from the ER. Yeast cells carrying a mutation in the inheritance process depend exclusively on the de novo biogenesis.

Wild type yeast cells accumulate a fluorescent peroxisomal protein in peroxisomes.





20

M A R C E L A H E R M A N N

LDL-R gene family, apolipoproteins and lipid transfer

In this context, the roles of the LDL receptor gene family members, apolipoproteins and lipid transfer proteins are studied. The developing avian embryo constitutes an excellent system for these studies. During oocyte growth in the chicken, the yolk accumulates via uptake from the circulation of precursor proteins and serves as the sole source of lipid, carbohydrate, and protein. Only 350 mg of the 5-6 g of lipid in the yolk are mobilized during the first two weeks of embryogenesis, the major portion is transported during the final week. Such uptake, to a large part, occurs via the yolk sac, which utilizes the yolk lipoprotein components,

following their degradation or modification, for re-synthesis of lipoproteins which are subsequently secreted and delivered to the embryo through the embryonic circulatory system.

The yolk-sac derived lipoproteins, mainly VLDL, contain much higher proportions of cholesteryl esters than yolk VLDL and harbor the intact form of apoB-100 rather than proteolytic fragments thereof. Furthermore, they lack apoVLDL-II, which is synthesized by laying hens and is present in yolk VLDL. These findings suggest that processing of yolk components inside the yolk sac proceeds in controlled fashion, initially involving degradation of their constituents.

We also focus on the role of LDL modification in atherogenesis. There is now some evidence that the modification of LDL may play a key role in early atherogenic events. Modifed LDL activates endothelial cells to attract and bind monocytes. Consecutively foam cells are formed, leading to the appearance of the fatty streak lesion. Various dis-eases such as diabetes, chronic renal insufficiency and obesity come along with elevated levels of blood cholesterol and different modified LDL. We are interested in identifying compounds (synthetic, natural) with the potential to act as catalysts or inhibitors of the atherogenic modification of LDL.

Marcela Hermann

TEAMMax StraussBarbara WalderFlorian MittermayerMiriam Kamper

Our studies focus on the biology of the growing chicken oocyte and the developing chicken embryo. Specifically, we are interested in unraveling molecular mechanisms involved in the transport of VLDL from the egg yolk to the embryo proper.

SELECTED PUBLICATIONSEresheim C, Plieschnig J, Ivessa NE, Schneider WJ, Hermann M. Expression of microsomal triglyceride transfer protein in lipoprotein-synthesizing tissues of the developing chicken embryo. Biochimie 2014;101:67-74. PMID: 24394625 Nikolay B, Plieschnig JA, Šubik D, Schneider JD, Schneider WJ, Hermann M. A novel estrogen-regulated avian apolipoprotein. Biochimie 2013;95:2445-2453. PMID: 24047540 Praschberger M, Hermann M, Wanner J, Jirovetz L, Exner M, Kapiotis S, Gmeiner BM, Laggner H. The uremic toxin indoxyl sulfate acts as a pro- or antioxidant on LDL oxidation. Free Radic Res 2014;48(6):641-648. PMID: 24568219

During oogenesis in the chicken, the yolk precursors (e.g., vitellogenin and VLDL) are synthesized by the maternal liver under stringent hormonal control (E2) and taken up into the oocyte via receptor-mediated endocytosis (LRs). After ovulation and fertilization, a major feature of development is the formation of a series of extraembryonic structures including the amnion, chorion, allantois and yolk sac membranes (modified from http://chickscope. beckman.uiuc.edu/). Inset: The yolk sac is a layer of tissue growing over the surface of the yolk containing area vasculosa with blood vessels (bv), endothelial cells (EC), and an inner single layer of endodermal epithelial cells (EEC) with endocytic LRs and basement membrane (bm). A major role of the yolk sac is the uptake of nutrients from the yolk, their deg-radation and/ or modification for resynthesis and secretion into the embryonic circulation.





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J O A C H I M H E R M I S S O N

Theoretical population genetics

Our aim is to design advanced mathemati-cal methods and models that account for the biological complexity involved in most evolutionary processes. Complexity arises on all levels of bio-logical organization: molecular, organismal, and ecological. The key issues of evolutionary research are usually addressed in special sub-disciplines, i.e. molecular population genetics, quantitative genetics, and evolutionary ecology. We work on all three fields with the special goal to create an integrative approach, using a combination of different models, concepts, and methods. Methods include analytical work, extensive computer simulations and statistical data analysis. Molecular approaches

The availability of DNA polymorphism data on a genome-wide scale (“population genomics”) is arguably the most significant development in evolutionary research today. In this context, the characterization of the adaptive process on the level of the molecular genotype is a primary research focus in our group. Our aim is to extend the popu-lation genetic theory of molecular adaptation to a broader range of biological scenarios. Quantities of interest are fixation probabilities and fixation times

and the expected footprint of selection on linked neutral variation. Phenotypic approaches

It is widely appreciated that the genetic basis of most quantitative traits consists of complex gene networks. However, when and how gene interac-tions (epistasis) affect evolutionary processes is far less clear. We have studied the evolutionary role of epistasis in equilibrium and non-equilibrium systems. A special research focus is on the effects on genetic variation and the adaptive process and on the evolution of the genotype-phenotype map. Ecologically motivated approaches

The vast majority of population genetic models work under the assumption of a constant fitness landscape. However, natural fitness landscapes will change over space and time. And because an im-portant aspect of an individual’s environment is the composition of phenotypes in its own population, fitness will also depend on allele frequencies. The aim of this third line of our research is to com-bine genetic models with ecological factors. Recent studies have focused on conditions for speciation in spatially structured populations with gene-flow.

Joachim Hermisson

TEAMAndrea FulgioneChristian Huber

MaBS members at Mathematics Department:Alexandre BlanckaertIlse HöllingerSebastian MatuszewskiAgnes RettelbachDerek Setter

The work of the Mathematics and Biosciences Group (MaBS) is on theoretical population genetics and evolutionary ecology.

SELECTED PUBLICATIONSUecker H, Otto SP, Hermisson J. Evolutionary rescue in structured populations. Am Nat. 2014; 183(1):E17-E35. PMID: 24334746 Bank C, Bürger R, Hermisson J. The limits to parapatric speciation: Dobzhansky-Muller incompatibilities in a continent-island model. Genetics. 2012;191(3):845-63. PMID: 22542972 Hancock AM, Brachi B, Faure N, Horton MW, Jarymowycz LB, Sperone FG, Toomajian C, Roux F, Bergelson J. Adaptation to climate across the Arabidopsis thaliana genome. Science. 2011;334(6052):83-6. PMID: 21980108

Evolutionary Rescue: When a population experiences a severe environmental challenge it may face extinction unless it is able rescue itself by rapid adaptation. In Uecker et al. (2014), rescue probabilities are derived for complex ecological scenarios. An important finding is that more severe changes in the environment can sometimes lead to higher rescue probabilities if rescue mutations already exist (at low frequency) prior to the environmental change. The figure shows the rescue probability Presc as a function of the severity r of the environmental change. Full line: analytical prediction for Presc. Dashed: rescue from pre-existing variation, dotted: rescue from new mutations only, symbols: computer simulations.





22

R E I N H O L D H O F B A U E R

Consequences of carnitine deficiency and CSF-1 inhibition

Our group revealed that L-carnitine acts as a nutrigenomical metabolite upon gene expres-sion. The clinical condition of carnitine deficiency defines a very critical condition, we therefore use it in a well defined cell culture model system to investigate the effects of carnitine supplementation in human liver, fibroblast, endothelial, nerve and muscle cells. By chip screen and promoter stud-ies we established a solid basis to study changes on mRNA expression levels and promoter factors. We identified genes being directly involved in the transcriptional regulation of the “L-carnitine effect”, thus being able to approach clinical pathologies of hyperlipidemia, insulin resistance and type 2 diabetes mellitus. We want to reveal

“candidate or susceptibility” genes, which are associated with these diseases and have an increased sensitivity to diet. The results of this research will provide better insight in metabolic aspects of pathologies and their regulation as well as mitochondrial function.

In a separate effort we are tracing the effects associated with inhibition of CSF-1. It primarily acts on cells of the mononuclear phagocyte lin-eage by controlling the differentiation, prolifera-tion and survival of precursor cells as well as the activation of mature macrophages. CSF-1 also has a pivotal role in the pathogenesis of several disorders including cancers, as it regulates the production genes involved in tissue remodeling and tumor invasion. We investigated whether inhibition of CSF-1 expression can serve as a valuable tool to fight tumor growth and metastasis. Microarray analyses have revealed very promising candi-date genes that are re- or induced during CSF-1 inhibition. Their inhibition should enhance the inhibitory effect of CSF-1 specific antibodies or specific RNAi. Preclinical animal studies with monoclonal antibodies/RNAis are the next experimental aims.

Recently we characterized potentially cy-to-toxic genes (e.g. Tmem66, superactive thymi-dine kinase 1) identified or generated in my group. These genes were cloned into an inducible vector system (pUHD-Hygr) and then transfected into model tumor cells (MCF-7 pretransfected with a puromycin resistance gene carrying a silencer construct). The final aim is to develop a genetic approach to attack tumor cells in mammalian organisms.

Reinhold Hofbauer

TEAMKlemens KienesbergerEva SteinerClemens JobstAlexander PanzenböckStefan Sack

Signaling processes are very important for the transcriptional activation of genes taking place under carnitine deprivation and CSF-1 inhibition.

SELECTED PUBLICATIONSKienesberger K, Pordes AG, Völk TG, Hofbauer R. L-carnitine and PPARα-agonist fenofibrate are involved in the regulation of Carnitine Acetyltransferase (CrAT) mRNA levels in murine liver cells. BMC Genomics 2014;15:514-524. PMID: 24962334 Blake SM, Strasser V, Andrade N, Duit S, Hofbauer R, Schneider WJ, Nimpf J. Thrombospondin-1 binds to ApoER2 and VLDL receptor and functions in post-natal neuronal migration. EMBO J. 2008;27(22):3069-80. PMID: 18946489 Godárová A, Litzlbauer E, Brunner S, Agu AC, Lohninger A, Hofbauer R. L-Carnitine regulates mRNA expression levels of the carnitine acyltransferases CPT I, CPT II and CRAT. Chem. Monthly 2005;136:1349-1363.

The pivotal role of L-carnitine for the mitochondrial lipid metabolism.




23

N . - E R W I N I V E S S A

Protein biogenesis and degradation from the ER

Misfolded polypeptides are retro-translocated from the ER to the cytosol, where they become poly-ubiquitinated and destructed by proteasomes. ER-associated degradation (ERAD) is of relevance for a variety of genetically inherited, neurodegenerative, and virally transmitted diseases with protein folding defects.

We have previously shown that a truncated form of ribophorin I, a model glycoprotein for ERAD, is degraded by the ubiquitin/proteasome system. The role of N-linked glycans in ERAD was pinpointed as temporary retention devices in the ER. Thus, interaction of N-glycosylated substrates with the calnexin cycle appears to prolong their half lives. Furthermore, the requirement of N-linked glycan trimming for ERAD was shown, and from studies with mutant cell lines with defects in N-glycan assembly the activities of one or more ER α1,2- mannosidases could be implicated in ERAD.

Interaction partners of ERAD substrate proteins in these mutant cell lines will be identified and further characterized.

Another aspect of this project deals with the precise intracellular localization of the ERAD pathway of glycoproteins by indirect immunofluorescence and confocal laser scanning microscopy using appropriate marker proteins.The role of MTP and PDI in the assembly and secre-tion of atherogenic lipoprotein particles

Microsomal triglyceride transfer protein (MTP) is a lipid transfer protein required for the assembly and secretion of very low density lipoproteins (VLDL). Ac-tive MTP is a heterodimer containing a 58 kDa sub-unit identified as protein disulfide isomerase (PDI). The MTP complex catalyzes the loading of apolipo-protein B (apoB) with lipids and/or the translocation of apoB into the lumen of the ER. We are studying the effect of estrogen treatment on MTP activity and on the regulation of VLDL secretion that is also de-termined by lipid availability and apoB degradation. In this context, the consequence of altered intracellular MTP activity on VLDL assembly and secretion is being analyzed. Another aspect of the project is concerned with the mechanism of retention of the MTP complex in the ER.

N.-Erwin Ivessa

TEAMKitty KollMichael Paal

We are interested in the molecular characterization of a quality control system that operates in the endoplasmic reticulum (ER) to ensure that only properly folded proteins will be released.

SELECTED PUBLICATIONSIvessa NE, Rehberg E, Kienzle B Seif F, Hermann R, Hermann M, Schneider WJ, Gordon DA. Molecular cloning, expression, and hormonal regulation of the chicken microsomal triglyceride transfer protein. Gene. 2013;523(1):1-9. PMID: 23542778 Kitzmüller C, Caprini A, Moore SE, Frénoy JP, Schwaiger E, Kellermann O, Ivessa NE, Ermonval M. Processing of N-linked glycans during endoplasmic-reticulum-associated degradation of a short-lived variant of ribophorin I. Biochem J. 2003;376(Pt 3):687-96. PMID: 12952521 Hermann M, Foisner R, Schneider WJ, Ivessa NE. Regulation by estrogen of synthesis and secretion of apolipoprotein A-I in the chicken hepatoma cell line, LMH-2A. Biochim Biophys Acta. 2003;1641(1):25-33. PMID: 12788226

The ERAD substrate RI332-6HA (B, D) co-localizes with the ER marker calnexin (A, C), but hardly with the marker for the ER-Golgi-intermediate-compartment, ERGIC53 (E, F) in Chinese hamster fibroblast cells.





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M I C H A E L F . J A N T S C H

Transcriptome diversification through RNA editing

One mechanism to achieve this, is RNA editing by adenosine deaminases that act on RNA (ADARs). ADARs convert adenosines to inosines in structured and double-stranded RNAs.

As inosines are interpreted as guanosines by most cellular processes, this type of editing can affect the coding potential of an RNA, its folding, stablility, or localization. ADAR mediated editing is widespread in metazoa and affects thousands of transcripts in the human transcriptome. Together with alternative splicing, RNA-editing therefore leads to massive diversification of the proteome. This is exemplified best by the fact that both RNA-editing and alternative splicing are most abundant in the mammalian brain. Consistent with its important function in modulating the tran-scriptome, RNA editing is essential for normal life and development in many organisms. Our research is focused on topics related to this type of RNA editing and aims at understanding the biochem-ical, cellular, and organismic consequences of A to I conversion, as well as its interplay with other post-transcriptional RNA-processing events. Editing in protein coding mRNAs

A handful of highly conserved protein coding targets for A to I editing are known today. To un-derstand the impact of editing on these RNAs and their encoded proteins we are generating trans-genic mice that are impaired in specific editing events. Our studies show that lack of editing of the mRNAs encoding the actin crosslinking pro-tein filamin A leads to altered smooth muscle cell contraction, cell motility and has a dramatic impact on cellular physiology.

Repetitive elements as modulators of gene expression

Massive editing can be found in structured 3’ ends of mRNAs. The editing sites are localized in basepaired regions formed between inverted repetitive elements. Analysis of these untranslated regions in reporter gene assays demonstrates that basepaired elements dramatically reduce gene expression. This phenomenon is independent of RNA-editing but is triggered by the double stranded structure formed in the 3’ UTR. Understanding the mechanisms by which these 3’ ends control gene expression is another goal of our research.

Michael F. Jantsch

TEAMCeline BrunnerLaura CimattiEmilio CusanelliEric HöhenwarterMamta JainKonstantin LichtMaja StulicMansoureh Tajaddod

The number of genes found in different organismic groups does not reflect their biological complexity. Instead, transcriptome diversification can increase biological complexity.

SELECTED PUBLICATIONSBarraud P, Banerjee S, Mohamed WI, Jantsch MF, Allain FH. A bimodular nuclear localization signal assembled via an extended double-stranded RNA-binding domain acts as an RNA-sensing signal for transportin 1. Proc Natl Acad Sci U S A. 2014;111(18):E1852-1861. PMID: 24753571 Vesely C, Tauber S, Sedlazeck FJ, Tajaddod M, Haeseler AV, Jantsch MF. ADAR2 induces reproducible changes in se-quence and abundance of mature microRNAs in the mouse brain. Nucleic Acids Res. 2014;42(19):12155-68. PMID: 25260591 Vesely C, Tauber S, Sedlazeck FJ, Haeseler AV, Jantsch MF. Adenosine deaminases that act on RNA induce reproducible changes in abundance and sequence of embryonic miRNAs. Genome Res. 2012;22(8):1468-76. PMID: 22310477

Editing of repetitive elements in mRNAs. Repetitive elements (such as Alu elements) can basepair if inserted in opposite orientation. The basepaired regions are recognized by ADAR enzymes and can thus provide a substrate for RNA editing.





25

V E R E N A J A N T S C H

Meiosis in Caenorhabditis elegans

Research in my lab is directed towards the identi-fication of genes and processes essential in meiotic prophase in an animal model system. Excellent for-ward and reverse genetics, transparency and easy cytological observation of all meiotic stages make the nematode Caenorhabditis elegans a powerful system for our studies.

During the first meiotic division, faithful chromo-some segregation is facilitated by the formation of a physical tether between the parental homologs (me-diated by crossover recombination and cohesion). Crossovers require the introduction of DNA dou-ble-strand breaks, chromosome pairing, formation of the synaptonemal complex, and double-strand break repair by homologous recombination using the homolog as a repair template.

In meiotic prophase I chromosomes are moved by cytoplasmic forces transferred to the nucleus via the SUN/KASH protein module (components of the outer and inner nuclear envelope that connect chromo-somes to cytoplasmatic microtubules). Abrogation of chromosome movement, as we demonstrated with the sun-1(jf18) allele, leads to precocious syn-apsis involving non-homologous chromosomes. We study the nature of chromosome movement and its regulation. These forces stir chromosomes, helping to bring homologs together and to inhibit unde-sired interactions. In addition we discovered that SUN-1 is an integral part of a meiotic surveillance mechanism that coordinates chromosome synapsis

and recombination with meiotic progression and chromosome movement. This checkpoint system monitors establishment of the obligate crossover, inducible only in leptotene/zygotene. Unrepaired DSBs and unsynapsed chromosomes maintain this checkpoint, but a crossover intermediate is neces-sary to shut it down.

Finally, we study a component of a putative double holiday junction dissolvase complex. During meiosis programmed double strand break induction leads to formation of about 12 breaks per chromo-some in C. elegans. In worms strictly only one of these matures into a crossover product. The rest of these gets resolved or dissolved in a non-crossover manner. To understand the interplay between these pathways will be essential to understand how a balanced number of crossovers will be established, since too few crossovers or unrepaired DNA double strand breaks are detrimental to gametes.

Verena Jantsch

TEAMAnahita DaryabeigiMaria Rosaria Dello StrittoMarlene JagutJana LinkMartin MiklSophia MilloniggDimitra PaouneskouJudith Windisch

Meiosis is the specialized cell division that generates haploid germ cells. It not only halves the chromosome content but also ensures genetic diversity by recombination. Errors in meiosis lead to infertility, pregnancy loss and clinical syndromes linked to mental retardation.

SELECTED PUBLICATIONSWoglar A, Daryabeigi A, Adamo A, Habacher C, Machacek T, La Volpe A, Jantsch V. Matefin/SUN-1 Phosphorylation Is Part of a Surveillance Mechanism to Coordinate Chromosome Synapsis and Recombination with Meiotic Progression and Chromosome Movement. PLoS Genet. 2013;9(3):e1003335. PMID: 23505384 Baudrimont A, Penkner A, Woglar A, Machacek T, Wegrostek C, Gloggnitzer J, Fridkin A, Klein F, Gruenbaum Y, Pasierbek P, Jantsch V. Leptotene/zygotene chromosome movement via the SUN/KASH protein bridge in Caenorhabditis elegans. PLoS Genet. 2010;6(11):e1001219. PMID: 21124819 Penkner AM, Fridkin A, Gloggnitzer J, Baudrimont A, Machacek T, Woglar A, Csaszar E, Pasierbek P, Ammerer G, Gruenbaum Y, Jantsch V. Meiotic chromosome homology search involves modifications of the nuclear envelope protein Matefin/SUN-1. Cell 2009;139(5):920-33. PMID: 19913286

In early C. elegans meiosis one end of each chromosome attaches to the nuclear envelope via meio-sis-specific protein complexes (filled blue, yellow and orange circles). Cytoplasmic tubulin (pink bars) provide the driving forces that move chromosomes (blue and brown lines) vigorously along the surface of the inner nuclear envelope. Cytoplasmic driving forces are transmitted to the nucleus via SUN-KASH protein com-plexes (green and magenta ellipses). Concomitantly the synaptonemal complex forms between homologous chromosomes (pink ladder like lines).





26

F R A N Z K L E I N

Chromosome structure and meiotic recombination

During meiosis, the genetic content of a dip-loid cell is reduced to half, a prerequisite for the production of gametes and for sexual reproduction. Our experience shows that the important processes are conserved between yeast and man, so that many of our findings can be generalized.

Chromosomes are organized as dynamic structures with distinct micro-domains, such as axis and loop regions, as well as macro-domains, such as recombination rich – or poor regions, centromeres, telomeres and others. In meiosis, cohesins and axial element proteins shape the chromosome and

mediate recombination as well as correct chro-mosome segregation. We have established a high- resolution map of protein-chromosome interactions by microarray-analysis, which we currently improve using deep sequencing. We also study in detail the process of chromosome synapsis, a structure that mainly exists to fine-tune recom-bination pathways. Eventually, these pathways decide over the integrity of the resulting gamete’s genome, and thus over the health of the new individual that originates from these gametes.

Franz Klein

TEAMCeline BrunnerLaura CimattiEmilio CusanelliEric HöhenwarterMamta JainKonstantin LichtMaja StulicMansoureh Tajaddod

We study meiotic recombination and chromosome segregation in S. cerevisiae as model organism to understand the interplay between chromosome structure and recombination.

SELECTED PUBLICATIONSXaver M, Huang L, Chen D, Klein F. Smc5/6-Mms21 prevents and eliminates inappropriate recombination intermediates in meiosis. PLoS Genet. 2013;9(12):e1004067. PMID: 24385936 Klug H, Xaver M, Chaugule VK, Koidl S, Mittler G, Klein F, Pichler A. Ubc9 sumoylation controls SUMO chain formation and meiotic synapsis in Saccharomyces cerevisiae. Mol Cell. 2013;50(5):625-36. PMID: 23644018 Panizza S, Mendoza MA, Berlinger M, Huang L, Nicolas A, Shirahige K, Klein F. Spo11-accessory proteins link double-strand break sites to the chromosome axis in early meiotic recombination. Cell. 2011;146(3):372-83. PMID: 21816273

The machine for DSB formation (three coloredballs symbolize three located components) waslocalized to the base of DNA loops (Panizza et al., Cell, 2011). In contrast, DSBs are made on chromosome loops (Blatt et al., Cell, 2002).

Glimpse into the nano-world – section of a chromosome with “DNA-break machines“. Loops of sister chromatids (blue and turquoise) are linked and held in shape by ring-molecules. The machines are anchored between the DNA-loops at the axis of the chromosome. The model is illustrative, but postulates unknown details, such as the way the ring-molecules hold DNA-loops.





27

A L W I N K Ö H L E R

Gene expression and chromosome dynamics

We focus on two areas: First, we explore the role of nuclear pore complexes (NPCs) in genome regulation. Interphase chromosomes are not ran-domly spread throughout the nucleus but are fairly well organized, with different gene loci found in different regions of the nucleus. At the same time, chromatin can undergo extensive motion. In fact, some inducible genes dramatically change nuclear positions depending on whether they are active or not. A fascinating new line of research suggests that activated genes can become hooked to nuclear pores – large transport channels, which protrude into the nuclear interior with a basket-like structure. According to this view, NPCs serve as anchors for the gene expression machineries and play a role in tuning gene activities. We would like to understand which factors mediate chromatin-NPC interactions, how these links are formed and broken and how they contribute mechanistically to transcription, RNA processing and export. Ultimately, our goal is to unravel basic principles of how nuclear architecture determines cellular function.

Secondly, we investigate how ubiquitin signaling controls gene expression. While ubiquitin is well-known for tagging proteins for destruction by the

proteasome, its role in regulating chromatin is far less understood. We are particularly interested in the enzymatic toolkit for histone ubiquitination (ligases & deubiquitinases). When appended to histones ubiquitin can function as a reversible molecular switch to regulate transcription, gene silencing and DNA repair. Recently, we have determined the structure of a histone deubiquitinase together with our collaborators and uncovered its sophisticated activation mechanism. Intriguingly, the Ubp8 deubiquitinase forms a pro-tein module with three co-factors, which act in concert to assemble the module, shape the catalytic center and recognize the substrate. The deubiquitinase module is part of SAGA, a multifunctional transcription co-activator. Our studies serve as a paradigm to explain how a deubiquitinase is switched on at the right time and place inside the cell. In addition, we aim to discover novel ubiquitin functions related to RNA and chromatin biology.

Alwin Köhler

TEAMLaura Gallego ValleMaria Jose MendiburoNoemi MeszarosMaren SchneiderTobias SchubertEleonora TurcoAnete RomanauskaXiao Yin LeeJakub Cibulka

My group is broadly interested in genome organization and the mechanisms of gene expression.

SELECTED PUBLICATIONSKöhler A, Zimmermann E, Schneider M, Hurt E, Zheng N. Structural basis for assembly and activation of the heterotetrameric SAGA histone H2B deubiquitinase module. Cell. 2010; 141(4):606-17. PMID: 20434206 Köhler A, Hurt E. Gene regulation by nucleoporins and links to cancer. Mol Cell. 2010;38(1):6-15. PMID: 20385085 Köhler A, Schneider M, Cabal GG, Nehrbass U, Hurt E. Yeast Ataxin-7 links histone deubiquitination with gene gating and mRNA export. Nat Cell Biol. 2008; 10(6):707-15. PMID: 18488019

Multi-step activation and structure of a Ubiquitin Pac-Man (Köhler et al., Cell, 2010).

NPCs and proteins of the inner nuclear membrane partition the genome into areas of silent (yellow) and active chromatin (green). Gene-NPC interactions require various adaptors including the SAGA histone acetyltransferase (HAT) (Köhler & Hurt, Mol Cell, 2010).





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G O T T F R I E D K Ö H L E R

Biomolecular optical spectroscopy

Methods include fluorescence and time resolved techniques performed over a wide range of time resolution. Studies by optical spectroscopy are complemented by biocalorimetry (DSC).

Quantitative studies on molecular dynamics on a single molecule level are performed using advanced fluorescence correlation techniques. Among others, these methods are applied on studies of ligand- receptor interactions relevant for hormone regu-lation and of the mechanisms of endocytosis and transport in single living cells. These measurements provide the basis for mathematical modelling of complex dynamic behaviour in biosystems, im-ple-mented in close cooperation with other re-search groups.

Translational research related to clinical diagnostics is another area of focus. Our aim is to establish and to optimize novel and highly sensitive detection tools for inflammatory biomarkers such as interleukins in body fluids (serum, saliva and tears) using fluorescence correlation techniques. It will help to develop cost effective, highly sen-sitive and portable diagnostic/screening devices that can be incorporated into clinics. Other project deals with the detection of malignant cells in body fluids such as urine by fluorescence imaging. This technology will also allow detecting circulating malignant cells (CTC) in plasma in case of cancer metastasis.

Gottfried Köhler

TEAMErwin GaubitzerMartin Gerald PuchingerAamir ShahzadDavid Weichselbaum

Biophysical characterization of biomolecules and of their interactions in solution as well as on a live cell level represents the main object of our research.

SELECTED PUBLICATIONSEdetsberger M, Knapp M, Gaubitzer E, Miksch C, Gvichiya KE, Koehler G. Effective staining of tumor cells by coumarin-6 depends on the stoichiometry of cyclodextrin complex formation. J Incl Phenom Macrocycl Chem. 2012;70(3):327-331. Agócs G, Szabó BT, Köhler G, Osváth S. Comparing the Folding and Misfolding Energy Landscapes of Phosphoglycerate Kinase. Biophys J. 2012;102(12):2828-34. PMID: 22735533 Puchinger MG, Zarzer CA, Kügler P, Gaubitzer E, Köhler G. In vitro detection of adenocorticotropic hormone levels by fluorescence correlation spectroscopy immunoassay for mathematical modeling of glucocorticoid-mediated feedback mechanisms. EURASIP J Bioinform Syst Biol. 2012;2012(1):17. PMID: 23102048

Consecutive threading of cyclodextrin macrocycles on green fluorescent coumarin dye was studied using flu-orescence correlation spectroscopy. Inclusion complexes of up to three macrocycles could be resolved. This construct becomes an ideal water-soluble stain for lipid structures in live cell imaging.




29

R O B E R T K O N R A T

Computational biology and biomolecular NMR spectroscopy

In order to cope with this huge number of targets we introduced a new theoretical conception of protein structural biology (meta-structure) that can be used for protein sequence-to-function annotation and drug design. A hallmark of our research is the integrative application of this novel conception and sophisticated NMR spectroscopy

directed towards a better understanding of fundamental biological processes. Finally, as much of protein function is predicated on dynamics, we are developing novel methodological approaches that combine biochemistry, bioorganic chemistry and NMR spectroscopy to unravel the microscopic details of functionally important protein plasticity. Robert Konrat

TEAMNicolas CoudevylleAndrea FlammLeonhard GeistTanja GesellGeorg KontaxisDennis KurzbachKarin LedolterTomas SaraThomas SchwarzAgathe Vanas

The sequencing of the human genome has provided a “parts list” of the human inventory comprising potential therapeutic targets for the pharmaceutical and biotechnology industry.

SELECTED PUBLICATIONSHenen MA, Coudevylle N, Geist L, Konrat R. Toward Rational Fragment-Based Lead Design without 3D Structures. J Med Chem. 2012;55(17):7909-19. PMID: 22889313 Brüschweiler S, Konrat R, Tollinger M. Allosteric Communication in the KIX Domain Proceeds through Dynamic Repacking of the Hydrophobic Core. ACS Chem Biol. 2013;8(7):1600-10. PMID: 23651431 Kurzbach D, Schwarz TC, Platzer G, Höfler S, Hinderberger D, Konrat R. Compensatory adaptations of structural dynamics in an intrinsically disordered protein complex. Angew Chem Int Ed Engl. 2014;53(15):3840-3. PMID: 24604825

The individual stages of fragment-based lead drug design (FBLD). Starting from a suitable chosen small molecule fragment library, biophysical techniques are used to identify weak binders. (A) Structure-based FBLD exploits 3-D structural information about ligand binding modes to rationally evolve starting fragments in iterative rounds of optimizations. (B) Fragment evolution is performed by either merging individual fragments binding to different interaction sites or by ligand extension using me-dicinal chemistry substitution. (C,D) Meta-structure based fragment-based lead drug design strategies for ligand merging (C) and extension (D). (C) Meta-structure homologies are used to discern putative binding modes based on available 3-D structure information of the homologue. (D) Suitable sites for ligand derivatization are identified using ligand-based NMR spectroscopy (AFP-NOESY). Protons exposed to the solvent exhibit a sign inversion with increasing spin lock power (red). In contrast protons embedded in hydrophobic clusters display a markedly different behavior (blue) due to spin diffusion. This differential behavior can be used to identify suitable sites for ligand derivatization.





30

P A V E L K O V A R I K

Signaling and gene expression in inflammation

Inefficient or uncontrolled responses may result in infectious or inflammatory diseases. We investigate both the basic principles of balanced innate immune responses as well as aspects relevant for immune disorders. The molecular mechanisms that allow a robust yet temporally precisely restricted inflammatory reaction are studied at the level of transcription, mRNA stability and pathogen recognition. Turning on/off and resetting inflammatory gene transcription

The Stat transcription factors play a central role in the immune system. An open question is the mo-lecular mechanism that determines how often one activated Stat molecule can initiate transcription before becoming inactivated. Our recent studies suggest that the still not well understood S727 kinase is a chromatin-associated enzyme involved in the feedback control of the transcription cycle at

the targeted gene. We are currently characterizing the kinases and their role in the Stat-dependent transcription cycle. Control of immune homeostasis by mRNA stability

A considerable proportion of the genes in-duced during the acute phase of inflammation are strongly regulated at the level of mRNA stability. Many proinflammatory mRNAs contain regulatory elements (AREs) that are targeted by RNA-sta-bilizing and -destabilizing proteins. The RNA-de-stabilizing protein TTP plays a fundamental role in attenuating inflammation. Our newest findings revealed that TTP is one of the effector molecules of the anti-inflammatory cytokine IL-10. We are currently studying the role of TTP in inflammatory diseases using animals with conditional ablation of the TTP gene. Responses of innate immune cells to Streptococcus pyogenes

S. pyogenes is a Gram-positive human pathogen causing mild (e.g. tonsillitis) as well as severe (e.g. toxic shock) diseases. It is still not known how this bacterium is recognized by the innate immune system. We have recently shown that, surprisingly, S. pyogenes is recognized by a receptor that is distinct from any so far described receptors for bacterial pathogens. The identification of the re-ceptor for S. pyogenes and the elucidation of in-flammatory signaling cascades in the host cells are currently the major goals of the project.

Pavel Kovarik

TEAMVirginia Castiglia Ursula Damböck Florian Ebner Clemens Gaumannmüller Masa Ivin Marton Janos Klemens Pranz Vitaly Sedlyarov Iris Steinparzer

The innate immune system dynamically responds to infecting pathogens by initiating protective host responses and rapidly terminating these responses once the infection agent is no longer present.

SELECTED PUBLICATIONSBancerek J, Poss ZC, Steinparzer I, Sedlyarov V, Pfaffenwimmer T, Mikulic I, Dölken L, Strobl B, Müller M, Taatjes DJ, Kovarik P. CDK8 Kinase Phosphorylates Transcription Factor STAT1 to Selectively Regulate the Interferon Response. Immunity 2013;38(2):250-62. PMID: 23352233 Kratochvill F, Machacek C, Vogl C, Ebner F, Sedlyarov V, Gruber AR, Hartweger H, Vielnascher R, Karaghiosoff M, Rülicke T, Müller M, Hofacker I, Lang R, Kovarik P. Tristetraprolin-driven regulatory circuit controls quality and timing of mRNA decay in inflammation. Mol Syst Biol. 2011;7:560. PMID: 22186734 Gratz N, Hartweger H, Matt U, Kratochvill F, Janos M, Sigel S, Drobits B, Li XD, Knapp S, Kovarik P. Type I interferon production induced by Streptococcus pyogenes-derived nucleic acids is required for host protection. PloS Pathog. 2011;7(5):e1001345. PMID: 21625574

CDK8 occupancy at IFN-γ-regulated genes is STAT1-dependent. CDK8 controls IFN-stimulated transcription in two ways: (i) CDK8 signals upstream by phosphorylating STAT1 in the transactivation domain (S727) resulting in gene expression changes. (ii) CDK8 directly regulates the transcription machinery. Together, these regulatory steps are required for antiviral response.





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H E I N R I C H K O W A L S K I

Molecular and structural biology of picornaviruses

Infections by these viruses are normally re-stricted to the upper respiratory tract leading to the “common cold”, but they are also implicated in more severe pathologies such as bronchiolitis and ex-acerbation of asthma upon spreading to the lung. Due to the ~160 different rhinovirus genotypes, vaccination is not practical and no approved anti- rhinoviral drug is currently available. By molecularly characterizing as yet ill-defined steps in the life-cy-cle of these prototypic enteroviruses we also hope to contribute to the discovery of novel drug targets.

We are presently studying the following topics funded by the Austrian Science Fund (FWF): Uncoating of the rhinovirus RNA genome

Primary aim of the project is to determine the conformation of the viral RNA in- and outside the virion, to dissect essential interactions with the cap-sid and to elucidate their dynamic changes during uncoating. In addition, we noticed that uncoating is promoted by host factor(s), which will be identified using novel experimental setups. The results will considerably advance our understanding of the rules governing the ordered disassembly of virus particles for successful invasion of their host cell.

Strategies towards analysis of non-cultivatable viruses

Focusing on C-type rhinoviruses, we have started to generate virus-like particles as surrogates for structural analysis. In addition, we conduct reverse genetics for targeting these viruses towards certain integrins expressed on many cells in tissue culture. By deliberately changing their host tropism, we hope to expedite and facilitate exploration of these fas-tidious viruses. Role of N-myristyltransferase family members in the covalent modification of rhinovirus proteins

Furthermore, we seek to uncover the role of N-myristoyltransferase family members NMT1 and NMT2 in the myristoylation of rhinoviral proteins, a vital modification in most picornaviruses. Due to its better growth in the haploid cell line HAP1 (www.haplogen-genomics.com/faq/hap1) the work has been expanded to Coxsackievirus employing knock-out HAP1 cells lacking NMT1 or NMT2.

Heinrich Kowalski

TEAMIrena Corbic-Ramljak

Our research is dedicated to unraveling the molecular mechanisms and structural changes underlying the invasion of host cells by small RNA viruses and the role of host factors for their efficient replication, mainly using human rhinoviruses as a model.

SELECTED PUBLICATIONSHarutyunyan S, Kowalski H, Blaas D. The Rhinovirus Subviral A-Particle Exposes 3‘-terminal Sequences of its Genomic RNA. J Virol 2014; 88(11):6307-17. PMID: 24672023 Harutyunyan S, Kumar M, Sedivy A, Subirats X, Kowalski H, Koehler G, Blaas D. Viral Uncoating Is Direc-tional: Exit of the Genomic RNA in a Common Cold Virus Starts with the Poly-(A) Tail at the 3´-End. PLoS Pathog. 2013;9(4):e1003270. PMID: 23592991 Weiss VU, Subirats X, Kumar M, Harutyunyan S, Gösler I, Kowalski H, Blaas D. Capillary Electrophoresis, Gas-Phase Electrophoretic Mobility Molecular Analysis, and Electron Microscopy: Effective Tools for Quality Assessment and Basic Rhinovirus Research. Meth Mol Biol 2015;1221:101-128. PMID: 25261310

(a) N-myristoyltransferase isozyme NMT1 or NMT2 knock-out in Hap1 cells results in fewer eGFP-positive (virus-producing) cells upon infection with re-combinant Coxsackievirus-B3 (CVB3-eGFP) at different virus-cell-ratio (MOI) compared to wild type. (b) RT-qPCR shows drastic reduction of viral progeny.

http://www.haplogen-genomics.com/faq/hap1

http://www.haplogen-genomics.com/faq/hap1





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C L A U D I N E K R A F T

Regulation and signaling in autophagy

One of the major cellular responses during fasting is the activation of the lysosomal degradation pathway of autophagy, a process in which the cell digests its own components. By this mechanism, the cell not only provides nutrients to maintain vital cellular functions during fasting but can also rid the cell of superfluous or damaged organelles, misfolded proteins, and invading microorganisms. Interestingly, autophagy is now emerging as a central biological pathway which functions to promote health and longevity. It has been found to play an important role in several human diseases including neurodegeneration and cancer.

One of the key regulators of autophagy is the TOR kinase, which is the major inhibitory signal that shuts off autophagy in the presence of nutrients and growth factors. Under nutrient limiting con-ditions, when TOR kinase is inactive, autophagy is induced, which results in the sequestration of cytosol and organelles into a double-membrane compartment followed by their subsequent

delivery to the vacuole/lysosome for breakdown and recycling.

Our understanding, however, of the mechanics of autophagy regulation, as well as its role in human health and disease, is still at an early stage. The major upstream regulator of autophagy is the Atg1 kinase complex, which is believed to receive directly the nutrient signal from TOR, and in response, to activate or inhibit the autophagic machinery. • We are studying the role and function of

Atg1 kinase in the regulation of autophagy in budding yeast, using phospho-proteomic approaches, genetic screening techniques as well as modern biochemical and cell biological methods. We are mainly focusing on two questions:

• How does the Atg1 kinase regulate autophagy function, e.g. what are the physiological substrates?

• How is the Atg1 kinase regulated?

These approaches aim in understanding the mechanistics of a central player in autophagy regulation. Since the Atg proteins are highly conserved in other eukaryotes, this work will also help us to better understand diseases associated with autophagy misregulation.

Claudine Kraft

TEAMLevent BasThorsten BrachAndrea BrezovichTamara MatzholdDaniel PapinskiThaddaeus PfaffenwimmerMartina SchuschnigRaffaela Torggler

Fasting has been part of health practices from ancient times to the present. This tradition may be partially rooted in a cellular process we are now beginning to understand in modern scientific terms.

SELECTED PUBLICATIONSPfaffenwimmer T, Reiter W, Brach T, Nogellova V, Papinski D, Schuschnig M, Abert C, Ammerer G, Martens S, Kraft C. Hrr25 kinase promotes selective autophagy by phosphorylating the cargo receptor Atg19. EMBO Rep. 2014;15(8):862-70. PMID: 24968893 Papinski D, Schuschnig M, Reiter W, Wilhelm L, Barnes CA, Majolica A, Hansmann I, Pfaffenwimmer T, Kijanska M, Stoffel I, Lee SS, Brezovich A, Lou JH, Turk BE, Aebersold R, Ammerer G, Peter M, Kraft C. Early Steps in Autophagy Depend on Direct Phosphorylation of Atg9 by the Atg1 Kinase. Mol Cell. 2014;53(3):471-83. PMID: 24440502 Kraft C, Kijanska M, Kalie E, Siergiejuk E, Lee SS, Semplicio G, Stoffel I, Brezovich A, Verma M, Hansmann I, Ammerer G, Hofmann K, Tooze S, Peter M. Binding of the Atg1/ULK1 kinase to the ubiquitin-like protein Atg8 regulates autophagy. EMBO J 2012;31(18):3691-703. PMID: 22885598

Schematic of the autophagy pathway. The nutrient signal is sensed by the PAS. Autophagy is then induced, and membranes are formed at the PAS. Membranes grow and engulf cytoplasmic components, which can include entire organelles. Membranes fuse to form a double-membraned autophagosome. The outer membrane fuses with the vacuolar membrane, releasing the inner autophagic vesicle into the lumen of the vacuole, where it is degraded by the vacuolar proteases. The Atg1 kinase is part of the PAS core and regulates the shuttling of membrane vesicles between the PAS and peripheral membrane structures.





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K A R L K U C H L E R

Host-pathogen interactions & mechanisms of fungal virulence

First, we use reverse genetics approaches to identify virulence and antifungal drug resistance genes in most prevalent human fungal pathogens such as Candida glabrata and C. albicans. For instance, we have generated a genome-scale gene deletion col-lection of C. glabrata currently comprising some 700 single deletions strains suitable to identify novel drug resistance genes.

Further, we decipher the roles of histone mod-ification in morphogenetic switching, cell fate determination and fungal virulence. We define the chromatin-related genetic networks and signaling pathways facilitating immune evasion and driving invasion of host cells. We also study the genomic and genetic adaptations occurring in pathogen genomes during host niche or organ colonization and systemic dissemination.

On the host side, we are investigating the mechanisms of host-pathogen interaction and cytokine signaling response in primary dendritic cells/macrophages related to the inflammatory immune response. Moreover, we exploit mouse infection models to decipher the role of host immune signaling in fungal pathogenesis. Along this line, we delineate the interplay of adaptive and innate immunity in immune surveillance, and particularly the role of Tec kinases and type I interferons in microbial inflammation and dissemination in the host.

Finally, we study the molecular structure-function relationships of eukaryotic ABC multidrug tran- sporters, especially those implicated in hepatic detoxification processes.

Karl Kuchler

TEAMAnna CsekeAybala EkerFabian IstelSabrina JenullNarakorn KhunweeraphongRegina KlausFilomena NogueiraLeonel PereiraAndriy PetryshynMichael RiedelbergerRaju ShivarathriSaren TasciyanMichael TschernerJing XieFlorian Zwolanek

We study the molecular mechanisms of fungal pathogenicity and fundamental problems in infection biology, using a combination of both molecular and genome-wide systems biology approaches.

SELECTED PUBLICATIONSHnisz D, Bardet A, Nobile C, Schoeck U, Petryshin A, Glaser W, Stark A, Kuchler K. Histone deacetylation at coding sequences adjusts tran-scription kinetics during Candida albicans morphogenesis. PLoS Genet. 2012;8(12):e1003118. PMID: 23236295 Majer O, Bourgeois C, Zwolanek F, Lassnig C, Kerjaschki D, Mack M, Müller M, Kuchler K. Type I interferons promote fatal immunopathology by regulating inflammatory monocytes and neutrophils during Candida infections. PLoS Pathog. 2012;8(7):e1002811. PMID: 22911155 Tierney L, Linde J, Müller S, Brunke S, Molina JC, Hube B, Schöck U, Guthke R, Kuchler K. An Interspecies Regulatory Network Inferred from Simultaneous RNA-seq of Candida albicans Invading Innate Immune Cells. Front Microbiol. 2012;3:85. PMID: 22416242

Candida albicans cells forming colonies of markedly different phenotypes on agar plates due to distinct chromatin modifications, which modulate transcriptional regulatory networks controlling morphogenesis.





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T H O M A S L E O N A R D

Structural biology of lipid-activated signal transduction

Dysregulation of signal transduction at these sites is the cause of a number of hereditary and non-hereditary diseases, including Coffin-Lowry syndrome, spinocerebellar ataxia, myotonic dystrophy, and various cancers. Over 500 kinases and 130 phosphatases regulate signal transduction, and whilst much is known about how these pro-teins are targeted to cellular membranes, very little is known about how membrane engagement is coupled to signal transduction. We are addressing two questions central to signal transduction at membranes.

One of the most important consequences of the activation of cell surface receptors is the generation of small molecule second messengers.

A number of cellular second messengers are lipids. Many of the lipid responsive human protein kinases belong to the AGC family of kinases. We would like to understand how lipid-engagement by these protein kinases is coupled to their activation at the molecular level. We hope to elucidate common principles of the molecular mechanisms that govern lipid-mediated signal transduction.

Our most recent work focuses on how the Rho-associated kinases (ROCK) control acto-myosin contraction in the actin cytoskeleton, a process that permits cells to move and change their shape according to their needs. We have recently determined the structure of human ROCK2, characterized it biochemically, and are currently exploring the mechanism by which it transduces membrane-based signals in vivo.

The second question relates to how signal transduction pathways are organized. Scaffolding of signaling proteins in the same pathway enhances specificity, promotes signal amplification by reducing noise, and improves signal propagation through the pathway. Membranes act as the scaffolds for many signaling reactions, including those involved in regulating cellular growth processes. Our studies are aimed at understanding how diverse signals are integrated, how substrate specificity is encoded not just at the kinase level, and the influ-ence of the membrane environment on multi- component signaling hubs. This is an exciting area of research with frontiers in ageing, cancer, as well as metabolic diseases such as diabetes and obesity.

Thomas Leonard

TEAMDaniel ElsnerIva LucicLinda Trübestein Freia von Raußendorf

Membranes are sites of intense signaling activity in eukaryotic cells. Essential processes such as autophagy, cytokinesis, exo- and endocytosis, and cytoskeletal remodeling depend on signal propagation at cellular membranes.

SELECTED PUBLICATIONSLeonard TA, Hurley JH. Regulation of protein kinases by lipids. Curr Opin Struct Biol. 2011;21(6):785-91. PMID: 22142590 Leonard TA, Thomas A; Rózycki B, Saidi LF, Hummer G, Hurley JH. Crystal structure and allosteric activation of protein kinase C βII. Cell. 2011; 144(1):55-66. PMID: 21215369 Oliva MA, Halbedel S, Freund SM, Dutow P, Leonard TA, Veprintsev DB, Hamoen LW, Löwe J. Features critical for membrane binding revealed by DivIVA crystal structure. EMBO J 2010;29(12):1988-2001. PMID: 20502438

A. Model of ROCK activation. ROCK is maintained in an inactive, autoinhibited state in the cytosol by interactions between its regulatory and kinase domains. Upon membrane binding and Rho activation, the inhibitory interactions are displaced, rendering ROCK active. B. ROCK regulates actomyosin contraction by phosphorylating myosin II. ROCK stimulates actomyosin contraction by phosphorylating the N-terminus of the regulatory myosin light chain (RMLC) of myosin II. Phosphorylated RMLC stimulates the ATPase activity of the myosin motor domain, resulting in contraction.





35

J O S E F L O I D L

Meiotic chromosome pairing and recombination

During meiosis, homologous chromosomes of paternal and maternal origin juxtapose and become connected by a protein structure, the synaptonemal complex (SC). They then exchange parts and segregate to different daughter nuclei. Thus, the function of meiosis is twofold: it compensates for the doubling of the chromosome number at fertilization and it provides the progeny with newly assorted sets of alleles, which is the basis of their genetic heterogeneity. Failures in meiosis may lead to gametes with aberrant chromosome numbers and thus to progeny with congenital defects.

We are studying various aspects of meiotic chromosome organization and behavior in evolutionarily divergent organisms such as yeasts and ciliates, to learn which adaptations and amendments have occurred during the evolution of extant meiosis.

In particular, we are investigating meiotic chromosome pairing and recombination in the SC-less ciliate Tetrahymena. Spo11-dependent DNA double- strand breaks cause an enormous elongation of meiotic nuclei. The ensuing close parallel and polarized arrangement of chromosomes within

the tubular nucleus, which is reminiscent of the conserved bouquet organization, promotes homologous pairing and recombination. A similar, although less pronounced, change in nuclear shape is observed in the fission yeast. Thus, this adaptation may have evolved to compensate for the lack of an SC in these two organisms. Moreover, the absence of the SC may have led to the processing of meiotic recombination intermediates by a single simplified crossover pathway. This combines features of two distinct pathways, which prevail in the majority of eukaryotes. Currently we are performing mutant analyses to identify novel meiotic genes (example shown in the figure), which have no homologs in other organisms, to reach a better understanding of the adaptations that have allowed Tetrahymena to undergo its streamlined meiosis.

Ultimately, our studies will help to understand the origin and function of conserved meiotic features such as the SCs, the chromosomal bouquet, and the regulation of meiotic recombination.

Josef Loidl

TEAMEmine I. AliRachel Howard-TillAnura Shodhan

Meiosis is a pivotal process in the sexual reproduction cycle: it compensates for the doubling of the chromosome number at fertilization and it provides the progeny with newly assorted sets of alleles, which is the basis of their genetic heterogeneity.

SELECTED PUBLICATIONSLoidl J. (2013). The hidden talents of SPO11. Dev Cell. 2013;24(2):123-124. PMID: 23369711 Shodhan A, Lukaszewicz A, Novatchkova M, Loidl J. Msh4 and Msh5 function in SC-independent chiasma formation during the streamlined meiosis of Tetrahymena. Genetics 2014;198(3):983-93. PMID: 25217051 Chi J, Mahé F, Loidl J, Logsdon J, Dunthorn M. Meiosis gene inventory of four ciliates reveals the prevalence of a synaptonemal complex-independent crossover pathway. Mol Biol Evol. 2014;31(3):660-72. PMID: 24336924

Conjugated pairs of Tetrahymena cells undergo synchronous meioses. Whereas in the wild type, meiosis I bivalents congress at a metaphase plate (and will subsequently separate), the knockout of a novel meiotic gene causes the 5 bivalents of each cell to arrange in a tandem configuration and prevents them from separation. (The large round structures are the somatic nuclei, which do not undergo meiosis.)





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S A S C H A M A R T E N S

Molecular mechanisms of autophagy

Autophagy plays an essential role during starva-tion, the defense against pathogenic microorgan-isms, the removal of protein aggregates and the degradation of damaged organelles. Misregulated or defective autophagy can result in neurodegeneration and premature aging and is thus highly relevant to a plethora of human diseases.

Autophagy is induced by an upstream signal such as starvation, the detection of pathogenic microorganisms in the cytosol or by damaged mito- chondria. This signal triggers the most enigmatic and fascinating step of autophagy, the de novo formation of autophagosomes. Initially a small double membrane bound structure is formed, which grows and adopts the shape of a cup. This cup-shaped structure eventually fuses at its rims to form a double membrane bound organelle enclos-ing a part of the cell’s cytoplasm. The autophago-

some then fuses with components of the classical endosomal system thereby maturing to an autoly-sosome within which the content is degraded. The degraded content can subsequently be used for the synthesis of factors that are essential for the survival of the cell.

Although many genes that are important for autophagy have been identified we have only a very limited understanding of how this important and fascinating process is regulated and executed. Thus, the challenge now is to assign functions to these genes in order to gain a better understanding of the mechanisms that orchestrate autophagy.

We are a multidisciplinary team employing cell biology, biochemistry, light- and electron microscopy as well as structural biology ap-proaches. Our findings will give important insights into mechanisms of autophagy.

Sascha Martens

TEAMChristine AbertDorotea FracchiollaNina HoblVeronika NogellovaJulia RomanovJustyna Sawa-MakarskaBettina WurzerGabriele ZaffagniniBettina Zens

Autophagy is an evolutionarily conserved and important process during which our cells cannibalize small parts of themselves.

SELECTED PUBLICATIONSSawa-Makarska J, Abert C, Romanov J, Zens B, Ibiricu I, Martens S. Cargo binding to Atg19 unmasks additional Atg8 binding sites to mediate membrane-cargo apposition during selective autophagy. Nat Cell Biol. 2014;16(5):425-33. PMID: 24705553 Romanov J, Walczak M, Ibiricu I, Schüchner S, Ogris E, Kraft C, Martens S. Mechanism and functions of membrane binding by the Atg5-Atg12/Atg16 complex during autophagosome formation. EMBO J. 2012;31(22):4304-17. PMID: 23064152 Kraft C, Martens S. Mechanisms and regulation of autophagosome formation. Curr Opin Cell Biol. 2012;24(4):496-501. PMID: 22664348

Scheme showing the generation of autophagosomes. Initially a small double membrane-bound structure called isolation membrane is formed. This structure expands to adopt a cup-like shape thereby gradually enclosing cytoplasmic cargo. This structure fuses at its rims giving rise to the mature autophagosome. Subsequently, autophagosomes fuse with lysosomes. Within these so-called autolysosomes the inner membrane and the cargo are degraded.

(A) A picture taken by confocal microscopy showing giant unilamellar vesicles (GUVs). The membrane of the GUVs was labelled by incorporation of a fluorescent lipid. (B) A picture showing human cells which express green and red labelled proteins that are targeted to autophagosomes.





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I S A B E L L A M O L L

Bacterial stress response and ribosome heterogeneity

Decoding mRNA-based information into the corresponding sequence of amino acids is per-formed by a complex ribonucleoprotein particle, the ribosome. Traditionally, the ribosome is viewed as highly conserved machinery with an invariable RNA and protein complement. The main aim of my group is to decipher molecular mechanisms that lead to modulation of the translational program by ribosome heterogeneity. Our initial studies revealed that antibiotic treatment of Escherichia coli can result in the formation of ribosomes that lack several proteins. Intriguingly, these ribosomes are selective for translation of leaderless mRNAs, which lack a 5´-untranslated region.

Subsequently, we have elucidated a novel post-transcriptional stress response mechanism in E. coli, which is based on the functional specialization of ribosomes by the stress-induced endoribo-nuclease MazF, the toxin component of a toxin- antitoxin system. Upon activation, MazF targets the 16S rRNA in the context of the ribosome and removes its 3´-terminus harboring the anti- Shine-Dalgarno (SD) sequence. As the SD-aSD interaction is required for translation initiation on canonical mRNAs, the modified ribosomes exclusively translate leaderless mRNAs that lack the SD-sequence. As MazF con-comitantly generates distinct leaderless transcripts by removing their 5´-UTR, the translational program is modulated to adapt protein synthesis to the given conditions. Currently, we are studying a mechanism that allows the reversion of the 16S rRNA processing, facilitating the growth recovery when conditions ameliorate.

Besides, we aim to structurally and functionally characterize the essential ribosomal protein S1, which is absent in the high-resolution images of the E. coli ribosome. Recently, we solved the

structure of the boundary between S1 and the ribo-some in detail, showing that S1 is anchored primarily via its short flexible N-terminal segment, stabilized by salt bridges that involve the zinc binding pocket of protein S2. Overall, this work provides one hitherto enigmatic piece in the “ribosome puzzle”, and suggests novel mechanisms entailing the modulation of protein synthesis in response to environmental cues by changing the affinity of S1 for the ribosome.

Collectively, our studies challenge a long term paradigm in biology by raising the physiological role of the ribosome from the general protein synthesis machinery to a central control unit of gene expression with an intrinsic regulatory capacity.

Isabella Moll

TEAMTanino AlbaneseGisela GrögerNela NikolicMartina SauertHannes Temmel

One of the most intricate and fundamental processes of life is the translation of the genetic code into proteins.

SELECTED PUBLICATIONSByrgazov K, Grishkovskaya I, Arenz S, Coudevylle N, Temmel H, Wilson DN, Djinovic-Carugo K, Moll I. Structural basis for the interaction of protein S1 with the Escherichia coli ribosome. Nucleic Acids Res. 2014; [Epub ahead of print]. PMID: 25510494 Byrgazov K, Manoharadas S, Kaberdina AC, Vesper O, Moll I. Direct Interaction of the N-Terminal Domain of Ribosomal Protein S1 with Protein S2 in Escherichia coli. PLoS One. 2012;7(3):e32702. PMID: 22412910 Vesper O, Amitai S, Belitsky M, Byrgazov K, Kaberdina AC, Engelberg-Kulka H, Moll I. Selective Translation of Leaderless mRNAs by Specialized Ribosomes Generated by MazF in Escherichia coli. Cell. 2011;147(1):147-57. PMID: 21944167

(A) Interaction between the N-terminal domain of S1 (S1NTD, blue) and protein S2 (yellow) shown by crystal structure analysis. (B) Cryo-EM structure of a translating 70S ribosome containing additional density for S1 domains D1 (S1D1, blue) and D2 (S1D2, cyan). (Byrgazov et al., Nucleic Acids Res., 2014)





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E R N S T M Ü L L N E R

Erythrocyte (patho)physiology

In one project, we investigate the autonomous physiological impact of erythrocytes in thrombus formation, so far exclusively ascribed to platelets. According to preliminary data red blood cells (RBCs) may be central to the pathophysiology of cancer-associated thromboembolism. The lipid mediator lysophosphatidic acid (LPA) is not only secreted locally from activated platelets but also found systemically in the blood of tumor patients. LPA induces accumulation of phosphatidylserine (PS) at the extracellular leaflet of the erythrocyte membrane. This generates binding sites for the pro-thrombinase complex and suggests an important active role for erythrocytes in thromboembolism. We identified a G-protein coupled receptor-based activation of the PI3K and the PKC pathway inducing calcium influx into erythrocytes. This abrogates the phospholipid asymmetry between the membrane leaflets. Apart from elucidating molecular mechanisms we aim at identifying inhibitors of that process that may have potential for future clinical interventions against thrombo-embolism in cancer patients.

The second approach aims to delineate molecular mechanisms underlying the pathology of rare con-

genital neurodegenerative disorders, i.e. chorea ac-anthocytosis, McLeod syndrome and pantothenate kinase associated neurodegeneration. They are caused by defects in the chorein/VPS13A protein, the XK protein and the Pank2 protein, respectively, and are associated with acanthocytosis, the occurrence of misshaped erythrocytes with thorny protrusions. Hence these disorders are collectively classified as neuroacanthocytosis syndromes. The functions of these proteins in RBCs or neu-rons and the respective mechanisms underlying neurodegeneration are largely unknown. Studying patient blood samples, we explore commonalities and differences in RBC properties between these three syndromes. Thereby, we found altered responses in endocytosis and, again, identified LPA signaling as being dysregulated. This results in aberrant calcium uptake and PS exposure (Siegl et al., 2013). Currently we are identifying the molecular details of these pathological changes to understand the morphological alterations during acanthocytosis and thus provide evidence for a possibly reduced functionality/stability of erythrocytes or even participation in the neurodegeneration process.

Contrary to common belief erythrocytes are far more than bags full of hemoglobin. They are equipped with a large set of signaling components, emerging as important players in various physiologic processes.

SELECTED PUBLICATIONSKostan J, Salzer U, Orlova A, Toro I, Hodnik V, Senju Y, Zou J, Schreiner C, Steiner J, Merilainen J, Nikki M, Virtanen I, Carugo O, Rappsilber J, Lappalainen P, Lehto VP, Anderluh G, Egelman EH, Djinovic-Carugo K. Direct interaction of actin filaments with F-BAR protein pacsin2. EMBO Rep. 2014;15(11):1154-62. PMID: 25216944 Siegl C, Hamminger P, Jank H, Ahting U, Bader B, Danek A, Gregory A, Hartig M, Hayflick S, Hermann A, Prokisch H, Sammler EM, Yapici Z, Prohaska R, Salzer U. Alterations of red cell membrane properties in neuroacanthocytosis. PloS One. 2013;8(10):e76715. PMID: 24098554 Kerenyi MA, Grebien F, Gehart H, Schifrer M, Artaker M, Kovacic B, Beug H, Moriggl R, Mullner EW. Stat5 regulates cellular iron uptake of erythroid cells via IRP-2 and TfR-1. Blood. 2008;112(9):3878-88. PMID: 18694996

Lysophosphatidic acid (LPA) promotes active participation of erythro-cytes in coagulation by rapid externalization of phosphatidylserine in a pathway involving G proteins, PI3K and PKC.

Reduced calcium uptake into ChAc (red) as compared to control (blue) erythrocytes upon stimulation with LPA. Flow cytometry of intracellular calcium levels was done with Fluo-3.

Ernst Müllner

TEAMSebastian GranitzerNina KüntzelThomas ÖhlingerUlrich SalzerMaike Werning





39

J O H A N N E S N I M P F

ApoER2 and VLDL receptor

The biological systems we are working with are the chicken oocyte and the mammalian brain. These two systems reflect the functional dichotomy of the receptors which function in endocytosis (follicles) and signal transduction (brain development). The best characterized function of VLDLR in follicles of egg laying species is endocytosis of yolk precursors into the growing oocyte.

These yolk precursors (VLDL and Vitellogenin) are synthesized in the liver and rapidly taken up by the growing oocyte. Recently, we have started to elucidate cell signaling functions of VLDLR and ApoER2 in granulosa cells which support the maturation of oocytes within the follicle. In respect to brain development both receptors act as Reelin- signal transducers. The Reelin signal orchestrates the correct positioning of newly generated neurons within laminated structures of the brain. In the development of the olfactory system in rodents, the

structure of the olfactory bulb depends on neurons generated throughout life in the subventricular zone. These neurons migrate via the rostral migratory stream towards the olfactory bulb. This migration also depends on the presence of ApoER2 and VLDLR but seems to be independent on Reelin. To this end we have characterized thrombospondin-1 and clusterin as novel ligands for ApoER2 and VLDLR.

Currently, we are focusing on a potential in-volvement of the receptors in the development of Alzheimer’s disease. A key feature of this disease is an imbalance of production and clearance of the Aβ-peptide which results in amyloid deposits in the brain. Since clusterin forms complexes with the Aβ-peptide, these complexes might get removed from the extracellular space by the action of ApoER2 and VLDLR.

Johannes Nimpf

TEAMPaula DlugoszSimon NimpfHarald Rumpler

We study the biology of LDL receptor related proteins (VLDL receptor and ApoER2), a group of cell surface receptors which mediate transport of macromolecules across cell membranes and play important roles in signal transduction.

SELECTED PUBLICATIONSRiegler B, Besenboeck C, Bauer R, Nimpf J, Schneider WJ. Enzymes involved in hepatic acylglycerol metabolism in the chicken. Biochem Biophys Res Commun. 2011; 406(2):257-61. PMID: 21316342 Duit S, Mayer H, Blake SM, Schneider WJ, Nimpf J. Differential functions of ApoER2 and VLDL receptor in Reelin signaling depend on differential sorting of the receptors. J Bio Chem. 2010;285(7):4896-908. PMID: 19948739 Hong C, Duit S, Jalonen P, Out R, Scheer L, Sorrentino V, Boyadjian R, Rodenburg KW, Foley E, Korhonen L, Lindholm D, Nimpf J, van Berkel TJ, Tontonoz P, Zelcer N. The E3 ubiquitin ligase IDOL induces the degradation of the low density lipoprotein receptor family members VLDLR and ApoER2. J Biol Chem. 2010;285(26):19720-6. PMID: 20427281

Model of the intracellular fates of ApoER2 and VLDLR upon Reelin stimulation. Upon binding of Reelin, both ApoER2 and VLDLR mediate phosphorylation of Dab1 (1). VLDLR internalizes Reelin rapidly via Clathrin-mediated endocytosis (2) and is separated from Reelin in the compartment of uncoupling of receptor and ligand (3). VLDLR then recycles back to the plasma membrane (4) while Reelin is delivered to the lysosome for degradation (5). ApoER2 internalizes Reelin via the same pathway although the receptor originally resides in lipid rafts and endocytoses its ligand with a much slower rate. In contrast to VLDLR, ApoER2 is not recycled but ends up in the lysosome together with Reelin (6). As an additional feedback mechanism, Reelin stimulation induces secretase-mediated cleavage of ApoER2, thereby generating a soluble extracellular fragment (8). This fragment can, together with another N-terminal fragment produced from an ApoER2 isoform by furin cleavage (9), inhibit the Reelin signal by sequestering free Reelin in the cell’s sur- rounding. The function of the soluble intracellular domain of ApoER2 is not well understood yet.





40

E G O N O G R I S

PP2A enzyme biogenesis and monoclonal antibodies

A major phosphoserine/threonine phosphatase in the cell is protein phosphatase 2A (PP2A), a known tumor suppressor and target of cancer causing viruses. PP2A comprises a family of many different multisubunit holoenzymes, each of which possesses unique substrate specificity dependent on its subunit composition. Decreased PP2A activity is associated with the development of hu-man diseases indicating an important role of prop-erly regulated PP2A for cellular and organismal homoeostasis. Our goal is to understand the molecular mechanisms of PP2A holoenzyme assembly and to identify the substrates regulated by PP2A.

The interactions between PP2A and its substrates are short-lived and thus hard to detect with standard biochemical methods. By using a detection method, termed M-Track (for Methyl-Tracking), which by itself is based on a short-lived enzyme-substrate reaction we are able now to capture PP2A “in flagrante” with substrates and we are using this method for PP2A substrate validation and identification.

Our study of PP2A biogenesis in yeast led to a

model, in which a chaperone-dependent activation step is coupled to methylation-dependent holoenzyme assembly and is under the surveillance of the PP2A methylesterase PPE1. How PP2A biogenesis is regulated, for example by subunit phosphorylation, is unknown and currently investigated in the lab. Another level of control occurs through PP2A specific inhibitors such as endosulfines. Despite the high degree of conservation not much is known about PP2A biogenesis in higher eukaryotes. How-ever, there are indications that dysfunctional PP2A biogenesis might be causally involved in disease development. Thus, we are studying PP2A biogenesis by biochemical, immunological and genetic approaches in mammalian cells.

A second business‐oriented research direction deals with the generation of high quality mono-clonal research antibodies such as monoclonals specific for human disease-linked proteins and posttranslational modifications. Moreover, we are developing novel immunoblotting tools such as the “Light Your Marker” technology (patent application filed) and are providing a monoclonal antibody service to the research community.

Egon Ogris

TEAMPeter AndorferBhumika BhattIngrid FrohnerElias KremmingerStephanie KronlachnerThomas KupkaMarie LangFlorian MartysIngrid MudrakFranziska MüllerStefan SchüchnerJiri Veis

Cells employ reversible protein phosphorylation to regulate the functional state of their proteins. The enzymes catalyzing these reactions, the protein kinases and phosphatases, are important regulators of almost all aspects of life.

SELECTED PUBLICATIONSZuzuarregui A, Kupka T, Bhatt B, Dohnal I, Mudrak I, Friedmann C, Schüchner S, Frohner IE, Ammerer G, Ogris E. M-Track: detecting short-lived protein-protein interactions in vivo. Nature Methods. 2012;9(6):594-6. PMID: 22581371 Juanes MA, Khoueiry R, Kupka T, Castro A, Mudrak I, Ogris E, Lorca T, Piatti S. Budding Yeast Greatwall and Endosulfines Control Activity and Spatial Regulation of PP2A(Cdc55) for Timely Mitotic Progression. PLoS Genet. 2013;9(7):e1003575. PMID: 23861665 Sontag JM, Nunbhakdi-Craig V, Mitterhuber M, Ogris E, Sontag E. Regulation of protein phosphatase 2A methylation by LCMT1 and PME-1 plays a critical role in differentiation of neuroblastoma cells. J Neurochem. 2010;115(6):1455-65. PMID: 21044074

Cartoon of an M-Track assay: Bait protein “Y”, prey protein “X” human histone lysine methyltransferase (HKMT), N terminus of histone H3 (H3K9). Upon interaction with the bait,the prey is stably marked by methylation (M, methyl group).

Cartoon of “Light Your Marker” technology. An antibody specific for the dye of the protein molecular weight markers allows visualization of the marker in the western blot analysis.





41

F R I E D R I C H P R O P S T

The neuronal cytoskeleton in axon guidance

These transformations are guided by extracellular signals which ultimately need to be translated into the rearrangement of the neuronal cytoskeleton. We study signaling mechanisms and posttranslational modifications of microtubule-associated proteins and other components of the cytoskeleton that regulate the orchestrated reorganization of microtubules and actin in response to extracellular signals. Our approach combines gene ablation in the mouse with cell biological and molecular analyses in cultured neurons and other primary cells.

One focus of our research is the role of micro-tubule-associated proteins of the MAP1 family. In a recent study we found that MAP1B is necessary for nitric oxide signaling to the cytoskeleton in axon retraction. Posttranslational modification of MAP1B by S-nitrosylation changes its interaction with microtubules. Thus, we showed that MAP1B is a component of a pathway that links calcium influx and activation of neuronal nitric oxide synthase to reconfiguration of axonal microtubules and might contribute to the physiological and pathological effects of nitric oxide in the brain. We have since expanded our investigation towards the role of S-nitrosylation of tubulin in neuronal morphogenesis. Another current topic is the role of MAP1B in repulsive axon guidance in the brain.

We have obtained evidence that MAP1B is essential for signal transduction of several unrelated repulsive axon guidance cues. We have also analyzed the functional properties of other MAP1 proteins and found that the light chains of these proteins determine to some extent their functional charac-teristics. Moreover, we characterized a novel mem-ber of the MAP1 family, which we termed MAP1S. MAP1S is expressed not only in the brain, but also in a wide range of other tissues and represents the non-neuronal counterpart of MAP1A and MAP1B.

We have generated MAP1S deficient mice and are exploring the role of this protein in cell division, cell migration, and tumorigenesis.

Friedrich Propst

TEAMRajeshwari MeliPetronela Weisova

Axon extension, axon branching, and axon retraction are major morphological changes that neurons have to execute to accomplish correct wiring of the nervous system during development and during regeneration after injury.

SELECTED PUBLICATIONSStroissnigg H, Trancíková A, Descovich L, Fuhrmann J, Kutschera W, Kostan J, Meixner A, Nothias F, Propst F. S-nitrosylation of microtubule-associated protein 1B mediates nitric-oxide-induced axon retraction. Nat Cell Biol. 2007;9(9):1035-45. PMID: 17704770 Barnat M, Enslen H, Propst F, Davis RJ, Soares S, Nothias F. Distinct roles of c-Jun N-terminal kinase isoforms in neurite initiation and elongation du-ring axonal regeneration. J Neurosci 2010; 30(23):7804-16. PMID: 20534829 Cheng L, Desai J, Miranda CJ, Duncan JS, Qiu W, Nugent AA, Kolpak AL, Wu CC, Drokhlyansky E, Delisle MM, Chan WM, Wei Y, Propst F, Reck-Peterson SL, Fritzsch B, Engle EC. Human CFEOM1 mutations attenuate KIF21A autoinhibition and cause oculomotor axon stalling. Neuron. 2014;82(2):334-49. PMID: 24656932

MAP1B is essential for nitric oxide-induced axonretraction. a) Cultured neurons from adult wild-type mice (dorsal root ganglion) were treated with the nitric oxide donor SNAP, fixed, stained for tubulin and analyzed by confocal fluorescence microscopy. Cellular morphology was scored as unchanged (left) or retracted (right). b) Cultured neurons from adult MAP1B+/+ or MAP1B-/- mice were treated with SNAP for 1 h and processed as above. Microtubule configuration was classified as unchanged (compared to untreated cells) or displaying retraction hallmarks (sinusoidal bends along the axon, a trailing remnant, and a re-traction bulb). MAP1B-/- neurons displayed a severely reduced capacity to respond to SNAP by axon retraction.





42

F L O R I A N R A I B L E

Hormonal control of animal energy expenditure

Our past work has shown that this marine worm exhibits a unique combination of ancestral-type genomic characteristics not found in insect and nematode model species. Moreover, we have iden-tified numerous components of ancestral-type hor-mone pathways in Platynereis.The hormonal control of reproduction and regen-eration

What could be the function of ancestral-type hormones in Platynereis? One of the systems that we aim to dissect is the hormonal machinery or-chestrating reproduction and regeneration. Platy-nereis is an excellent object for this analysis: it has been a central model for regeneration, and for the link between chronobiology and reproduction. Our molecular analyses have identified a spectrum of conserved hormones in Platynereis. Thanks to the establishment of new molecular tools, we are now able to systematically assess the impact of these candidates on the development and maturation of the animals.

These experiments are supported by an ERC starting grant (HOR.MOON) as well as the inter-

disciplinary research platform “Marine rhythms of life”, and provide us with first molecular insight into the enigmatic hormone machinery underlying lunar reproductive periodicity. Exploring a new marine model system

Over the past years, Platynereis has emerged as a very promising “next-generation” model system. We have pioneered both transgenic technology and genome engineering in Platynereis, allowing us to interrogate cell types as well as individual genes on the functional level with unprecedented pre-cision. Furthermore, we combine transcriptomics, proteomics and quantitative mass spectrometry to dissect the logic of hormonal regulation. Finally, we make use of the remarkable transparency of Platy-nereis to observe cells and molecules in action. These approaches provide entry points into the fascinating biology of a new marine model species. Besides the action of hormones, we are actively in-vestigating the evolution of gene-regulatory logic and the regulation of stem cells in worms and sponges. Our vision is to firmly establish Platynereis as a reference species for marine biology.

Florian Raible

TEAMStephanie BannisterAndrej BelokurovMingliu DuAlessandra PoloRoger Revilla-i-DomingoAlexandra SchauerSven SchenkPoonam SharmaDidi-Andreas SongKarim Piyarali VadiwalaAgnė ValinčiūtėJuliane Zantke

Hormones orchestrate key switches in animal metabolism and physiology. We uncover main principles of hormone function in a novel invertebrate model system, Platynereis dumerilii.

SELECTED PUBLICATIONSBannister S, Antonova O, Polo A, Lohs C, Hallay N, Valinciute A, Raible F, Tessmar-Raible K. TALENs mediate efficient and heritable mutation of endogenous genes in the marine annelid Platynereis dumerilii. Genetics. 2014;197(1):77-89. PMID: 24653002 Backfisch B, Veedin Rajan VB, Fischer RM, Lohs C, Arboleda E, Tessmar-Raible K, Raible F. Stable transgenesis in the marine annelid Platynereis dumerilii sheds new light on photoreceptor evolution. Proc Natl Acad Sci USA. 2013;110(1):193-8. PMID: 23284166 Tessmar-Raible K, Raible F, Arboleda E. Another place, another timer: Marine species and the rhythms of life. Bioessays. 2011;33(3):165-72. PMID: 21254149

Ancestral-type hormones in a simple invertebrate. Individual hormone-producing cells are visualized (green color) in an adult Platynereis brain. Whereas cell bodies (“mc”) localize to the medial brain, neuronal projections (np) project into the region of the infracerebral gland, an annelid neurohemal organ.

Hormonal orchestration of regeneration and re-production by the medial Platynereis brain. (A) Classical transplantation results indicating the presence of an endocrine brain hormone inhibiting maturation and pro-moting regeneration. (B) Transplantation of smaller pieces (blue) refining the source of the hormone activity to the medial brain. For the first time, we can now dissect the involved machinery on a molecular and functional level.





43

J O H A N N R O T H E N E D E R

Cell cycle regulation and DNA damage response

Dysfunction of these mechanisms often results in the malignant transformation of a cell and the development of cancer. E2F is a family of transcription factors that integrate cell-cycle progression with transcription through cyclical interactions with important cell cycle regulators.

We have recently identified and characterized a protein that we called EAPP (E2F Associated PhosphoProtein). EAPP interacts with E2F1-3, com-prising the activator group of E2F proteins, and modulates E2F-dependent transcription. Tumour cells often overexpress EAPP, indicating that it confers a selective advantage to these cells. EAPP levels increase upon DNA damage and higher EAPP levels seem to protect cells from apoptosis. This protection can also be achieved by ectopic ex-pression of EAPP and correlates with an increased number of cells in G1 phase and an upregulation of p21. Increased p21 inhibits cyclin/cdk activity which is required for cell cycle progression, but has also interferes with apoptosis. The RNAi-mediated knock

down of p21 reduces the anti-apoptotic activities of overexpressed EAPP. This suggests that p21 at least in part mediates this activity of EAPP.

EAPP stimulates p21 expression by binding to its promoter and seems to be required for the assembly of the transcription initiation. The knock down of EAPP facilitates apoptosis and goes along with reduced p21.

Our findings suggest that EAPP is indispensable for the survival of a cell. The required amount of EAPP seems to depend on the environmental con-ditions. Preliminary evidence suggests that the role of EAPP in transcription is not limited to the p21 promoter. Active promoters are occupied by multiple types of complexes and EAPP seems to be an im-portant component of at least some of them. Lower-ing EAPP levels influences the expression of some of the genes examined including important cell-cycle regulators. In the future, we will examine which genes are influenced by EAPP and scrutinize the biochemical details of its activity.

Johann Rotheneder

My laboratory is focused on the mechanisms controlling growth and cell cycle of the mammalian cell. They respond to perturbations like replication errors or DNA damage by inducing cell cycle arrest, senescence, or apoptosis.

SELECTED PUBLICATIONSAndorfer P, Rotheneder H. EAPP: gatekeeper at the crossroad of apoptosis and p21-mediated cell-cycle arrest. Oncogene 2011;30(23):2679-90. PMID: 21258403 Schwarzmayr L, Andorfer P, Novy M, Rotheneder H. Regulation of the E2F-associated phosphoprotein promoter by GC-box binding proteins. Int J Biochem Cell Biol. 2008;40(12):2845-53. PMID: 18588995 Andorfer P, Schwarzmayr L, Rotheneder H. EAPP modulates the activity of p21 and Chk2. Cell Cycle. 2011;10(13):2077-82. PMID: 21572256

A model showing three different scenarios with elevated, normal and reduced levels of EAPP.





44

M A T T H I A S S C H Ä F E R

RNA modifications: their impact on gene expression and innate immunity

In contrast to the limited number of known DNA modifications (< 10), RNAs are “decorated” with (at least) 13 distinct post-transcriptional modifications (both at terminal and internal positions). The chemical nature of these modifications, especially in non-coding RNAs, can be diverse and very complex. However, the majority involves methylation reactions indicating the importance of methylated nucleotides for the function of RNA or DNA molecules. For instance, it is now well established that DNA is methylated in most organisms in form of 5-methylcytosine (m5C). This modification is also found in RNAs but its biological function is mostly unknown.

Importantly, studies of DNA modifications contributed greatly to the concept of epigenetic regulation of gene expression. Recent findings point towards a role for RNA, and possibly RNA mod-ifications, for the regulation of gene expression, which has set the stage for defining an exciting new concept: “RNA epigenetics”.

My group applies genetic and biochemical tools in the model organism Drosophila melanogaster to understand how m5C in RNA • Impacts on RNA stability and function• Controls stress-induced RNA processing• Affects the interaction with RNA-binding

proteins• Contributes to the regulation of gene

expression

In particular, we are interested in:• The characterization of m5C RNA methy-

lomes by using various methods to map this modification systematically and transcriptome- wide in different tissues and during various stress conditions. CRISPR-mediated genome editing is used to tag and manipulate the func-tion of various m5C RNA methyltransferases in an attempt to understand the nature of m5C RNA methylation systems in Drosophila;

• The impact of m5C RNA methylation on the innate immune response. We are using virus-infection paradigms to determine how m5C RNA methyltransferases affect anti-viral responses in Drosophila;

• The biological function of stress-induced tRNA fragments. Using inducible and tissue- specific expression of tRNA anticodon nucleases allows to study the cellular pathways that are affected by tRNA fragments, to measure tRNA fragment metabolism and to analyze their movement between different tissues and their inheritance into the next generation.

Our long-term goal is to characterize how RNA modifications contribute to genome re-gulation and, importantly, how they influence the reprogramming of gene expression under environmental impact such as stress conditions.

Matthias Schäfer

TEAMBianca GenenncherSophie Juliane VeiglDaniela Zinkl

RNAs carry post-transcriptional modifications. More than 130 distinct modifications are have been detected but their biological functions remain mostly elusive.

m5C RNA methyltransferase activity has been connected to RNA stability and to mobile element control. It is presently unclear how exactly m5C-methylated RNA impacts on these and other processes.

SELECTED PUBLICATIONSDurdevic Z, Mobin MB, Hanna K, Lyko F, Schaefer M. The RNA methyltransferase Dnmt2 is required for efficient Dicer-2-dependent siRNA pathway activity in Drosophila. Cell Rep. 2013;4(5):931-7. PMID: 24012760 Durdevic Z, Hanna K, Gold B, Pollex T, Cherry S, Lyko F, Schaefer M. Efficient RNA virus control in Drosophila requires the RNA methyltransferase Dnmt2. Embo Rep. 2013;14(3):269-75. PMID: 23370384 Schaefer M, Pollex T, Hanna K, Lyko F. RNA cytosine methylation analysis by bisulfite sequencing. Nucleic Acids Res. 2009;37(2):e12. PMID: 19059995


45

P E T E R S C H L Ö G E L H O F E R

Meiotic recombination

Our research efforts are well embedded in the Department of Chromosome Biology with five other groups performing meiosis research in various organisms.

Meiosis is a specialized, two-step cell division that ensures the reduction of the genome prior to the formation of generative cells. During meiosis, homologous centromers are segregated during the first, and sister centromers during the second division. As there is no intervening DNA replica-tion between the two meiotic divisions, each of the final division products contains only half of the initial DNA content. For a given diploid organism the developing generative cells are then haploid. It is important to note that during meiosis, genetic information between maternal and paternal chro-mosomes is mutually exchanged, leading to novel combinations of genetic traits in the following generation. Two genetically diverse generative cells fuse during the process of fertilization, re-estab-lish the species-specific original genome content

and constitute an individual with a unique genetic set-up.

Novel combinations between parts of paternal and maternal chromosomes are generated through the process of homologous recombination (HR). A pre-requisite for HR are DNA double strand breaks (DSBs), generated by a protein complex with the conserved protein SPO11 being its catalytically active subunit. DSBs are formed at non-random sites throughout the genome, known as hot spots of meiotic recombination. We are interested in 1) cis and trans acting factors that mediate meiotic DSB formation, 2) mechanisms of meiotic DSB process-ing, 3) the biochemical details of subsequent DSB repair and 4) the coordination of all these events. We use a broad range of techniques (molecular biology, cytology, biochemistry and genetics) and take advantage of the on-site facilities (BioOptics, deep- sequencing, mass spectrometry, bioinformatics). .

Peter Schlögelhofer

TEAMInes DittrichMichael JanisiwMarie-Therese KurzbauerZsuzsanna Orban-NemethKatja SchneiderKatharina SchroppJason SimsMona von HarderFaye Wheeler

We focus our research on meiotic recombination, mainly working with the model plant Arabidopsis thaliana and to some extent with the yeast Saccharomyces cerevisiae.

SELECTED PUBLICATIONSCabral G, Marques A, Schubert V, Pedrosa-Harand A, Schlögelhofer P. Chiasmatic and achiasmatic inverted meiosis of plants with holo-centric chromosomes. Nat Commun. 2014;5:5070. PMID: 25295686 Uanschou C, Ronceret A, Von Harder M, De Muyt A, Vezon D, Pereira L, Chelysheva L, Kobayashi W, Kurumizaka H, Schlögelhofer P, Grelon M. Sufficient Amounts of Functional HOP2/MND1 Complex Promote Interhomolog DNA Repair but Are Dispensable for Intersister DNA Repair during Meiosis in Arabidopsis. Plant Cell. 2013;25(12):4924-40. PMID: 24363313 Kurzbauer MT, Uanschou C, Chen D, Schlögelhofer P. The Recombinases DMC1 and RAD51 Are Functionally and Spatially Separated during Meiosis in Arabidopsis. Plant Cell. 2012;24(5):2058-70. PMID: 22589466

The panel shows a preparation of meiotic chromosomesisolated from meiocytes of a mutant Arabidopsis plant.The depicted stage of meiosis is called “pachytene” withall five chromosome pairs in close alignment, stabilized bya protein complex known as the “synaptonemal complex”(SC). To visualize the DNA and associated proteins aspecific DNA dye (DAPI) and antibodies (coupled tofluorescent molecules) specifically detecting certainmeiotic proteins have been applied. The DNA is stainedin blue, a protein of the SC is stained in green, and a DNArepair protein is stained in red.





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R E N É E S C H R O E D E R

RNA aptamers that regulate the transcriptome

To achieve this goal we adapted the classical SELEX procedure to be used in combination with genome sequences and deep sequencing. Genomic systematic evolution of ligands by exponential enrichment (SELEX) allows the isolation of protein binding RNAs independently of computational predictions and expression conditions.

We used genomic SELEX with an E. coli library to isolate RNA aptamers against RNA polymerase and the regulator protein Hfq. We further selected RNA polymerase II binding aptamers from the yeast and human genomes. These experiments delivered thousands of genomic RNA aptamers that regulate the activity of RNA polymerases. We named these regulatory RNAs RAPs for RNA polymerase aptamers or PBEs polymerase binding elements. RAPs and PBEs interact with the RNA polymerase and regulate the expression of the nascent transcripts.

We are analyzing the transcriptome of E. coli in depth to understand how Hfq and RNA aptamers fine tune gene expression.

Renée Schroeder

TEAMLucia AronicaIvana BilusicKrzysztof ChylinskiKasimir KienbeckMeghan LybeckerKatarzyna MatyllaMaximilian RadtkeDavid Fima Ruchman IndacocheaNatascha RzihaUrsula SchöberlNadia SedlyarovaJohanna StrannerAdam WeissLucie WeissMichael Thomas WolfingerRobert Paul Zimmermann

RNA is at the center of all steps of gene expression. Cells can be defined by their transcriptomes, not by their genomes. We are interested in discovering many regulatory elements that are part of the RNA regulon and in identifying their interacting partners and their targets.

SELECTED PUBLICATIONSMatylla-Kulinska K, Tafer H, Weiss A, Schroeder R. Functional repeat-derived RNAs often originate from retrotransposon-propagated ncRNAs. Wiley Interdiscip Rev RNA. 2014;5(5):591-600. PMID: 25045147 Bilusic I, Popitsch N, Rescheneder P, Schroeder R, Lybecker M. Revisiting the coding potential of the E. coli genome through Hfq co-immunoprecipitation. RNA Biol. 2014;11(5):641-54. PMID: 24922322 Lybecker M, Zimmermann B, Bilusic I, Tukhtubaeva N, Schroeder R. The double-stranded transcriptome of Escherichia coli. Proc Natl Acad Sci U S A. 2014;111(8):3134-9. PMID: 24453212

Overview of SELEX procedure. A genomic RNA library derived from DNA is submitted to several rounds of selection (b-e) and amplification (f-g,a) until a pool enriched in desired sequences is obtained. Deep sequencing of enriched pools allows the annotation of genomic aptamers to analyzed genomes.

RepRNAs: Repeat-derived RNAs often originate from retrotransposed non-coding. Upon retrotransposi-tion, ncRNAs are highly amplified, and as they spread throughout the genome, they diversify in sequence (depicted as bands of different shades of the same color). Some copies evolve new functions (depicted as a band with a changed color) giving rise to new classes of repRNAs. Therefore, non-coding transcripts derived from highly repetitive regions can be a rich reservoir for the evolution of novel functional RNAs( Matylla-Kulinska et al., WIREs RNA, 2014).





47

C H R I S T I A N S E I S E R

Chromatin modifiers in development and disease

Our group is specifically interested in the role of histone deacetylases (HDACs) in development and disease. HDAC inhibitor treatment of tumor cells leads to cell cycle arrest, differentiation or apoptosis. Therefore, HDACs are potential targets for anti-tumor drugs and several HDAC inhibitors are currently tested in clinical trials. HDACs are considered as transcriptional co-repressors and 18 members of the HDAC family have been identified in mammalian cells.

We have originally identified mouse HDAC1 as growth factor inducible gene in cytolytic T cells. In 2002, our lab published the first histone deacetylase knock-out mouse. HDAC1 gene disruption in mice leads to reduced proliferation and severe develop-mental problems resulting in embryonic lethality of HDAC1 knockout mice. In collaboration with Pat-rick Matthias, FMI Basel, we have generated mice with conditional Hdac1/Hdac2 alleles and published during the last years several studies on HDAC1/HDAC2 function in teratoma, brain and epidermis development. Together with Wilfried Ellmeier, Medical University of Vienna, we are studying the role of HDAC enzymes during T cell development. In

these studies we have identified HDAC1/HDAC2 as crucial regulators of cell proliferation, cell survival and differentiation.

In a second project, we examine the role of histone phosphorylation during the activation of mammalian genes by stress and growth factors. The presence of histone H3 phosphorylation marks at the regulatory regions of several mammalian genes correlates with the induced expression of dozens of target genes in mammalian cells. We have shown that 14-3-3 zeta can act as reader protein for S10- and S28-phosphorylated histone H3. We have identified the phosphatase PP2A as chromatin associated transcriptional repressor, which removes the active chromatin mark from specific target genes. Recently, we have started to analyze the genome wide presence of H3 phosphorylation marks and their impact on global gene expression in mammalian cells.

Christian Seiser

TEAMBrigitte GundackerAstrid HagelkruysChristina HumerTina MeischelVerena MoosMirjam MoserAnna Sawicka

DNA, the carrier of genetic information in our cells, is organized with the help of histone proteins as chromatin. Histone modifications affect the chromatin structure and thereby important biological processes such as transcription, replication and DNA repair.

SELECTED PUBLICATIONSSawicka A, Hartl D, Goiser M, Pusch O, Stocsits RT, Tamir I, Mechtler K, Seiser C. H3S28 phosphorylation is a hallmark of the transcriptional response to cellular stress. Genome Res. 2014;24(11):1808-20. PMID: 25135956 Hagelkruys A, Lagger S, Krahmer J, Leopoldi A, Artaker M, Pusch O, Zezula J, Weissmann S, Xie Y, Schöfer C, Schlederer M, Brosch G, Matthias P, Selfridge J, Lassmann H, Knoblich JA, Seiser C. A sin-gle allele of Hdac2 but not Hdac1 is sufficient for normal mouse brain development in the absence of its paralog. Development 2014; 141(3):604-16. PMID: 24449838 Winter M, Moser MA, Meunier D, Fischer C, Machat G, Mattes K, Lichtenberger BM, Brunmeir R, Weiss-mann S, Murko C, Humer C, Meischel T, Brosch G, Matthias P, Sibilia M, Seiser C. Divergent roles of HDAC1 and HDAC2 in the regulation of epidermal development and tumorigenesis. EMBO J. 2013;32(24):3176-91. PMID: 24240174

Deletion of HDAC1 and HDAC2 in the mouse brain leads to severe devel-opmental problems and embryonic lethality. Hematoxylin/eosin staining of representative paraffin sections from wildtype (A) and HDAC1/HDAC2-deficient (B) mice at embryonic day 18.5. Simultaneous loss of HDAC1 and HDAC2 results in reduced proliferation, increased apoptosis, DNA damage, impaired differentiation and severe cerebral hemorrhage.

Transient transcriptional derepression by histone H3 phosphorylation. In the absence of signals the gene is silent due to recruitment of HP1γ by local H3K9-dimethylation. Signal-dependent phosphorylation of H3S10 leads to dissociation of the HP1γ repressor, acetylation of H3K14 and binding of 14-3-3ζ. The resulting recruitment of RNA polymerase II and transcriptional activation occurs in the presence of the repressive H3K9me2 mark. Deacetylation, dephosphorylation and subsequent dissociation of 14-3-3ζ and binding of HP1γ reconstitute the repressed state of the promoter.





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T I M S K E R N

Interactions between viruses and cells

eIF4G is involved in recruiting capped cellular mRNA to the ribosomes for protein synthesis. Cleavage of this molecule during replication of the above mentioned viruses thus prevents capped cellular mRNA being translated. Viral protein synthesis is unaffected as it initiates internally downstream of the 5’ end of its RNA. We have determined the molecular structures of three of these proteinases and investigated the sites at which they interact with eIF4G. The leader proteinase is a relative of the plant cysteine proteinase papain. However, in contrast to papain, the leader proteinase is very specific, with only three target proteins identified at present. Nevertheless, a consensus sequence representing the cleavage site has been difficult to define as the three known cleavage sites show considerable differences. We have determined the structure of this proteinase in complex with an inhibitor to show the reasons why this proteinase is specific.

In addition, we have also used nuclear magnetic resonance to see how a short fragment of eIF4G is recognized by the proteinases.

Vaccinia virus is the viral vaccine that was used to eradicate smallpox virus. Both vaccinia virus and smallpox virus encode many proteins that re-duce the affectivity of the host immune system. One such protein is vaccinia virus A46, a protein that counteracts several immune regulators of the infected cell to prevent the inflammation re-sponse. We have determined the three-dimensional structure of residues 87to 229 of A46 and in-vestigated the interaction of this protein with purified host cell proteins. The structure suggests how the A46 specifically interacts with its targets in the host cell and why its spectrum of interaction partners is different to those of closely related vaccina virus proteins. Further investigation should help to illuminate how the cellular immune regulators interact with each other.

Tim Skern

TEAMMartina Aumayr Sofiya FedosyukKatharina RadakovicsJutta Steinberger

Most viruses interfere with or modulate host systems to ensure successful replication. My group looks at interactions between the leader proteinase of foot-and-mouth disease virus and the 2A proteinase of the common cold virus with the cellular protein called eukaryotic initiation factor 4G (eIF4G) as well as investigating the immunemodulator protein A46 of vaccinia virus.

SELECTED PUBLICATIONSFedosyuk S, Grishkovskaya I, De Almeida Ribeiro E, Skern T. Characterisation and structure of the vaccinia virus NF- κB antagonist A46. J Biol Chem. 2014;289(6):3749-62. PMID: 24356965 Steinberger J, Kontaxis G, Rancan C, Skern T. Comparison of self-processing of foot-and-mouth disease virus leader proteinase and porcine reproductive and respiratory syndrome virus leader proteinase nsp1α. Virology 2013;443(2):271-7. PMID: 23756127 Steinberger J, Grishkovskaya I, Cencic R, Juliano L, Juliano MA, Skern T. Foot-and-mouth disease virus leader proteinase: Structural insights into the mechanism of intermolecular cleavage. Virology. 2014;468-470:397-408. PMID: 25240326

The foot-and-mouth disease leader proteinase. Structure of A46 protein from vaccinia virus.





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D E A S L A D E

DNA damage response

The dynamics of the DDR protein network is gov-erned by posttranslational modifications including phosphorylation, methylation, acetylation, ubiq-uitination, sumoylation and poly(ADP-ribosyl)ation (PARylation). They regulate the recruitment of DNA repair factors to the sites of DNA damage, their enzymatic activity, interactions and the choice of the DNA repair pathway. Our research focuses on the regulation of DDR by reversible PARylation and acetylation modifications.

Poly(ADP-ribose) (PAR) is rapidly produced in re-sponse to DNA damage by PARP polymerases and elicits the recruitment of different DDR factors to DNA damage sites. PAR is rapidly degraded by the PARG glycosylase to ensure a transient effect. While PAR is the largest posttranslational protein modifi-cation, acetylation is the most prevalent. Among numerous deacetylases, in recent years sirtuins (SIRTs) have emerged as crucial regulators of gene expression, metabolism and genome integrity that interconnect with the PAR processes. As NAD-con-suming enzymes, PARPs and sirtuins are activated in conditions of genotoxic, oxidative, metabolic and inflammatory stress, which alter NAD levels. They regulate cellular response to stress through changes in chromatin structure, gene expression, DNA repair, cell cycle, metabolism and cell fate.

Among 17 PARP family members and seven sirtuins in mammals, PARP1, PARP2, PARP3, SIRT1, and SIRT6 are hitherto known to affect DDR. PARP1/2 and SIRT1 deficiencies sensitize the cells to DNA-damaging agents and result in embryonic lethality due to genomic instability. However, little is known about DNA repair processes regulated by sirtuins and the relationship between sirtuin- and PARP-mediated modifications of DNA repair proteins. The observations that PARPs and sirtuins

regulate each other’s levels and activities and have opposite effects on the same pathway such as cell death suggest a functional interplay between these NAD-consuming enzyme families.

Our aim is to identify unknown DDR protein targets regulated by PARPs and sirtuins and characterize their functional interplay by using biochemical and cell biological techniques in mammalian systems.

DNA damage response (DDR) is a complex regulatory network that involves DNA damage sensing, signaling and repair. These processes are carried out by diverse enzymatic activities that must be precisely coordinated as to ensure the efficient, accurate and timely repair of DNA damage and the preservation of genomic integrity.

SELECTED PUBLICATIONSSlade D, Dunstan MS, Barkauskaite E, Weston R, Lafite P, Dixon N, Ahel M, Leys D, Ahel I. The structure and catalytic mechanism of a poly(ADP-ribose) glycohydrolase. Nature 2011;477(7366):616-20. PMID: 21892188 Chen D, Vollmar M, Rossi MN, Phillips C, Kraehenbuehl R, Slade D, Mehrotra PV, von Delft F, Crosthwaite SK, Gileadi O, Denu JM, Ahel I. Identification of macrodomain proteins as novel O-acetyl-ADP-ribose deacetylases. J Biol Chem. 2011;286(15):13261-71. PMID: 21257746 Slade D, Radman M. Oxidative stress resistance in Deinococcus radiodurans. Microbiol Mol Biol Rev. 2011;75(1):133-91. PMID: 21372322

PARPs and sirtuins regulate stress response after DNA damage.

PARPs and sirtuins modify proteins: PARPs synthesize poly(ADP-ribose) , while sirtuins remove acetyl groups.

Dea Slade

TEAMAndreas BrachnerMelania BrunoTanja KaufmannSebastian KostrhonEva Kukolj





50

K R I S T I N T E S S M A R - R A I B L E

Lunar periodicity and inner brain photoreceptors

The function of vertebrate inner brain opsins - Nature’s own optogenetics?

Starting with experiments at the beginning of the 20th century it has become apparent that light can be perceived by cells in the inner brain of vertebrates, independent of eyes and pineal organs. Subsequent studies established that measurable amounts of light can penetrate deep inside the brain of vertebrates, including mammals.

Our study of non-visual photoreceptors in fishes recently revealed that vertebrate brains harbor directly light-sensory motor- and interneurons. This led us to hypothesize that environmental light impacts on the information processing and output in the vertebrate brain by modulating the membrane potential of Opsin-expressing inter- and motorneurons, and hence leads to light- dependent behavioral alterations. We now test this hypothesis.

Molecular neurobiology of a moonlight entrained circalunar clock

While the function of vertebrate non-visual opsins is likely connected to solar light perception, lunar light is also a strong environmental stimulus for animals. The lunar cycle synchronizes reproduc-tive behavior and sexual maturation of animals as diverse as corals, midges, polychaetes and fishes. In animals such as the annelid Platynereis dumerilii or the midge Clunio marinus, dim nocturnal light serves as entrainment cue for an endogenous oscillator – a circalunar clock – that orchestrates reproductive and behavioral cycles. As circalunar clocks run with a (semi-)monthly period, they represent fundamental biological phenomena clearly distinct from the widely studied, solar light-entrained circadian (24h) clocks. Jointly with Florian Raible’s group we established sev-eral critical functional tools for Platynereis du-

merilii, including stable transgenesis and TALEN-mediated targeted genome muta-genesis. Using Platynereis dumerilii, my group has started to obtain first insights into the circalunar clock and its interac-tions with the circadian clock.

Differening from Platynereis dumeri-lii (whose worm ancestors never left the oceans), the midge Clunio marinus likely acquired its circalunar clock de novo during the past 20.000 years. Strains of different geographic origins exhibit dif-ferences in their circalunar and circadian timing (“time races”). Clunio marinus thus represents an ideal model to understand the evolution of circalunar clocks across species (in comparison to Platynereis du-merilii), but also across the population of its own species (“time races”).

Kristin Tessmar-Raible

TEAMOlga AntonovaThomas AyersMarcus DekensBruno FontinhaStefan HajnyMaximilian HofbauerVerena HöllbacherJoanna Jagoda Sandra PflüglerBirgit PoehnMirta ResetarPrabhavathi Talloji Vinoth Babu Veedin Rajan

The main interest of my lab is to investigate how solar and lunar light are sensed by the nervous system and how this light information impacts on the animals’ information processing and endogenous clocks.

SELECTED PUBLICATIONSZantke J, Ishikawa T, Arboleda E, Lohs C, Schipany K, Hallay N, Straw A, Todo T, Tessmar-Raible K. Circadian and circalunar clock interactions in a marine annelid. Cell Rep. 2013;5:1-15. PMID: 24075994 Fischer RM, Fontinha BM, Kirchmaier S, Steger J, Bloch S , Inoue D, Panda S, Rumpel S, Tessmar-Raible K. Co-Expression of VAL- and TMT-Opsins Uncovers Ancient Photosensory Interneurons and Motorneurons in the Vertebrate Brain. PLoS Biology. 2013;11(6):e1001585. PMID: 23776409 Raible F, Tessmar-Raible K. Platynereis dumerilii. Curr Biol. 2014;24(15):R676-7. PMID: 25093553

Lunar reproductive periodicity of Platynereis dumerilii is synchronized by light and controlled by a clock mechanism.





51

A L I P A S H A V A Z I R I

Dynamics of coupled biological systems: methods and phenomena

The focus of the Vaziri lab lies at the intersection between physics, neuroscience and information theory. We are interested in understanding how the information processing capabilities of the brain emerges from the dynamic interaction of the neuronal networks. Ultimately, our aim is discover the computational algorithms underlying object recognition, generalization, learning and decision- making. Addressing these questions has been hampered by lack of appropriate tools and methods that allow parallel and spatiotemporally specific application of excitation patterns onto neuronal populations while capturing the dynamic activity of the entire network at high spatial and temporal resolution.

Taking a multidisciplinary and reverse engineering approach we develop and apply new techniques aimed at addressing the above questions. Over the last few years we have developed two different new high-speed calcium imaging techniques that allow for the first time to capture the dynamic of neuronal network at the single cell resolution in vivo and on the level of whole brains at up to tens of Hertz for small model organism such as the nematode C. elegans and zebrafish larva. This provides for the first time the opportunity to capture the flow

of information from the primary sensory neurons across different stages of representation, to their processing and interaction with internal brain states for generating behavior in real time.

One approach relies on “sculpting” the excitation volumes in biological samples using non-linear op-tics and the other relies on light-field imaging, a tomography type approach for simultaneous readout of neuronal activity in 3-D. Using these techniques we have recently shown brain-wide functional imaging of entire nervous systems at single cell resolution. Further, we demonstrate intrinsically simultaneous volumetric Ca-imaging in the entire brain of larval zebrafish during sensory stimulation. We are able to track the activity of 5000 neurons distributed throughout the brain at 20Hz volume rate. The simplicity of this technique and the possibility of the integration into conven-tional microscopes make it an attractive tool for high-speed volumetric functional-imaging. These tools combined with high speed optogenetic control of neuronal circuits, advanced statistics tools and mathematical modeling will be crucial to move from an anatomical wiring map towards a dynamic map of neuronal circuits.

How do the dynamics of coupled biological systems confined by their structure lead to function? While we investigate this question in different systems and at different scales, a major goal is to understand how sensory inputs are represented across brain hierarchies and how their processing generates innate and learnt motor output. To answer these questions we have a major focus on development of optical technologies for high-speed, single cell specific and brain-wide modulation and functional imaging of neuronal circuits.

SELECTED PUBLICATIONSPrevedel R, Yoon YG, Hoffmann M, Pak N, Wetzstein G, Kato S, Schrödel T, Raskar R, Zimmer M, Boyden ES, Vaziri A. Simultaneous whole-animal 3D imaging of neuronal activity using light-field microscopy. Nat Meth. 2014;11(7):727-30. PMID: 24836920 Schrödel T, Prevedel R. Aumayr K, Zimmer M, Vaziri A. Brain-wide 3D imaging of neuronal activity in Caenorhabditis elegans with sculpted light. Nat Meth. 2013;10(10):1013-20. PMID: 24013820 Andrasfalvy B, Zemelman BV, Vaziri A. Two-photon Single Cell Optogenetic Control of Neuronal Activity by Sculpted Light. Proc Natl Acad Sci U S A. 2010;107(26):11981-6. PMID: 20543137

High-speed whole-brain calcium imaging in larval zebrafish via light-field deconvolution microscopy. (a) Maximum-intensity pro-jection (MIP) of a light-field deconvolved volume, (b) Extracted Ca2+ intensity signal (∆F/F 0) of GCaMP5 fluo-rescence using spatial filters shown in a. Each row shows a time-series heat map. Color bars denote encircled regions in b, which include the olfac-tory epithelium, olfactory bulb and telencephalon. The arrow at 15 s denotes the addition of an aversive odor. A close-up of the dashed box is shown (right, lower panel).

Alipasha Vaziri

TEAMDavid CisnerosMartin FrankChristoph GoetzMagdalena HelmreichMaxim MolodtsovTobias NöbauerRobert PrevedelFriederike SchlummStephanie Walcher





52

G I J S V E R S T E E G

Ubiquitin-mediated regulation of immune signaling

Precise regulation and fine-tuning of immune signaling pathways is critical to strike the right bal-ance between conferring sufficient antimicrobial activity during infection and preventing hyper-im-mune activation resulting in auto-immunity. The molecular mechanisms regulating these signaling molecules in different cell types during the innate immune response remain relatively poorly defined.

The post-translational modifier ubiquitin is essential for both positive and negative immune regulation. Ubiquitin can be covalently attached to target molecules by so-called E3 ligases, after which the properties of these targets are dramati-cally changed. An important family of E3 ligases is formed by the 75-member tri-partite motif (TRIM) proteins.

We recently demonstrated that half of the 75-member family of human TRIM ubiquitin E3 ligases positively regulates innate immune cytokine expression. In addition, preliminary data implicate several other TRIM proteins as negative regulators. Our data show that individual TRIMs act at various different stages of immune signaling, suggesting that many of them act on different target mol-ecules.

My lab focuses on identifying the molecular mechanisms through which TRIM E3 ligases act to balance innate immune cytokine responses us-ing biochemical, proteomic and cell biology ap-proaches. This will be done at a TRIM family-wide scale, after which the significance of individual molecules will be assessed in reconstituted in vitro models, relevant primary human cells (e.g. macro-phages, dendritic cells) and ultimately small animal models.

Gijs Versteeg

TEAMStefan BenkeThomas HahnMartin Stöger

We are interested in understanding how the immune system is regulated during infection and inflammatory disease.

SELECTED PUBLICATIONSRajsbaum R, Versteeg GA, Schmid S, Maestre AM, Belicha-Villanueva A, Martínez-Romero C, Patel JR, Morrison J, Pisanelli G, Miorin L, Lau-rent-Rolle M, Moulton HM, Stein DA, Fernandez-Sesma A, tenOever BR, García-Sastre A. Unanchored K48-linked polyubiquitin synthesized by the E3-ubiquitin ligase TRIM6 stimulates the interferon-IKKε kinase-mediated antiviral response. Immunity 2014;40(6):880-95. PMID: 24882218 Versteeg GA, Rajsbaum R, Sánchez-Aparicio MT, Maestre AM, Valdiviezo J, Shi M, Inn KS, Fernandez-Sesma A, Jung J, García-Sastre A. The E3-ligase TRIM family of proteins regulates signaling pathways triggered by innate immune pattern-recognition receptors. Immunity 2013;38(2):384-98. PMID: 23438823 Mata MA, Satterly N, Versteeg GA, Frantz D, Wei S, Williams N, Schmolke M, Peña-Llopis S, Brugarolas J, Forst CV, White MA, García-Sastre A, Roth MG, Fontoura BM. Chemical inhibition of RNA viruses reveals REDD1 as a host defense factor. Nat Chem Biol. 2011;7(10):712-9. PMID: 21909097

Balanced immune regulation by ubiquitin is critical for preventing insufficient immune activation or autoimmunity.





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A R N D T V O N H A E S E L E R

Bioinformatics

Besides the collaborative efforts, the CIBIV pur-sues its own research agenda. The group’s main effort is to understand the evolutionary processes that have shaped the genomes of contemporary species. To this end, the CIBIV applies methods from statistics, computer sciences, and mathematics to detect the traces ancient evolutionary events have left in modern genomes. With the advancement of sequencing technologies, inference of evolutionary history faces new computational challenges. Consequently we have developed more efficient tools for phylogenomics, one of them speeds up the bootstrap procedure considerably. We are also involved in data analysis of phylogenomics data (see figure below).

More recently we expanded our research interests to address mathematically and computation-ally tractable problems that may help to assist

in conservation decisions. We have employed the integer linear programming paradigm to explore conservation scenarios in the presence of external constraints.

Finally, we develop tools to efficiently analyze deep sequencing data that pose a new challenge to Bioinformatics. To this end we have developed an efficient optimal local alignment tool, which maps millions of reads to a reference genome in a few seconds. The mapping of reads to a reference genome is the first, and possibly crucial step for any further analysis. We also aim to understand the sampling properties of RNA-seq approaches. Recently, we have shown that the Pitman-sampling formula allows statements about the coverage.

The development of efficient algorithms and further statistical tools to analyze the data will be a major research focus of the CIBIV during the next years.

The Center for Integrative Bioinformatics Vienna (CIBIV, www.cibiv.at) serves as a central facility to coordinate the Bioinformatics activities at the MFPL. Moreover it provides infrastructure and Bioinformatics expertise for the various research groups at MFPL and on campus

SELECTED PUBLICATIONSMinh BQ, Nguyen MA, von Haeseler A. Ultra-Fast Approximation for Phylogenetic Bootstrap. Mol Biol Evol. 2013;30(5):1188-95. PMID: 23418397 Sedlazeck FJ, Rescheneder P, von Haeseler A. NextGenMap: Fast and accurate read mapping in highly polymorphic genomes. Bioinformatics. 2013;29(21):2790-1. PMID: 23975764 Tauber S, von Haeseler A. Exploring the sampling universe of RNA-seq. Stat Appl Genet Mol Biol. 2013;12(2):175-88. PMID: 23629158

The phylogenetic profiles of the yeast ribosome biogenesis factors and their Pfam domains (Ebersberger et al., Nucleic Acids Res., 2014).

Arndt von Haeseler

TEAMQuang Minh BuiOlga ChernomorMaurits EversMiguel GallachIris GruberChristian HarlanderTobias KaiserMilica KrunicKonstantina KyriakouliLam Tung NguyenLuis Felipe Paulin PazSusanne PfeiferNiko PopitschCeline PrakashPhilipp ReschenederHeiko SchmidtFritz SedlazeckMoritz SmolkaDavid SzkibaPeter Venhuizen

http://www.cibiv.at





54

C H R I S T I N A W A L D S I C H

Exploring RNA folding: from structure to function

Although RNAs are highly diverse and fulfill very different tasks, they share their strict dependence on acquiring a specific 3-D architecture to be functional. Our research focuses on understand-ing this most essential aspect of RNA function by investigating RNA structure and folding pathways. Specifically, we aim to provide novel insights into protein-facilitated RNA folding and how RNAs fold in the living cell. Little is known about how RNA folds in vivo and how they interact with their tar-gets despite RNA’s importance for cell viability. Therefore, it is of fundamental importance to gain insights into the forces driving RNA folding in vivo and to establish the contribution and impact of the cellular environment.

Catalytic RNAs, in particular group II introns, are the best-suited model system to study RNA folding in the living cell, as their structure and folding pathways are well characterized in vitro and formation of the native conformation can be measured as a function of catalysis. Therefore, we investigate the intracellular folding pathway of

the Sc. ai5g group II intron. Importantly, Sc. ai5g and other yeast mitochondrial introns depend on trans-acting protein factors for efficient splicing in vivo. Consequently, we are interested in explor-ing how these proteins shape folding of its target RNAs. This allows us to derive principles govern-ing in vivo RNA folding facilitated by proteins and other cellular factors.

We are also fascinated by telomerase; an RNP that has received considerable attention because of its significant up-regulation in the majority of cancer cells and its role in preventing chromosomal instability and senescence as well as in inherited human disorders. We are interested in exploring the structure of telomerase RNA and in studying the interplay of RNA folding and RNP assembly. Ultimately, deciphering the rules governing RNA folding will advance our understanding of the ba-sic mechanism of RNA-dependent processes, like self-splicing and telomere addition, and the role of RNA in disease.

Christina Waldsich

TEAMStefan HandlVerena HeisigElzbieta KowalskaAndreas LiebegNora SachsenmaierMichael WildauerGeorgeta Zemora

RNAs regulate biology: In the past years it has become increasingly evident that RNA is the driving force in most cellular processes.

SELECTED PUBLICATIONSCruz JA, Blanchet MF, Boniecki M, Bujnicki JM, Chen SJ, Cao S, Das R, Ding F, Dokholyan NV, Flores SC, Huang L, Lavender CA, Lisi V, Major F, Mikolajczak K, Patel DJ, Philips A, Puton T, Santalucia J, Sijenyi F, Hermann T, Rother K, Rother M, Serganov A, Skorupski M, Soltysinski T, Sripakdeevong P, Tuszynska I, Weeks KM, Waldsich C, Wildauer M, Leontis NB, Westhof E. RNA-Puzzles: a CASP-like evaluation of RNA three-dimensional structure prediction. RNA. 2012;18(4):610-25. PMID: 22361291 Sachsenmaier N, Waldsich C. Mss116p: A DEAD-box protein facilitates RNA folding. RNA Biol. 2013;10(1):71-82. PMID: 23064153 Tian N, Yang Y, Sachsenmaier N, Muggenhumer D, Bi J, Waldsich C, Jantsch MF, Jin Y. A structural determinant required for RNA editing. Nucleic Acids Res. 2011;39(13):5669-81. PMID: 21427087

The folding pathway of the Sc. ai5γ group II ribozyme. In the unfolded state only the secondary structure is formed, while in case of the intermediate state Domain 1 (blue) compacts and forms a tertiary structure thereby providing the scaffold for docking of Domains 3 (green) and 5 (red), which completes folding to the native conformation (Pyle, Fedorova and Waldsich, TiBS, 2007).





55

G R A H A M W A R R E N

Golgi biogenesis

For membrane-bound organelles, such as the Golgi, the mechanism underlying this duplication and segregation remains controversial. The primary aim of our research is to understand how the cell copies the complex architecture of the Golgi during the cell cycle and partitions one copy to each of the two daughter cells

The Golgi lies at the heart of the secretory path-way, modifying and sorting the transiting cargo. Despite a deep mechanistic understanding of the underlying mechanisms of selective cargo transport we know much less about the process of biogenesis, which duplicates and partitions the Golgi once per cell cycle.

The large number of Golgi stacks in mammalian cells has made it difficult to address mechanistic

questions about Golgi biogenesis. To circumvent this problem, we exploit the simplicity and genetic tractability of the protozoan parasite Trypanosoma brucei, the causative agent of sleeping sickness. This organism has a single Golgi, the duplication of which can be tracked using (photoactivatable) fluorescent proteins and time-lapse fluorescence microscopy. Using this approach, we investigated whether the existing Golgi contributes to the forma-tion of the new Golgi. We showed that components of the existing Golgi can indeed be tracked to the newly-forming Golgi. In order to manipulate the system more directly, we have established a perme-abilized cell assay using T. brucei cells and we are currently trying to determine the factors essential for this process.

During normal growth and division, cells double in mass and divide into two equally-sized daughters. All cell constituents must be duplicated and then segregated equally during mitosis.

SELECTED PUBLICATIONSMorriswood B, Havlicek K, Demmel L, Yavuz S, Sealey-Cardona M, Vidilaseris K, Anrather D, Kostan J, Djinovic-Carugo K, Roux K, Warren G. Novel bilobe components in Trypanosoma brucei identified using proximity-dependent biotinylation. Eukaryot Cell. 2013;12(2):356-67. PMID: 23264645 Misteli T, Warren G. 25 Years of Current Opinion in Cell Biology. Curr Opin Cell Biol. 2013;25(1):1-2. PMID: 23352256 Esson HJ, Morriswood B, Yavuz S, Vidilaseris K, Dong G, Warren G. Morphology of the trypanosome bilobe, a novel cytoskeletal structure. Eukaryot Cell. 2013;11(6):761-772. PMID: 22327007

Trypanosoma brucei, stably expressing two Golgi markers: GRASP-mCherry (the reference marker) and GntB-PAGFP, the retention domain of a resident Golgi enzyme coupled to photo-activatable GFP. By photo-activating the region containing the old Golgi (open arrow) fluorescence is seen over time to appear in the region containing the new Golgi (closed arrow), suggesting that material from the old Golgi is used to construct the new.

Graham Warren

TEAMEmine Sevil DavidsonKyojiro IkedaMartina MitterhuberBrooke MorriswoodIsabelle Walters





56

G E O R G W E I T Z E R

Somatic stem cells of the heart

Stem cells and progenitor cells are supposed to exist in niches, where they remain in an undifferentiated and quasi-dormant state until external signals stimulate commitment and differentiation to specific somatic cells which may contribute to the repair or maintenance of homeostasis of an organ. Mimicking a stem cell niche of the heart in vitro, we succeeded in the isolation of somatic stem cells from postnatal murine hearts and could maintain these cells as monoclonal self-renewing cells lines expressing Oct4, Sox2 and Nanog for several years.

These cells obviously committed to the meso- dermal lineage and expressing early myocardial transcription factors Brachyury, Nkx2.5, GATA4, and Isl1 exclusively differentiate to cardiomyocytes, smooth muscle cells, and vascular endothelial cells and thus were named cardiovascular progenitor cells (CVPCs).

Cardiomyogenic progenitors further differentiate to equal numbers of functional pacemakers, atrial and ventricular cardiomyocytes with a near-adult action potential. Stimulation of CVPCs with Activin A and Retinoic Acid did not yield any cell types of the endodermal and ectodermal lineage, respectively. Addition of BMP2 and SPARC promoted cardiomyogenesis and led to the upregulation of genes for the mesoderm specific transcription factor Brachyury and the early myocardial transcription factor Nkx2.5.

In our ongoing research, we try to reveal the molecular pathways which allow SPARC, BMP2 and Nodal to activate Brachyury and Nkx2.5 expression in CVPCs and how Brachyury, Nanog and Nkx2.5 interact on the transcriptional level in undifferentiated and differentiating CVPCs. Our long term scientific goal is to understand early cardiomyogenesis and how somatic stem cells may contribute to homeostasis of the heart.

Understanding the molecular and cellular interplay regulating stem cell self-renewal and differentiation may contribute to future targeted therapies utilizing growth factors or small molecules for the temporal endogenous activation of the stem cell pool.

Georg Weitzer

TEAMSabrina BailMario IvankovicJulia Klammer Marie-Thérèse Konrad

In recent years, numerous groups provided compelling evidence for the existence of somatic stem cells in the heart of different mammalian species.

SELECTED PUBLICATIONSHoebaus J, Heher P, Gottschamel T, Scheinast M, Auner H, Walder D, Wiedner M, Taubenschmid J, Miksch M, Sauer T, Schultheis M, Kuzmenkin A, Seiser C, Hescheler J, Weitzer G. Embryonic Stem Cells Facilitate the Isolation of Persistent Clonal Cardiovascular Progenitor Cell Lines and Leukemia Inhibitor Factor Maintains Their Self-Renewal and Myocardial Differentiation Potential in vitro. Cells Tissues Organs. 2013;197(4):249-68. PMID: 23343517 Fuchs C, Scheinast M, Pasteiner W, Lagger S, Hofner M, Hoellrigl A, Schultheis M, Weitzer G. Self-Organization Phenomena in Embryonic Stem Cell-Derived Embryoid Bodies: Axis Formation and Breaking of Symmetry during Cardiomyogenesis. Cells Tissues Organs 2012;195(5):377-91. PMID: 21860211 Taubenschmid J, Weitzer G. Mechanisms of cardiogenesis in cardiovascular progenitor cells. Int Rev Cell Mol Biol. 2012;293:195-267. PMID: 22251563

Localization of Oct4 protein during cell division of CVPCs. Immunofluorescence microscopy of CVPCs with Oct4 antibodies (green), and DAPI (blue). Bar: 15 μm.Arrows, top, Metaphase; middle, Anaphase, and bottom, Telophase. Asterisks, Oct4 negative nucleus of a SNL76/7 feeder cell.





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G E R H A R D W I C H E

Cytolinker proteins in signaling and disease

We are interested in cytoskeletal linker proteins (cytolinkers), a family of multi-modular, highly versatile proteins of exceptional size, that by networking and anchoring intermediate (10 nm) filaments (IFs) regulate the dynamics and archi-tecture of the cytoskeleton. We are studying the role of cytolinkers in normal development, cellular stress response and disease, combining mouse genetics with cell and structural biology.

Several years ago we discovered plectin, a ubiquitous cytolinker that became the prototype of what meanwhile is a whole family of similar proteins. Plectin has key functions in shaping cell architecture, mechanical stabilization, po-larization, and migration of cells, positioning of organelles, signal transduction, nerve conduction, and others. Thus, loss or dysfunction of plectin leads to diseases affecting a variety of cell types and tissues. Plectin’s versatility is based on an unusual diversity of isoforms distinguished by short sequences that determine the differential targeting of IFs within cells. We have generated a panel of transgenic mouse lines, including full knockout (KO), and over a dozen single isoform and conditional/tissue-restricted KOs, and knock-in lines. This unique repertoire of genetically distinct mouse lines serves us and our collaborators around the world as animal models for a variety of plectin-related human diseases and source of cell systems for basic research topics with focus on:

Myofibrillar myopathiesthe hallmarks of which are formation of protein

aggregates in skeletal muscle, mitochondrial dys-function, and disturbed heart function;Skin blistering disease (EBS)

due to lack of hemidesmosome stabilization;Neuropathies and memory loss

caused by deregulated microtubule dynamics, reduced nerve cell branching and nerve conduction, as well as unbalanced metabolism; Mechanosensing and -transduction

with fibroblasts, keratinocytes, endothelia, cardiac/skeletal muscle used as model systems;Cancer

mechanisms of plectin-mediated metastasis in pancreatic and urinary bladder cancer;Gene therapy, disease treatment

utilization of mini-genes in mouse models, application of chemical chaperons in clinical trials.

The cytoskeleton provides the structural basis for physical robustness, shape, movement, and intracellular dynamics of eukaryotic cells.

SELECTED PUBLICATIONSWinter L, Staszewska I, Mihailovska E, Fischer I, Goldmann WH, Schröder R, Wiche G. Chemical chaperone ameliorates pathological protein aggregation in plectin-deficient muscle. J Clin Invest. 2014;124, 1144-1157. PMID: 24487589 Wiche G, Osmanagic-Myers S, Castañón MJ. Networking and anchoring through plectin: a key to IF functionality and mechanotransduction. Curr Opin Cell Biol. 2014;32C:21-29. PMID: 25460778 Shin SJ, Smith JA, Rezniczek GA, Pan S, Chen R, Brentnall TA, Wiche G, Kelly KA. Unexpected gain of function for the scaffolding protein, plectin, due to mislocation in pancreatic cancer. Proc Natl Acad Sci U S A. 2013;110(48):19414-9. PMID: 24218614

Images represent top (left) and inside (right) views of a neuromuscular synapse triple-labeled for desmin-filaments (red), the nerve terminal marker synaptophysin (blue), and AChRs (green). (MBoC cover, Dec 15, 2014)

Gerhard Wiche

TEAMIrmgard FischerEva MihailovskaIlona StaszewskaMaria J Wiche-Castanon

High-resolution immunofluorescence microscopy images of keratinocyte cytoskeleton visualizing IF network- and microtubule-associated cytolinker proteins (plectin and MAP2). (Histochem Cell Biol. cover, July, 2013)





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A N G E L A W I T T E

φCh1, a model for gene regulation in haloalkaliphilic archaea

This organism has an optimal growth at 3.5M NaCl and at a pH of 9.5. The virus itself is used as a model system to analyze gene expression in haloalkaliphilic organisms, facing with two extremes: a high pH and high concentrations of salt. The sequence of φCh1, infecting the haloalkaliphilic ar-chaeon Natrialba magadii, contains an open read-ing frame (int1) in the central part of its genome that belongs to the λ integrase family of site-spe-cific recombinases. Sequence similarities to known integrases include the highly conserved tetrad R-H-R-Y. The flanking sequences of int1 contain several direct repeats of 30 bp in length (IR-L and IR-R), which are orientated in an inverted direction. The invertible region encodes two structural pro-teins (gp34 and gp36, encoded by ORF34 and ORF36) expected to represent the viral tail fibre proteins.

In vitro experiments using purified protein variants gp341 and gp3452 (containing the C-terminus of gp36) revealed exclusive binding of gp3452 but not of gp341 to cells of the cured strain Nab. magadii L13. This specific binding could be inhibited by the addition of α-D-galactose. α-D-galactose also sig-nificantly reduced the infectivity of φCh1. Binding experiments employing distinct domains of gp341 and gp3452 indicated the C-terminus to be respon-sible for binding to the receptor on the cell surface of Nab. magadii L13. This C-terminus contains a domain with similarities to the super-family of “galactose-like binding” proteins. In summary, the experiments gave evidence that gp3452 represents the anti-receptor of φCh1 that binds to specific carbohydrate ligands located on the cell surface of Nab. magadii.

Currently the work concentrates on the identification and function of repressor and activator molecules encoded by the virus, gene regulation

due to a recombination event, identification of the receptor for the virus on the cell surface of Nab. magadii and the transformation/shuttle vector system developed by the group. In addition, the method is used to construct different mutants.

Angela Witte

TEAMTina GrohmannChristoph HofbauerAgnes KoglerKathrin Schönfelder

The virus φCh1 was found by spontaneous lysis of a culture of the haloalkaliphilic archaeon, Natrialba magadii, an isolate from the soda lake, Lake Magadii in Kenya.

SELECTED PUBLICATIONSMayrhofer-Iro M, Ladurner A, Meissner C, Derntl C.,Reiter M, Haider F, Dimmel, K. Rössler N, Klein R, Baranyi U, Scholz H, Witte A. Utilization of virus φCh1 elements to establish a shuttle vector system for halo(alkali)philic Archaea via transformation of Natrialba magadii. Appl Environ Microbial. 2013;79(8):2741-2748. PMID: 23416999 Klein R, Rössler N, Iro M, Scholz H, Witte A. Haloarchaeal myovirus φCh1 harbours a phase variation system for the production of protein variants with distinct cell surface adhesion specificities. Mol Microbiol. 2012;83(1):137-50. PMID: 22111759 Iro M, Klein R, Gálos B, Baranyi U, Rössler N, Witte A. The lysogenic region of virus phiCh1: identification of a repressor-operator system and determination of its activity in halophilic Archaea. Extremophiles. 2007;11(2):383-96. PMID: 17123129

Electron micrograph of φCh1 particle negatively stained with uranylacetate.





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I V A N Y U D U S H K I N

Functional imaging of signaling networks

Although many signaling molecules have been identified, how exactly the dynamics of their inter-actions ensure the ultimate specificity and adaptive character of cellular responses remains poorly un-derstood. How do changes in flux of substrates via an enzyme affect signaling specificity? How is the intracellular localization of a protein coupled to its signaling role? What is the function of a given protein in network regulation?

To address these questions, we employ a combi-nation of biochemical and advanced imaging tech-niques. We are developing tools to visualize the functional state of signaling molecules in live cells to determine how spatial distribution, regulation of specific activity and the corresponding changes in the flux of substrates via individual enzymes deter-mine their signaling function.

Furthermore, to examine the physiological rele-vance of individual enzymes in signaling, we will be developing new tools to selectively manipulate their localization and activity in live cells.

The ultimate specialization of T lymphocytes provides a cell biologist with a unique model sys-tem for functional studies. Specifically, we will be addressing the functional role of protein compart-mentalization in T cell signaling, probe the flux of substrates via the Src family kinases in T cells and develop methods to acutely destabilize select

signaling proteins to elucidate their physiological functions in T cells. The outlined approaches should also be broadly applicable in other model systems. Our findings will address a spectrum of fundamental questions of cellular physiology as well as provide insight into the molecular mechanisms of activation of T cells.

In the lab we are seeking to create a friendly, collaborative environment that promotes dynamic exchange of ideas and expertise between colleagues in and outside the lab and values initiative, origi-nality and scientific rigor. The group is looking for PhD students and Postdocs to pursue challenging, technology-oriented research projects.

Adaptive character of responses to signals from the environment is a fundamental property of all living organisms. At the cellular level, it is brought about by a highly integrated process of transmembrane and intracellular signal transduction.

SELECTED PUBLICATIONSYudushkin IA, Vale RD. Imaging T-cell receptor activation reveals accumulation of tyrosine-phosphorylated CD3ζ in the endosomal com-partment. Proc Natl Acad Sci U S A. 2010;107(51):22128-33. PMID: 21135224 De Graffenried CL, Anrather D, Von Raußendorf F, Warren G. Polo-like kinase phosphorylation of bilobe-resident TbCentrin2 facilitates flagellar inheritance in Trypanosoma brucei. Mol Biol Cell. 24(12):1947-63. PMID: 23615446 Yudushkin IA, Schleifenbaum A, Kinkhabwala A, Neel BG, Schultz C, Bastiaens PI. Live-cell imaging of enzyme-substrate interaction reveals spatial regulation of PTP1B. Science 2007;315(5808):115-9. PMID: 17204654

A) T cell receptor complex is phosphorylated on multipletyrosine residues upon binding of an antigen. This phosphorylation can be detected in live T cells using translocation- (A) or FRET-based(B) fluorescent reporters. Our data demonstrate that in T cells, active T cell receptor (C, green) localizes on endosomal vesicles (C, red) and may recruit and activate the essential downstream kinase ZAP-70.

Ivan Yudushkin

TEAMMichael EbnerNathan EhrlichFreia von Raußendorf





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B O J A N Ž A G R O V I Ć

Biophysics of macromolecular interactions

Conceptually, the research interests of our laboratory revolve around exploring different faces of this fundamental principle through the use of computational and theoretical methods in close collaboration with experimentalists. While experi-mental approaches are making breathtaking steps forward, currently the only way to probe biomolecular structure and dynamics at a single molecule level and with all-atom, femtosecond resolution is through molecular dynamics computer simulations, our primary workhorse.

Specifically, we are interested in the role of dynamics and conformational entropy in non-co-valent protein interactions. We develop new methods for calculating conformational entropy of biomolecules from computer simulations and for measuring it experimentally. In addition to function, dynamics also affects the very process of biomolecular structure determination. Namely, biomolecular structures are typically static models derived from experiments performed on many dynamic copies of the same molecule. We use MD simulations to help interpret such time- and ensemble-averaged experiments and analyze the impact of conformational averaging on the derived structures.

Second, all biomolecular processes occur in crowded, dynamic, constantly changing environments. We study how this affects protein- protein interactions and other basic processes such as protein folding or posttranslational modifications of proteins. In particular, we are interested in studying how binding partners find each other in the crowded cell and employ MD and Brownian dynamics simulations and structural bioinformatics methods to address this question.

Finally, we have recently discovered a remarkably robust correspondence between the nucleobase content of mRNAs and the propensity of cognate protein sequences to bind different nucleobases. We believe this provides strong support for the stereochemical hypothesis concerning the origin of the genetic code and suggests that cognate mRNAs and proteins may be physico-chemically complementary to each other. We use different computational methods including MD simulations, structural bioinformatics techniques and free energy methods as well as in vitro biochemistry to further explore this hypothesis.

Bojan Žagrović

TEAMAlexander BeribiskyAnita de RuiterMarkus FleckMatea HajnicMario HlevnjakGerald PlatzerAnton Polyansky

Biological function at the molecular level is directly related to the 3-dimensional structure of biomolecules, their dynamics and finally their interactions, both with the environment as well as with other biomolecules.

SELECTED PUBLICATIONSKuzmanic A, Pannu NS, Zagrovic B. X-ray refinement significantly underestimates the level of microscopic heterogeneity in biomole-cular crystals. Nat Commun. 2014;5:3220. PMID: 24504120 Polyansky AA, Zagrovic B. Evidence of direct complementary interactions between messenger RNAs and their cognate proteins. Nucleic Acids Res. 2013;41(18), 8434-8443. PMID: 23868089 Polyansky AA, Hlevnjak M, Zagrovic B. Analog encoding of physicochemical properties of proteins in their cognate messenger RNAs. Nat Commun. 2013;4:2784. PMID: 24253588

Linear correlations decrease the overall quasi-harmonic entropy change (ΔSQH) in protein-protein interactions by a remarkably constant amount as revealed by MD simulations of complexes of ubiquitin (UBQ) with its binding partners such as UBM2 (PD code: 2KTF) (Polyansky AA et al., J.Chem. Theory Comput., 2012).

The pyrimidine content profiles of mRNA coding sequences quantitatively match the profiles of their cognate proteins’ affinity for pyrimidine mimetics (“polar requirement”) as illustrated here for protein S100-A1 and its mRNA (Hlevn-jak M et al., Nucleic Acids Res., 2012).





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Publications

as of 19 December 2014

Velghe AI, Van Cauwenberghe S, Polyansky AA, Chand D, Montano-Almendras CP, Charni S, Hallberg B, Essaghir A, Demoulin JB. PDGFRA alterations in cancer: characterization of a gain-of-function V536E transmembrane mutant as well as loss-of-function and passenger mutations. Oncogene. 2014;33(20):2568-76. PMID: 23752188

Uecker H, Otto SP, Hermisson J. Evolutionary rescue in structured populations. Am Nat. 2014;183(1):E17-35. PMID: 24334746

Gregor M, Osmanagic-Myers S, Burgstaller G, Wolfram M, Fischer I, Walko G, Resch GP, Jörgl A, Herrmann H, Wiche G. Mechanosensing through focal adhesion-anchored intermediate filaments. FASEB J. 2014;28(2):715-29. PMID: 24347609

Vin H, Ching G, Ojeda SS, Adelmann CH, Chitsazzadeh V, Dwyer DW, Ma H, Ehrenreiter K, Baccarini M, Ruggieri R, Curry JL, Ciurea AM, Duvic M, Busaidy NL, Tannir NM, Tsai KY. Sorafenib suppresses JNK-dependent apoptosis through inhibition of ZAK. Mol Cancer Ther. 2014;13(1):221-9. PMID: 24170769

Polakova S, Benko Z, Zhang L, Gregan J. Mal3, the Schizosaccharomyces pombe homolog of EB1, is required for karyogamy and for promoting oscillatory nuclear movement during meiosis. Cell Cycle. 2014;13(1):72-7. PMID: 24196444

Märtens B, Manoharadas S, Hasenöhrl D, Zeichen L, Bläsi U. Back to translation: removal of aIF2 from the 5’-end of mRNAs by translation recovery factor in the crenarchaeon Sulfolobus solfataricus. Nucleic Acids Res. 2014;42(4):2505-11. PMID: 24271401

Bajari TM, Winnicki W, Gensberger ET, Scharrer SI, Regele H, Aumayr K, Kopecky C, Gmeiner BM, Hermann M, Zeillinger R, Sengölge G. Known players, new interplay in atherogenesis: Chronic shear stress and carbamylated-LDL induce and modulate expression of atherogenic LR11 in human coronary artery endothelium. Thromb Haemost. 2014;111(2):323-32. PMID: 24284991

Wienerroither S, Rauch I, Rosebrock F, Jamieson AM, Bradner J, Muhar M, Zuber J, Müller M, Decker T. Regulation of NO synthesis, local inflammation, and innate immunity to pathogens by BET family proteins. Mol Cell Biol. 2014;34(3):415-27. PMID: 24248598

Chi J, Mahé F, Loidl J, Logsdon J, Dunthorn M. Meiosis gene inventory of four ciliates reveals the prevalence of a synaptonemal complex-independent crossover pathway. Mol Biol Evol. 2014;31(3):660-72. PMID: 24336924

Dell’Ampio E, Meusemann K, Szucsich NU, Peters RS, Meyer B, Borner J, Petersen M, Aberer AJ, Stamatakis A, Walzl MG, Minh BQ, von Haeseler A, Ebersberger I, Pass G, Misof B. Decisive data sets in phylogenomics: lessons from studies on the phylogenetic relationships of primarily wingless insects. Mol Biol Evol. 2014;31(1):239-49. PMID: 24140757

Ebersberger I, Simm S, Leisegang MS, Schmitzberger P, Mirus O, von Haeseler A, Bohnsack MT, Schleiff E. The evolution of the ribosome biogenesis pathway from a yeast perspective. Nucleic Acids Res. 2014;42(3):1509-23. PMID: 24234440

Tobler R, Franssen SU, Kofler R, Orozco-Terwengel P, Nolte V, Hermisson J, Schlötterer C. Massive habitat-specific genomic response in D. melanogaster populations during experimental evolution in hot and cold environments. Mol Biol Evol. 2014;31(2):364-75. PMID: 24150039

Sára T, Konrat R, Skern T. Strategies for purifying variants of human rhinovirus 14 2C protein. Protein Expr Purif. 2014;95:28-37. PMID: 24316192

Fedosyuk S, Grishkovskaya I, de Almeida Ribeiro E Jr, Skern T. Characterization and structure of the vaccinia virus NF-κB antagonist A46. J Biol Chem. 2014;289(6):3749-62. PMID: 24356965

Rajsbaum R, García-Sastre A, Versteeg GA. TRIMmunity: the roles of the TRIM E3-ubiquitin ligase family in innate antiviral immunity. J Mol Biol. 2014;426(6):1265-84. PMID: 24333484

Vidilaseris K, Morriswood B, Kontaxis G, Dong G. Structure of the TbBILBO1 protein N-terminal domain from Trypanosoma brucei reveals an essential requirement for a conserved surface patch. J Biol Chem. 2014;289(6):3724-35. PMID: 24362019

Pfanzagl B, Andergassen D, Edlmayr J, Niespodziana K, Valenta R, Blaas D. Entry of human rhinovirus 89 via ICAM-1 into HeLa epithelial cells is inhibited by actin skeleton disruption and by bafilomycin. Arch Virol. 2014;159(1):125-40. PMID: 23913188

Lidke AK, Bannister S, Löwer AM, Apel DM, Podleschny M, Kollmann M, Ackermann CF, García-Alonso J, Raible F, Rebscher N. 17β-Estradiol induces supernumerary primordial germ cells in embryos of the polychaete Platynereis dumerilii. Gen Comp Endocrinol. 2014;196:52-61. PMID: 24287341

Zakrzewski AC, Weigert A, Helm C, Adamski M, Adamska M, Bleidorn C, Raible F, Hausen H. Early divergence, broad distribution, and high diversity of animal chitin synthases. Genome Biol Evol. 2014;6(2):316-25. PMID: 24443419





















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Hofbauer S, Gysel K, Bellei M, Hagmüller A, Schaffner I, Mlynek G, Kostan J, Pirker KF, Daims H, Furtmüller PG, Battistuzzi G, Djinović-Carugo K, Obinger C. Manipulating conserved heme cavity residues of chlorite dismutase: effect on structure, redox chemistry, and reactivity. Biochemistry. 2014;53(1):77-89. PMID: 24364531

Leeb C, Eresheim C, Nimpf J. Clusterin is a ligand for apolipoprotein E receptor 2 (ApoER2) and very low density lipoprotein receptor (VLDLR) and signals via the Reelin-signaling pathway. J Biol Chem. 2014;289(7):4161-72. PMID: 24381170

Wildauer M, Zemora G, Liebeg A, Heisig V, Waldsich C. Chemical probing of RNA in living cells. Methods Mol Biol. 2014;1086:159-76. PMID: 24136603

Sachsenmaier N, Handl S, Debeljak F, Waldsich C. Mapping RNA structure in vitro using nucleobase-specific probes. Methods Mol Biol. 2014;1086:79-94. PMID: 24136599

Begitt A, Droescher M, Meyer T, Schmid CD, Baker M, Antunes F, Knobeloch KP, Owen MR, Naumann R, Decker T, Vinkemeier U. STAT1-cooperative DNA binding distinguishes type 1 from type 2 interferon signaling. Nat Immunol. 2014;15(2):168-76. PMID: 24413774

Papinski D, Schuschnig M, Reiter W, Wilhelm L, Barnes CA, Majolica A, Hansmann I, Pfaffenwimmer T, Kijanska M, Stoffel I, Lee SS, Brezovich A, Lou JH, Turk BE, Aebersold R, Ammerer G, Peter M, Kraft C. Early Steps in Autophagy Depend on Direct Phosphorylation of Atg9 by the Atg1 Kinase. Mol Cell. 2014;53(3):471-83. PMID: 24440502

Eresheim C, Plieschnig J, Ivessa NE, Schneider WJ, Hermann M. Expression of microsomal triglyceride transfer protein in lipoproteinsynthesizing tissues of the developing chicken embryo. Biochimie 2014;101:67-74. PMID: 24394625

Hagelkruys A, Lagger S, Krahmer J, Leopoldi A, Artaker M, Pusch O, Zezula J, Weissmann S, Xie Y, Schöfer C, Schlederer M, Brosch G, Matthias P, Selfridge J, Lassmann H, Knoblich JA, Seiser C. A single allele of Hdac2 but not Hdac1 is sufficient for normal mouse brain development in the absence of its paralog. Development 2014;141(3):604-16. PMID: 24449838

Peter B, Polyansky AA, Fanucchi S, Dirr HW. A Lys-Trp cation-π interaction mediates the dimerization and function of the chloride intracellular channel protein 1 transmembrane domain. Biochemistry. 2014;53(1):57-67. PMID: 24328417

Polyansky AA, Chugunov AO, Volynsky PE, Krylov NA, Nolde DE, Efremov RG. PREDDIMER: a web server for prediction of transmembrane helical dimers. Bioinformatics. 2014;30(6):889-90. PMID: 24202542

Gibbs DJ, Md Isa N, Movahedi M, Lozano-Juste J, Mendiondo GM, Berckhan S, Marín-de la Rosa N, Vicente Conde J, Sousa Correia C, Pearce SP, Bassel GW, Hamali B, Talloji P, Tomé DF, Coego A, Beynon J, Alabadí D, Bachmair A, León J, Gray JE, Theodoulou FL, Holdsworth MJ. Nitric oxide sensing in plants is mediated by proteolytic control of group VII ERF transcription factors. Mol Cell. 2014;53(3):369-79. PMID: 24462115

Iantorno S, Gori K, Goldman N, Gil M, Dessimoz C. Who watches the watchmen? An appraisal of benchmarks for multiple sequence alignment. Methods Mol Biol. 2014;1079:59-73. PMID: 24170395

Orbán-Németh Z, Henen MA, Geist L, Żerko S, Saxena S, Stanek J, Koźmiński W, Propst F, Konrat R. Backbone and partial side chain assignment of the microtubule binding domain of the MAP1B light chain. Biomol NMR Assign. 2014;8(1):123-7. PMID: 23339032

Dönmez Cakil Y, Khunweeraphong N, Parveen Z, Schmid D, Artaker M, Ecker GF, Sitte HH, Pusch O, Stockner T, Chiba P. Pore-exposed tyrosine residues of P-glycoprotein are important hydrogen-bonding partners for drugs. Mol Pharmacol. 2014;85(3):420-8. PMID: 24366667

Carugo O, Djinovic-Carugo K. Packing bridges in protein crystal structures. J Appl Crystallogr. 2014;47:458–461.

Karstens K, Zschiedrich CP, Bowien B, Stülke J, Görke B. Phosphotransferase protein EIIANtr interacts with SpoT, a key enzyme of the stringent response, in Ralstonia eutropha H16. Microbiology. 2014;160(Pt 4):711-22. PMID: 24515609

Tomanov K, Luschnig C, Bachmair A. Ubiquitin Lys 63 chains - second-most abundant, but poorly understood in plants. Front Plant Sci. 2014;5:15. PMID: 24550925

Schmidt A, Trentini DB, Spiess S, Fuhrmann J, Ammerer G, Mechtler K, Clausen T. Quantitative phosphoproteomics reveals the role of protein arginine phosphorylation in the bacterial stress response. Mol Cell Proteomics. 2014;13(2):537-50. PMID: 24263382

Yelamanchi SK, Veis J, Anrather D, Klug H, Ammerer G. Genotoxic stress prevents Ndd1-dependent transcriptional activation of G2/M-specific genes in Saccharomyces cerevisiae. Mol Cell Biol. 2014;34(4):711-24. PMID: 24324010

Szkiba D, Kapun M, von Haeseler A, Gallach M. SNP2GO: functional analysis of genome-wide association studies. Genetics. 2014;197(1):285-9. PMID: 24561481

Praschberger M, Hermann M, Wanner J, Jirovetz L, Exner M, Kapiotis S, Gmeiner BM, Laggner H. The uremic toxin indoxyl sulfate acts as a pro- or antioxidant on LDL oxidation. Free Radic Res. 2014;48(6):641-8. PMID: 24568219

Kuzmanic A, Zagrovic B. Dependence of protein crystal stability on residue charge states and ion content of crystal solvent. Biophys J. 2014;106(3):677-86. PMID: 24507608

Kuzmanic A, Pannu NS, Zagrovic B. X-ray refinement significantly underestimates the level of microscopic heterogeneity in biomolecular crystals. Nat Commun. 2014;5:3220. PMID: 24504120

Göpel Y, Görke B. Lies and deception in bacterial gene regulation: the roles of nucleic acid decoys. Mol Microbiol. 2014;92(4):641-7. PMID: 24707963

























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Göpel Y, Khan M, Görke B. Ménage à trois: Post-transcriptional control of the key enzyme for cell envelope synthesis by a base-pairing small RNA, an RNase adaptor protein, and a small RNA mimic. RNA Biol. 2014;11(5):433-42. PMID: 24667238

Bannister S, Antonova O, Polo A, Lohs C, Hallay N, Valinciute A, Raible F, Tessmar-Raible K. TALENs mediate efficient and heritable mutation of endogenous genes in the marine annelid Platynereis dumerilii. Genetics. 2014;197(1):77-89. PMID: 24653002

Boucheron N, Tschismarov R, Goeschl L, Moser MA, Lagger S, Sakaguchi S, Winter M, Lenz F, Vitko D, Breitwieser FP, Müller L, Hassan H, Bennett KL, Colinge J, Schreiner W, Egawa T, Taniuchi I, Matthias P, Seiser C, Ellmeier W. CD4(+) T cell lineage integrity is controlled by the histone deacetylases HDAC1 and HDAC2. Nat Immunol. 2014;15(5):439-48. PMID: 24681565

Sawa-Makarska J, Abert C, Romanov J, Zens B, Ibiricu I, Martens S. Cargo binding to Atg19 unmasks additional Atg8 binding sites to mediate membrane-cargo apposition during selective autophagy. Nat Cell Biol. 2014; 16(5):425-33. PMID: 24705553

Harutyunyan S, Kowalski H, Blaas D. The Rhinovirus subviral a-particle exposes 3’-terminal sequences of its genomic RNA. J Virol. 2014;88(11):6307-17. PMID: 24672023

Backfisch B, Kozin VV, Kirchmaier S, Tessmar-Raible K, Raible F. Tools for gene-regulatory analyses in the marine annelid Platynereis dumerilii. PLoS One. 2014;9(4):e93076. PMID: 24714200

Oliveri P, Fortunato AE, Petrone L, Ishikawa-Fujiwara T, Kobayashi Y, Todo T, Antonova O, Arboleda E, Zantke J, Tessmar-Raible K, Falciatore A. The Cryptochrome/Photolyase Family in aquatic organisms. Mar Genomics. 2014;14:23-37. PMID: 24568948

Barraud P, Banerjee S, Mohamed WI, Jantsch MF, Allain FH. A bimodular nuclear localization signal assembled via an extended double-stranded RNA-binding domain acts as an RNA-sensing signal for transportin 1. Proc Natl Acad Sci U S A. 2014;111(18):E1852-1861. PMID: 24753571

Petrillo E, Godoy Herz MA, Fuchs A, Reifer D, Fuller J, Yanovsky MJ, Simpson C, Brown JW, Barta A, Kalyna M, Kornblihtt AR. A chloroplast retrograde signal regulates nuclear alternative splicing. Science. 2014;344(6182):427-30. PMID: 24763593

Mike AK, Koenig X, Koley M, Heher P, Wahl G, Rubi L, Schnürch M, Mihovilovic MD, Weitzer G, Hilber K. Small molecule cardiogenol C upregulates cardiac markers and induces cardiac functional properties in lineage-committed progenitor cells. Cell Physiol Biochem. 2014;33(1):205-21. PMID: 24481283

Pfaffenwimmer T, Reiter W, Brach T, Nogellova V, Papinski D, Schuschnig M, Abert C, Ammerer G, Martens S, Kraft C. Hrr25 kinase promotes selective autophagy by phosphorylating the cargo receptor Atg19. EMBO Rep. 2014;15(8):862-70. PMID: 24968893

Papinski D, Kraft C. Atg1 kinase organizes autophagosome formation by phosphorylating Atg9. Autophagy. 2014;10(7):1338-40. PMID: 24905091

Gallach M. Recurrent turnover of chromosome-specific satellites in Drosophila. Genome Biol Evol. 2014;6(6):1279-86. PMID: 24846631

Sorourian M, Kunte MM, Domingues S, Gallach M, Özdil F, Río J, Betrán E. Relocation facilitates the acquisition of short cis-regulatory regions that drive the expression of retrogenes during spermatogenesis in drosophila. Mol Biol Evol. 2014;31(8):2170-80. PMID: 24855141

Vidilaseris K, Dong G. Expression, purification and preliminary crystallographic analysis of the N-terminal domain of Trypanosoma brucei BILBO1. Acta Crystallogr F Struct Biol Commun. 2014;70(Pt 5):628-31. PMID: 24817725

Muggenhumer D, Vesely C, Nimpf S, Tian N, Yongfeng J, Jantsch MF. Drosha protein levels are translationally regulated during Xenopus oocyte maturation. Mol Biol Cell. 2014;25(13):2094-104. PMID: 24829383

Garg RK, Rennert RC, Duscher D, Sorkin M, Kosaraju R, Auerbach LJ, Lennon J, Chung MT, Paik K, Nimpf J, Rajadas J, Longaker MT, Gurtner GC. Capillary force seeding of hydrogels for adipose-derived stem cell delivery in wounds. Stem Cells Transl Med. 2014;3(9):1079-89. PMID: 25038246

Sealey-Cardona M, Schmidt K, Demmel L, Hirschmugl T, Gesell T, Dong G, Warren G. Sec16 determines the size and functioning of the Golgi in the protist parasite, Trypanosoma brucei. Traffic. 2014;15(6):613-29. PMID: 24612401

Erven C, Meyer-Scott E, Fisher K, Lavoie J, Higgins BL, Yan Z, Pugh CJ, Bourgoin JP, Prevedel R, Shalm LK, Richards L, Gigov N, Laflamme R, Weihs G, Jennewein T, Resch KJ. Experimental Three-Particle Quantum Nonlocality under Strict Locality Conditions. Nature photonics. 2014;8(4):292 - 296.

Fisher KA, Broadbent A, Shalm LK, Yan Z, Lavoie J, Prevedel R, Jennewein T, Resch KJ. Quantum computing on encrypted data. Nat Commun. 2014;5:3074. PMID: 24445949

Shanmugam R, Aklujkar M, Schäfer M, Reinhardt R, Nickel O, Reuter G, Lovley DR, Ehrenhofer-Murray A, Nellen W, Ankri S, Helm M, Jurkowski TP, Jeltsch A. The Dnmt2 RNA methyltransferase homolog of Geobacter sulfurreducens specifically methylates tRNA-Glu. Nucleic Acids Res. 2014;42(10):6487-96. PMID: 24711368

Villajuana-Bonequi M, Elrouby N, Nordström K, Griebel T, Bachmair A, Coupland G. Elevated salicylic acid levels conferred by increased expression of ISOCHORISMATE SYNTHASE 1 contribute to hyperaccumulation of SUMO1 conjugates in the Arabidopsis mutant early in short days 4. Plant J. 2014;79(2):206-19. PMID: 24816345























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Rajsbaum R, Versteeg GA, Schmid S, Maestre AM, Belicha-Villanueva A, Martínez-Romero C, Patel JR, Morrison J, Pisanelli G, Miorin L, Laurent-Rolle M, Moulton HM, Stein DA, Fernandez-Sesma A, tenOever BR, García-Sastre A. Unanchored K48-linked polyubiquitin synthesized by the E3-ubiquitin ligase TRIM6 stimulates the interferon-IKKε kinase-mediated antiviral response. Immunity. 2014;40(6):880-95. PMID: 24882218

Cheng L, Desai J, Miranda CJ, Duncan JS, Qiu W, Nugent AA, Kolpak AL, Wu CC, Drokhlyansky E, Delisle MM, Chan WM, Wei Y, Propst F, Reck-Peterson SL, Fritzsch B, Engle EC. Human CFEOM1 mutations attenuate KIF21A autoinhibition and cause oculomotor axon stalling. Neuron. 2014;82(2):334-49. PMID: 24656932

Preiner J, Kodera N, Tang J, Ebner A, Brameshuber M, Blaas D, Gelbmann N, Gruber HJ, Ando T, Hinterdorfer P. IgGs are made for walking on bacterial and viral surfaces. Nat Commun. 2014;5:4394. PMID: 25008037

Hämmerle H, Amman F, Večerek B, Stülke J, Hofacker I, Bläsi U. Impact of Hfq on the Bacillus subtilis transcriptome. PLoS One. 2014;9(6):e98661. PMID: 24932523

Sonnleitner E, Bläsi U. Regulation of Hfq by the RNA CrcZ in Pseudomonas aeruginosa carbon catabolite repression. PLoS Genet. 2014;10(6):e1004440. PMID: 24945892

Kienesberger K, Pordes AG, Völk TG, Hofbauer R. L-carnitine and PPARα-agonist fenofibrate are involved in the regulation of Carnitine Acetyltransferase (CrAT) mRNA levels in murine liver cells. BMC Genomics. 2014;15:514. PMID: 24962334

Sawicka A, Seiser C. Sensing core histone phosphorylation - a matter of perfect timing. Biochim Biophys Acta. 2014;1839(8):711-8. PMID: 24747175

Dinis M, Plainvert C, Kovarik P, Longo M, Fouet A, Poyart C. The innate immune response elicited by Group A Streptococcus is highly variable among clinical isolates and correlates with the emm type. PLoS One. 2014;9(7):e101464. PMID: 24991887

Prevedel R, Yoon YG, Hoffmann M, Pak N, Wetzstein G, Kato S, Schrödel T, Raskar R, Zimmer M, Boyden ES, Vaziri A. Simultaneous whole-animal 3D imaging of neuronal activity using light-field microscopy. Nat Meth. 2014;11(7):727-30. PMID: 24836920

Demmel L, Schmidt K, Lucast L, Havlicek K, Zankel A, Koestler T, Reithofer V, de Camilli P, Warren G. The endocytic activity of the flagellar pocket in Trypanosoma brucei is regulated by an adjacent phosphatidylinositol phosphate kinase. J Cell Sci. 2014;127(Pt 10):2351-64. PMID: 24639465

Rauch I, Hainzl E, Rosebrock F, Heider S, Schwab C, Berry D, Stoiber D, Wagner M, Schleper C, Loy A, Urich T, Müller M, Strobl B, Kenner L, Decker T. Type I interferons have opposing effects during the emergence and recovery phases of colitis. Eur J Immunol. 2014;44(9):2749-60. PMID: 24975266

Bilusic I, Popitsch N, Rescheneder P, Schroeder R, Lybecker M. Revisiting the coding potential of the E. coli genome through Hfq co-immunoprecipitation. RNA Biol. 2014;11(5):641-54. PMID: 24922322

Popitsch N. CODOC: efficient access, analysis and compression of depth of coverage signals. Bioinformatics. 2014;30(18):2676-7. PMID: 24872424

Mlynek G, Lehner A, Neuhold J, Leeb S, Kostan J, Charnagalov A, Stolt-Bergner P, Djinović-Carugo K, Pinotsis N. The Center for Optimized Structural Studies (COSS) platform for automation in cloning, expression, and purification of single proteins and protein-protein complexes. Amino Acids. 2014;46(6):1565-82. PMID: 24647677

Hofbauer S, Gruber C, Pirker KF, Schaffner I, Furtmuller PG, Obinger C, Hagmuller A, Gysel K, Mlynek G, Kostan J, Djinović-Carugo K, Bellei M, Battistuzzi G, Daims H. Understanding Chlorite Dismutase from Candidatus Nitrospira defluvii. J Biol Inorg Chem. 2014;19:253-253.

Kostan J, Salzer U, Orlova A, Törö I, Hodnik V, Senju Y, Zou J, Schreiner C, Steiner J, Meriläinen J, Nikki M, Virtanen I, Carugo O, Rappsilber J, Lappalainen P, Lehto VP, Anderluh G, Egelman EH, Djinović-Carugo K. Direct interaction of actin filaments with F-BAR protein pacsin2. EMBO Rep. 2014;15(11):1154-62. PMID: 25216944

Nikolay B, Fall G, Boye CS, Sall AA, Skern T. Validation of a structural comparison of the antigenic characteristics of Usutu virus and West Nile virus envelope proteins. Virus Res. 2014;189:87-91. PMID: 24874193

Vidilaseris K, Shimanovskaya E, Esson HJ, Morriswood B, Dong G. Assembly Mechanism of Trypanosoma brucei BILBO1, a Multidomain Cytoskeletal Protein. J Biol Chem 2014;289(34):23870-81. PMID: 25031322

Göhring J, Fulcher N, Jacak J, Riha K. TeloTool: a new tool for telomere length measurement from terminal restriction fragment analysis with improved probe intensity correction. Nucleic Acids Res. 2014;42(3):e21. PMID: 24366880

Sams M, Silye R, Göhring J, Muresan L, Schilcher K, Jacak J. Spatial cluster analysis of nanoscopically mapped serotonin receptors for classification of fixed brain tissue. J Biomed Opt. 2014;19(1):011021. PMID: 24297043

Singh A, Kanwar P, Yadav AK, Mishra M, Jha SK, Baranwal V, Pandey A, Kapoor S, Tyagi AK, Pandey GK. Genome-wide expressional and functional analysis of calcium transport elements during abiotic stress and development in rice. FEBS J. 2014;281(3):894-915. PMID: 24286292

Shi M, Cho H, Inn KS, Yang A, Zhao Z, Liang Q, Versteeg GA, Amini-Bavil-Olyaee S, Wong LY, Zlokovic BV, Park HS, García-Sastre A, Jung JU. Negative regulation of NF-κB activity by brain-specific TRIpartite Motif protein 9. Nat Commun. 2014;5:4820. PMID: 25190485





http://www.ncbi.nlm.nih.gov/pubmed/?term=Regulation+of+Hfq+by+the+RNA+CrcZ+in+Pseudomonas+aeruginosa+carbon+catabolite+repression.


















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Versteeg GA, Benke S, García-Sastre A, Rajsbaum R. InTRIMsic immunity: Positive and negative regulation of immune signaling by tripartite motif proteins. Cytokine Growth Factor Rev. 2014;25(5):563-76. PMID: 25172371

Weber C, Hartig A, Hartmann RK, Rossmanith W. Playing RNase P evolution: swapping the RNA catalyst for a protein reveals functional uniformity of highly divergent enzyme forms. PLoS Genet. 2014;10(8):e1004506. PMID: 25101763

Gigova A, Duggimpudi S, Pollex T, Schaefer M, Koš M. A cluster of methylations in the domain IV of 25S rRNA is required for ribosome stability. RNA. 2014;20(10):1632-44. PMID: 25125595

Boban M, Pantazopoulou M, Schick A, Ljungdahl PO, Foisner R. A nuclear ubiquitin-proteasome pathway targets the inner nuclear membrane protein Asi2 for degradation. J Cell Sci. 2014;127(Pt 16):3603-13. PMID: 24928896

Brachner A, Foisner R. Lamina-associated polypeptide (LAP)2α and other LEM proteins in cancer biology. Adv Exp Med Biol. 2014;773:143-63. PMID: 24563347

Göhring J, Jacak J, Barta A. Imaging of endogenous messenger RNA splice variants in living cells reveals nuclear retention of transcripts inaccessible to nonsense-mediated decay in Arabidopsis. Plant Cell. 2014;26(2):754-64. PMID: 24532591

Wögenstein KL, Szabo S, Lunova M, Wiche G, Haybaeck J, Strnad P, Boor P, Wagner M, Fuchs P. Epiplakin deficiency aggravates murine caerulein-induced acute pancreatitis and favors the formation of acinar keratin granules. PLoS One. 2014;9(9):e108323. PMID: 25232867

Vesely C, Tauber S, Sedlazeck FJ, Tajaddod M, Haeseler Av, Jantsch MF. ADAR2 induces reproducible changes in sequence and abundance of mature microRNAs in the mouse brain. Nucleic Acids Res. 2014;42(19):12155-68. PMID: 25260591

Moeton M, Kanski R, Stassen OM, Sluijs JA, Geerts D, van Tijn P, Wiche G, van Strien ME, Hol EM. Silencing GFAP isoforms in astrocytoma cells disturbs laminin-dependent motility and cell adhesion. FASEB J. 2014;28(7):2942-54. PMID: 24696300

Winter L, Staszewska I, Mihailovska E, Fischer I, Goldmann WH, Schröder R, Wiche G. Chemical chaperone ameliorates pathological protein aggregation in plectin-deficient muscle. J Clin Invest. 2014;124(3):1144-57. PMID: 24487589

Sutoh Yoneyama M, Hatakeyama S, Habuchi T, Inoue T, Nakamura T, Funyu T, Wiche G, Ohyama C, Tsuboi S. Vimentin intermediate filament and plectin provide a scaffold for invadopodia, facilitating cancer cell invasion and extravasation for metastasis. Eur J Cell Biol. 2014;93(4):157-69. PMID: 24810881

Molt S, Bührdel JB, Yakovlev S, Schein P, Orfanos Z, Kirfel G, Winter L, Wiche G, van der Ven PF, Rottbauer W, Just S, Belkin AM, Fürst DO. Aciculin interacts with filamin C and Xin and is essential for myofibril assembly, remodeling and maintenance. J Cell Sci. 2014;127(Pt 16):3578-92. PMID: 24963132

Cabral G, Marques A, Schubert V, Pedrosa-Harand A, Schlögelhofer P. Chiasmatic and achiasmatic inverted meiosis of plants with holocentric chromosomes. Nat Commun. 2014;5:5070. PMID: 25295686

Fieber C, Kovarik P. Responses of innate immune cells to group A Streptococcus. Front Cell Infect Microbiol. 2014;4:140. PMID: 25325020

Pollak DD, Minh BQ, Cicvaric A, Monje FJ. A novel fibroblast growth factor receptor family member promotes neuronal outgrowth and synaptic plasticity in aplysia. Amino Acids. 2014;46(11):2477-88. PMID: 25059541

Kaschner M, Loeschcke A, Krause J, Minh BQ, Heck A, Endres S, Svensson V, Wirtz A, von Haeseler A, Jaeger KE, Drepper T, Krauss U. Discovery of the first light-dependent protochlorophyllide oxidoreductase in anoxygenic phototrophic bacteria. Mol Microbiol. 2014;93(5):1066-78. PMID: 25039543

Sengoelge G, Winnicki W, Kupczok A, von Haeseler A, Schuster M, Pfaller W, Jennings P, Weltermann A, Blake S, Sunder-Plassmann G. A SAGE based approach to human glomerular endothelium: defining the transcriptome, finding a novel molecule and highlighting endothelial diversity. BMC Genomics. 2014;15:725. PMID: 25163811

Watanabe H, Schmidt HA, Kuhn A, Höger SK, Kocagöz Y, Laumann-Lipp N, Ozbek S, Holstein TW. Nodal signalling determines biradial asymmetry in Hydra. Nature. 2014;515(7525):112-5. PMID: 25156256

Mihailovska E, Raith M, Valencia RG, Fischer I, Banchaabouchi MA, Herbst R, Wiche G. Neuromuscular synapse integrity requires linkage of acetylcholine receptors to postsynaptic intermediate filament networks via rapsyn-plectin 1f complexes. Mol Biol Cell. 2014;25(25):4130-49. PMID: 25318670

Petrov D, Zagrovic B. Are current atomistic force fields accurate enough to study proteins in crowded environments? PLoS Comput Biol. 2014;10(5):e1003638. PMID: 24854339

Raible F, Tessmar-Raible K. Platynereis dumerilii. Curr Biol. 2014;24(15):R676-7. PMID: 25093553

Raible F, Falciatore A. It’s about time: rhythms as a new dimension of molecular marine research. Mar Genomics. 2014;14:1-2. PMID: 24952925

Zantke J, Bannister S, Rajan VB, Raible F, Tessmar-Raible K. Genetic and genomic tools for the marine annelid Platynereis dumerilii. Genetics. 2014;197(1):19-31. PMID: 24807110

























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Sarangi P, Altmannova V, Holland C, Bartosova Z, Hao F, Anrather D, Ammerer G, Lee SE, Krejci L, Zhao X. A versatile scaffold contributes to damage survival via sumoylation and nuclease interactions. Cell Rep. 2014;9(1):143-52. PMID: 25263559

Steinberger J, Grishkovskaya I, Cencic R, Juliano L, Juliano MA, Skern T. Foot-and-mouth disease virus leader proteinase: Structural insights into the mechanism of intermolecular cleavage. Virology. 2014;468-470:397-408. PMID: 25240326

Moral-López P, Alvarez E, Redondo N, Skern T, Carrasco L. L protease from foot and mouth disease virus confers eIF2-independent translation for mRNAs bearing picornavirus IRES. FEBS Lett. 2014;588(21):4053-9. PMID: 25268112

Tschismarov R, Firner S, Gil-Cruz C, Göschl L, Boucheron N, Steiner G, Matthias P, Seiser C, Ludewig B, Ellmeier W. HDAC1 controls CD8+ T cell homeostasis and antiviral response. PLoS One. 2014;9(10):e110576. PMID: 25333902

Scholz HC, Pearson T, Hornstra H, Projahn M, Terzioglu R, Wernery R, Georgi E, Riehm JM, Wagner DM, Keim PS, Joseph M, Johnson B, Kinne J, Jose S, Hepp CM, Witte A, Wernery U. Genotyping of Burkholderia mallei from an outbreak of glanders in Bahrain suggests multiple introduction events. PLoS Negl Trop Dis. 2014;8(9):e3195. PMID: 25255232

Khan M, Rozhon W, Unterholzner SJ, Chen T, Eremina M, Wurzinger B, Bachmair A, Teige M, Sieberer T, Isono E, Poppenberger B. Interplay between phosphorylation and SUMOylation events determines CESTA protein fate in brassinosteroid signalling. Nat Commun. 2014;5:4687. PMID: 25134617

Matuszewski S, Hermisson J, Kopp M. Fisher’s geometric model with a moving optimum. Evolution. 2014;68(9):2571-88. PMID: 24898080

Uecker H, Setter D, Hermisson J. Adaptive gene introgression after secondary contact. J Math Biol. 2014 [Epub ahead of print]. PMID: 24992884

Huber CD, Nordborg M, Hermisson J, Hellmann I. Keeping it local: evidence for positive selection in Swedish Arabidopsis thaliana. Mol Biol Evol. 2014;31(11):3026-39. PMID: 25158800

Cseh B, Doma E, Baccarini M. “RAF” neighborhood: protein-protein interaction in the Raf/Mek/Erk pathway. FEBS Lett. 2014;588(15):2398-406. PMID: 24937142

Baumgartner C, Baccarini M. p38 links RAS to GATA2. EMBO Rep. 2014;15(9):912-3. PMID: 25092790

Shodhan A, Lukaszewicz A, Novatchkova M, Loidl J. Msh4 and Msh5 Function in SC-Independent Chiasma Formation During the Streamlined Meiosis of Tetrahymena. Genetics. 2014;198(3):983-93. PMID: 25217051

Montgomery TR, Steinkellner T, Sucic S, Koban F, Schüchner S, Ogris E, Sitte HH, Freissmuth M. Axonal targeting of the serotonin transporter in cultured rat dorsal raphe neurons is specified by SEC24C-dependent export from the endoplasmic reticulum. J Neurosci. 2014;34(18):6344-51. PMID: 24790205

Mannion NM, Greenwood SM, Young R, Cox S, Brindle J, Read D, Nellåker C, Vesely C, Ponting CP, McLaughlin PJ, Jantsch MF, Dorin J, Adams IR, Scadden AD, Ohman M, Keegan LP, O’Connell MA. The RNA-Editing Enzyme ADAR1 Controls Innate Immune Responses to RNA. Cell Rep. 2014;9(4):1482-94. PMID: 25456137

Klepsatel P, Gáliková M, Huber CD, Flatt T. Similarities and differences in altitudinal versus latitudinal variation for morphological traits in Drosophila melanogaster. Evolution. 2014;68(5):1385-98. PMID: 24410363

Cieślik-Boczula K, Swiątek P, Jaszczyszyn A, Zawilska P, Gąsiorowski K, Malinka W, Köhler G. Phase separation in phosphatidylcholine membrane caused by the presence of a pyrimidine analogue of fluphenazine with high anti-multidrug-resistance activity. J Phys Chem B. 2014;118(13):3605-15. PMID: 24601791

Alaghaz AN, Ammar RA, Koehler G, Wolschann KP, El-Gogary TM. Synthesis, spectral and quantum chemical studies on NO-chelating sulfamonomethoxine-cyclophosph(V)azane and its Er(III) complex. Spectrochim Acta A Mol Biomol Spectrosc. 2014;128:724-9. PMID: 24699291

Kurzbach D, Schwarz TC, Platzer G, Höfler S, Hinderberger D, Konrat R. Compensatory adaptations of structural dynamics in an intrinsically disordered protein complex. Angew Chem Int Ed Engl. 2014;53(15):3840-3. PMID: 24604825

Konrat R. NMR contributions to structural dynamics studies of intrinsically disordered proteins. J Magn Reson. 2014;241:74-85. PMID: 24656082

Sára T, Schwarz TC, Kurzbach D, Wunderlich CH, Kreutz C, Konrat R. Magnetic resonance access to transiently formed protein complexes. ChemistryOpen. 2014;3(3):115-23. PMID: 25050230

Fenollar-Ferrer C, Stockner T, Schwarz TC, Pal A, Gotovina J, Hofmaier T, Jayaraman K, Adhikary S, Kudlacek O, Mehdipour AR, Tavoulari S, Rudnick G, Singh SK, Konrat R, Sitte HH, Forrest LR. Structure and regulatory interactions of the cytoplasmic terminal domains of serotonin transporter. Biochemistry. 2014;53(33):5444-60. PMID: 25093911

Shen C, Knapp M, Puchinger MG, Shahzad A, Gaubitzer E, Shen AD, Koehler G. Using fluorescence correlation spectroscopy (FCS) for IFN-g detection: a preliminary study. J Immunol Methods. 2014;407:35-9. PMID: 24685834
























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