Nonsense-MediatedmRNADecayBeginsWhere Translation Ends Nonsense-MediatedmRNADecayBeginsWhere Translation

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  • Nonsense-MediatedmRNADecay BeginsWhere Translation Ends

    Evangelos D. Karousis and Oliver Mühlemann

    Department of Chemistry and Biochemistry, University of Bern, CH-3012 Bern, Switzerland

    Correspondence: oliver.muehlemann@dcb.unibe.ch

    Nonsense-mediated mRNA decay (NMD) is arguably the best-studied eukaryotic messenger RNA (mRNA) surveillance pathway, yet fundamental questions concerning the molecular mechanism of target RNA selection remain unsolved. Besides degrading defective mRNAs harboring premature termination codons (PTCs), NMD also targets many mRNAs encoding functional full-length proteins. Thus, NMD impacts on a cell’s transcriptome and is implicat- ed in a range of biological processes that affect a broad spectrum of cellular homeostasis. Here, we focus on the steps involved in the recognition of NMD targets and the activation of NMD. We summarize the accumulating evidence that tightly links NMD to translation ter- mination and we further discuss the recruitment and activation of the mRNA degradation machinery and the regulation of this complex series of events. Finally, we review emerging ideas concerning themechanistic details of NMDactivation and the potential role of NMDas a general surveyor of translation efficacy.

    Originally conceived as a quality controlpathway that recognizes and specifically degrades aberrant messenger RNAs (mRNAs) with premature termination codons (PTCs), it has meanwhile become clear that nonsense- mediated mRNA decay (NMD) contributes to posttranscriptional gene regulation in a way that goes far beyond quality control. Exploring in which biological contexts NMD-mediated gene regulation plays an important role is a rel- atively new but rapidly expanding area of re- search that has been covered in recent reviews (Nasif et al. 2017; Nickless et al. 2017). Here, we summarize our current understanding regard- ing the molecular mechanism of NMD, with the focus on data obtained from mammalian sys- tems. Despite more than 25 years of research

    and a wealth of biochemical data characterizing interactions between different NMD factors, their enzymatic functions and posttranslational modifications, the mechanism and criteria for selection of an mRNA for the NMD pathway are still not well understood. The slow progress in deciphering the mechanism of NMD can at least partially be attributed to the lack of a suitable in vitro system that faithfully recapitu- lates the key steps of NMD. Nevertheless, work from many laboratories during the last few years has provided compelling evidence that NMD is tightly coupled to the process of translation termination. During translation ter- mination, it is decided whether the translated mRNA shall remain intact and serve as a tem- plate for additional rounds of translation or

    Editors: Michael B. Mathews, Nahum Sonenberg, and John W.B. Hershey Additional Perspectives on Translation Mechanisms and Control available at www.cshperspectives.org

    Copyright © 2018 Cold Spring Harbor Laboratory Press; all rights reserved Advanced Online Article. Cite this article as Cold Spring Harb Perspect Biol doi: 10.1101/cshperspect.a032862

    1

    on June 11, 2018 - Published by Cold Spring Harbor Laboratory Press http://cshperspectives.cshlp.org/Downloaded from

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  • whether it shall be degraded by the NMD path- way (He and Jacobson 2015). In a nutshell, the current view is that NMD ensues when ribo- somes at nonsense codons (hereafter called ter- mination codon [TC]) fail to terminate correct- ly. Because of the tight link between NMD and translation termination, we begin this review with a brief overview of eukaryotic translation termination. For a more detailed review of the mechanism of translation termination, see Hel- len (2018).

    THE MECHANISM OF EUKARYOTIC TRANSLATION TERMINATION AND RIBOSOME RECYCLING Translation termination is signaled by the pres- ence of one of the three TCs in the A site of the ribosome. Canonical translation termination is marked by three key events: (1) proper recogni- tion of the termination signal, (2) hydrolysis of the terminal peptidyl-tRNA bond and release of the nascent peptide, and (3) dissociation of the ribosome into its 60S and 40S subunits (Fig. 1A)

    Normal translation termination

    GDP+Pi ABCE1

    eRF1

    UAA

    UAA UAA UAA

    UAA UAA UAA UAA

    UAA

    eR F

    3

    GTP

    eRF1

    eR F

    3

    GTP

    eR F

    3

    UPF3 U P

    F 2

    UPF2

    UPF3

    UPF1UPF1 SMG6 UAAUAA

    NN CC

    SM G5

    SM G7

    UPF 1

    EJC UAA

    eR F

    3 U P

    F 3

    U P

    F2

    UPF 1

    SMG1

    SM G9

    SMG8

    SM G9

    SMG8

    Aberrant translation termination and NMD activation

    A

    B

    mRNA degradation

    Figure 1. Schematic illustration of the sequential events taking place in normal translation termination and aberrant translation termination resulting in the activation of nonsense-mediatedmRNA decay (NMD). (A) The presence of a termination codon (TC) in the A site of the ribosome provides the signal for translation termination that is marked by recognition of the TC by eukaryotic release factor (eRF)1 and eRF3, a conformational change of eRF1 that facilitates hydrolysis of the terminal peptidyl-transfer RNA (tRNA) bond and the release of the nascent peptide. Subsequently, release of the ribosome from the messenger RNA (mRNA) is facilitated by ABCE1. (B) When translation termination occurs in an unfavorable environment, UPF1 is activated by UPF2 and/or UPF3 that are free in the cytoplasm or positioned nearby on an exon junction complex (EJC). UPF1 activation involves its phosphorylation by SMG1, whose kinase activity is controlled by SMG8 and SMG9. The fate of the stalled ribosomes in the context of NMD activation remains unclear. Phosphorylated UPF1 (indicated by red dots) recruits the NMD-specific SMG6 endonuclease, which cleaves the targetedmRNAnearby the TC and the SMG5/ SMG7 heterodimer that stimulates the exonucleolytic degradation of the mRNA by recruiting the CCR4/NOT complex. For clarity, a part of the NMD-activating complex (SMG1, UPF2, and UPF3) is depicted in gray to highlight the factors triggering mRNA degradation in the lower part of the panel.

    E.D. Karousis and O. Mühlemann

    2 Advanced Online Article. Cite this article as Cold Spring Harb Perspect Biol doi: 10.1101/cshperspect.a032862

    on June 11, 2018 - Published by Cold Spring Harbor Laboratory Press http://cshperspectives.cshlp.org/Downloaded from

    http://cshperspectives.cshlp.org/

  • (Dever and Green 2012; Simms et al. 2017). In comparison to the initiation and elongation steps, the step of translation termination is less well studied and accordinglymuch less is known regarding its molecular mechanism. Instead of cognate aminoacylated (aa) transfer RNAs (tRNAs) that get recruited to the A site of the ribosome during elongation, eukaryotic release factor 1 (eRF)1 binds the A site when it harbors one of the three TCs. On terminating ribosomes, eRF1 is found as a ternary complex with the GTPase eRF3 and GTP. After its recruitment to the A site, GTP hydrolysis by eRF3 stimulates a large conformational change in eRF1 that enhances polypeptide release by engaging the active site of the ribosome. Despite the extended conformational change of the middle and car- boxy-terminal parts of eRF1, the amino-termi- nal part of the protein interacts stably with the TC throughout the process (Alkalaeva et al. 2006; Becker et al. 2012; Eyler et al. 2013; Brown et al. 2015; Shao et al. 2016).

    Following GTP hydrolysis, eRF3 dissociates from the termination complex, allowing for the subsequent interaction of eRF1 with the ABC- type ATPase ABCE1 (Rli1 in yeast), a factor that stimulates the recycling of the ribosome by split- ting the ribosomal subunits. ABCE1 contains two nucleotide-binding domains (NBDs) and a unique amino-terminal FeS cluster domain aligned by two diamagnetic [4Fe–4S]2+ clusters. ATP hydrolysis by ABCE1 causes extended con- formational changes that provide the mechani- cal force leading to the dissociation of the 60S from the 40S ribosomal subunit (Pisarev et al. 2010; Becker et al. 2012). Structural studies showed that an initial closure of the NBDs positions the FeS cluster domain toward eRF1, exerting an immediate force that destabilizes the intersubunit interactions (Heuer et al. 2017). Cross-linking and mass spectrometry ap- proaches showed that after ribosomal splitting, ABCE1 remains bound to the translational GTPase-binding site of the small ribosomal subunit, establishing major contacts with the S24e ribosomal protein mainly through NBD1 and FeS cluster domains. Structural data suggest that ABCE1 performs a tweezer-like movement that positions the FeS cluster domain in a cleft

    between S12 and rRNA on the small subunit (Kiosze-Becker et al. 2016). Additionally, ABCE1 associates with the 43S preinitiation complex (consisting of eIF1A, eIF2, eIF3, and initiator tRNA) but its role in translation initiation re- mains to be elucidated (Heuer et al. 2017). Bio- chemical data suggest thatABCE1alsoaccelerates the rate of the peptide release by eRF1 in an ATP-independentmanner, providing a function- al link between translation termination and ribo- some recycling (Shoemaker and Green 2011).

    Most of the information concerning trans- lation t