Candida albicansCandida albicans pathogenesis and systems ... · Candida albicans enjoys differing relationships with its human host … Aberdeen Fungal Group commensalism systemic

Aberdeen Fungal Group

AFG

Candida albicans pathogenesisCandida albicans pathogenesis and systems biology

Alistair BrownAlistair BrownAberdeen Fungal GroupUniversity of Aberdeen

Candida albicans enjoys differing relationships with its human host …


p

commensalism

systemic infections

mucosal infections

fungus hostg

… and occupies contrasting and complex niches within its human host

How does Candida albicans responddynamically to host niches?


y y

Major AIM:To understand the

fungus host

adaptation of the fungus to these

complex environmentsg complex environments


Experimental approaches

l l globallocal global

Dissection of specific target functions:Expression patterns

l l h

Exploration on genome-wide basis:Transcriptomics & ChIP on chip

Molecular phenotypes Cellular phenotypesPhysical interactions

Proteomics Genome-wide phenotypic screens Genome-wide interaction screensPhysical interactions

…Genome wide interaction screens …


Systems biology approaches


modeling of specific modules:Qualitative & semi-quantitative

( )

Network construction:Genetic networks

b d kQuantitative (ODE)Stochastic …

Expression-based networksInteraction networks …… …

LONG TERM AIMS: To define the networks that underpin cell biologyTo define the dynamic behaviour of these systems and how these behaviours contribute to cell adaptation, growth and development To explore the evolution of these biological systems

Aberdeen Fungal GroupMathematical modeling is in its infancy in C. albicansy



( )Quantitative (ODE)Stochastic ……

Aberdeen Fungal GroupThis talk …



( )ASPIRATIONS:

I d i i fQuantitative (ODE)Stochastic …

Improve our dynamic views of adaptive responsesIncrease our understanding of the contributions of specific components

GCN System:Rosa De Llanos Tao You… contributions of specific components

to dynamic behavioursExplanations for non-intuitive observations

Laura Selway Priya ChandrasekaranHelene Tournu George Coghill

THREE SHORT STORIES:Amino acid starvation (GCN)Osmotic stress

observationsInsights into the evolution of dynamic systemsInsights into the relevance of

Amino acid starvation (GCN)Osmotic stressHeat shock

Insights into the relevance of regulatory systems to pathogenicity

Aberdeen Fungal GroupAmino Acid Starvation (GCN) System

amino acidstarvation

Gcn2

[uncharged tRNAs]

eIF2α-P

GCN4 mRNA

Gcn4

amino acid

aa genes (HIS3)GCRE

amino acidbiosynthesis

Aberdeen Fungal GroupAmino Acid Starvation (GCN) Model

histidinemRNA

translationmodule

4 modules

module

3AT

histidine

GCN4 mRNA

histidine biosynthesis

GCN4 mRNAtranslational

control moduleGcn4

degradationTao You

Aberdeen Fungal GroupThe process – building the mRNA translation module

mRNA translation module:1. Build a reaction

network

You et al. (2010) Yeast 27, 785



network2. Biochemical 2. Biochemical

formulation of the model

You et al. (2010) Yeast 27, 785





3. Mathematical formulation of the model

You et al. (2010) Yeast 27, 785






4. Model t i ti dparameterization and

validation

You et al. (2010) Yeast 27, 785






4. Model t i ti dparameterization and

validation5. Module predictions

Translation initiation is robust to internal perturbations (e.g. changes in eIF concentrations)Translation initiation is sensitive to external perturbations (e.g. changes in histidine availability)

You et al. (2010) Yeast 27, 785

Aberdeen Fungal GroupThe process – building the Amino Acid Starvation (GCN) Model( )

histidine1 A bl th d l1. Assemble the modules2. Reparameterize and

validate

3AT3AT

3. Apply the model –built using artificial

di iconditions – to simulate a more physiologically relevant situationrelevant situation

Response to histidine starvation imposed by

Tao You

Response to histidine starvation imposed by 3-aminotriazole

Aberdeen Fungal GroupUse model to simulate natural amino acid starvation

1 A bl th d l

Gcn4eIf2αP

1. Assemble the modules2. Reparameterize and

validate

3. Apply the model –built using artificial

di iconditions – to simulate a more physiologically relevant situationrelevant situation

Tao You

Aberdeen Fungal GroupThe GCN Systems behave similarly. However, they operate differently, y p y

GCN4 is transcriptionally

CaGCN4 +/+ +/- -/- -/-/+

GCN43AT +- +- +- +-

GCN4 is transcriptionally activated in C. albicans

GCN4

ACT1HIS4

Gcn2-mediated translational regulation is NOT required for Gcn4-mediated

i d ti i C lbigene induction in C. albicans

Gcn2-mediated translational regulation is NOT essential for gadaptation in C. albicans

Tournu et al. (2005) Eukaryotic Cell, 4, 1687; Tripathi et al. (2002) EMBO J. 21, 5448

Aberdeen Fungal GroupAmino Acid Starvation (GCN) Systems


Gcn2

starvation[uncharged tRNAs]

S. cerevisiaeGcn2

GCN4 geneeIF2α-P

translational regulation

GCN4 mRNA

G 4

GCN4 mRNA

i id

aa genes (HIS3)GCRE

Gcn4

amino acidbiosynthesis

Aberdeen Fungal GroupAmino Acid Starvation (GCN) Systems


C. albicansGcn2

starvation[uncharged tRNAs]

transcriptional regulationGcn2

GCN4 geneeIF2α-P

translational regulation

regulation

GCN4 mRNA

G 4

GCN4 mRNA

In C. albicans:l l

i id

aa genes (HIS3)GCRE

Gcn4Translational regulation less importantT i ti l

Role of uORF spacing in alteringamino acid

biosynthesisTranscriptional regulation more important

altering translational regulation?


ScGCN4 translational regulation

GCN4 mRNA (A)nuORF1 uORF4

( )n

eIF2·GTPuORF4p v6

GTM

60S mRNAGCN4

mRNA v GTM60S

Gcn4p40S+

TC40S

Met-tRNAiMet Ribosome·mRNAGCN4

(uORF4)

48S RNA

v6

60S

mRNAGCN4

Ribosome·mRNAGCN4(GCN4)

60S

v8GTM

43S

48S·mRNAGCN4

mRNAGCN4

eIF2·GTP TC

48S·mRNAGCN4(<uORF4)

eIF2·GTP TC

48S·mRNAGCN4(<uORF4)

60S

v1GTM

Ribosome·mRNAGCN4(uORF1)

GCN4

60S

40S·mRNAGCN4(<uORF4)v GTM

v3GTM

Met-tRNAiMet

v5GTM

40S·mRNAGCN4( GCN4)

eIF2·GTP TC

Met-tRNAiMet

v7GTM

S. cerevisiae

(uORF1)

uORF1p

60S(<uORF4)v2

GTM

40Sv4

GTM(<GCN4)

Tao You


ScGCN4 translational regulation

GCN4 mRNA (A)nuORF1 uORF4

( )nn1 n2

Under replete conditions few ribosomes translate the main GCN4 ORF

Under starvation conditionsUnder starvation conditions ~25% ribosomes translate the main GCN4 ORF

G t t l (1994)

S. cerevisiaeTao You

Grant et al. (1994)Mol Cell Biol 14, 2616


CaGCN4 translational regulation

GCN4 mRNA (A)nuORF1 uORF3uORF2

( )nn1 n2

Could the differential arrangement ofCould the differential arrangement of uORFs in the C. albicans GCN4 mRNA account for the reduced impact of Gcn2-mediated translational regulation?

Under starvation conditions ~40% ribosomes translate the

mediated translational regulation?

40% ribosomes translate the main GCN4 ORF

Simple spatial effects unlikely!Alternative mechanisms probably account for the p yreduced translational regulation in C. albicans

C. albicansPriya Chandrasekaran




( )ASPIRATIONS:


Improve our dynamic views of adaptive responsesIncrease our understanding of the contributions of specific components… contributions of specific components to dynamic behavioursExplanations for non-intuitive observationsTHREE SHORT STORIES:

Amino acid starvation (GCN)Osmotic stress

observationsInsights into the evolution of dynamic systemsInsights into the relevance ofOsmotic stress

Heat shockInsights into the relevance of regulatory systems to pathogenicity



Osmotic Stress:Despoina Kaloriti Tao Youmodeling of specific modules:

Qualitative & semi-quantitative( )

ASPIRATIONS: I d i i f

Despoina Kaloriti Tao YouMette Jacobsen Piers IngramAnna Tillmann Wei PangNeil Gow Mamen RomanoQuantitative (ODE)

Stochastic …


Neil Gow Mamen RomanoKen Haynes Marco ThielEmily Cook Alessandro MouraMelanie Putnam Celso Grebogi… contributions of specific components


Melanie Putnam Celso GrebogiAndy McDonaghJanet Quinn




Aberdeen Fungal GroupOsmotic stress adaptation in C. albicans – the aspirationp

Klipp et al. (2002) Nature Biotechnology 23, 975-982.

Aberdeen Fungal GroupOsmotic stress adaptation in C. albicans – the problemp

Conservation of molecular systems in C. albicansOsmotic stress signalling network

Conservation of molecular systems in C. albicansTranslational apparatus

Many components of the osmotic stress signalling network have

Nikolaou et al. (2009) BMC Evol. Biol. 9, 44.

a y co po e s o e os o c s ess s g a g e o a ediverged significantly

Aberdeen Fungal GroupOsmotic stress adaptation in C. albicans – the problemp

The Ste11/Sho1 module is not required for osmotic stress signalling toHog1 in C. albicansHog1 in C. albicans

Tao You, Piers Ingram Cheetham et al. (2007) Mol Biol Cell 18, 4603

Aberdeen Fungal GroupOsmotic stress adaptation in C. albicans – the solution

Low granularity ODE models that reflect scarcity of information in C. albicans

The models capture essential features of the system, abstracting:Upstream signalling to Hog1Glycerol production machinery and its regulation Tao You, Piers Ingram

Aberdeen Fungal GroupModeling osmotic stress adaptation in C. albicansp

These low granularity models h h l d t

1.0

aCl (

M)

have helped us to understand global features of the system:

0Na

Integral control of glycerol

The existence of memory in the system

Integral control of glycerol production

Tao You, Mette Jacobsen

Aberdeen Fungal GroupModeling osmotic stress adaptation in C. albicansp

These low granularity models do NOT help us to understand the t ib ti f ifi l l t t t b h icontributions of specific molecular components to system behaviours




( )ASPIRATIONS:









( )ASPIRATIONS:

I d i i f

Michelle Leach @ 4:35pm on ThursdayQuantitative (ODE)Stochastic …


Heat Shock:Michelle LeachKatarzyna Tyc… contributions of specific components


y yEdda KlippLeah Cowen




Aberdeen Fungal GroupThe heat shock response in Candida albicans

Th H t Sh k T i ti F t (H f1) i ti t d bThe Heat Shock Transcription Factor (Hsf1) is activated by phosphorylation during heat shock Why is Hsf1 not phosphorylated during growth at elevated

Temperature (˚C)

Why is Hsf1 not phosphorylated during growth at elevated temperatures?

15 20 25 3530 40 HSp ( )

Hsf1Hsf1-P

Act1

Nicholls, Leach, et al. (2009) Molec. Micro. 74, 844.

Aberdeen Fungal GroupModeling the heat shock response in Candida albicans

Hsf1heat

HSP90

heatshock

autoregulatorycircuit

Hsp90 unfoldedproteins

Michelle Leach & Katarzyna Tyc

Aberdeen Fungal GroupThe heat shock system displays“Perfect Adaptation”p

simulations experimental datasimulations

P

experimental data

30-42˚C in tet-HSP90 knockdown

Hsf

1 P

- + - + - + - +- +Ppase

0 1 3 42 hrs

30 42 C in tet HSP90 knockdown

Time (min)

30-42˚C+ + + ++

Hsf1Hsf1-P

Hsf1 is activated by phosphorylation during heat shock But this activation is transient, Hsf1-P returning to basal levels

e ( )

, gonce cells have adapted to the elevated temperature“Perfect Adaptation” is dependent upon a negative feedback loopThis explains why Hsf1 is not phosphorylated during

Michelle Leach & Katarzyna Tyc

This explains why Hsf1 is not phosphorylated during protracted growth at elevated temperatures




( )ASPIRATIONS:









( )ASPIRATIONS:






Thanks to …Neil Gow


Neil GowFrank OddsGordon BrownCarol Munro

AFG

Carol MunroAlex BrandDonna MacCallum

GCN System:Rosa De Llanos Tao YouLaura Selway Priya Chandrasekaran

Heat Shock:Michelle LeachKatarzyna TycLaura Selway Priya Chandrasekaran

Helene Tournu George Coghill

Osmotic Stress:

Katarzyna TycEdda KlippLeah Cowen

Osmotic Stress:Despoina Kaloriti Tao YouMette Jacobsen Piers IngramAnna Tillmann Wei Pang Janet Q inngNeil Gow Mamen RomanoKen Haynes Marco ThielEmily Cook Alessandro Moura

Janet Quinn

Melanie Putnam Celso GrebogiAndy McDonagh

Documents

Candida albicansCandida albicans pathogenesis and systems ... · Candida albicans enjoys differing relationships with its human host … Aberdeen Fungal Group commensalism systemic