GIGYF2 and 4EHP Inhibit Translation Initiation of Defective Messenger RNAs to Assist Ribosome-Associated Quality Control

Ribosome-associated Quality Control (RQC) pathways protect cells from toxicity caused by incomplete protein products resulting from translation of damaged or problematic mRNAs. Extensive work in yeast has identified highly conserved mechanisms that lead to the degradation of the faulty mRNA and partially synthesized polypeptide. Here, we used CRISPR-Cas9-based screening to search for additional RQC strategies in mammals. We found that failed translation leads to specific silencing of translation initiation on that message. This negative feedback loop is mediated by two translation inhibitors, GIGYF2 and 4EHP, in part via the ribosome collision sensor ZNF598. Both model substrates and growth-based assays established that inhibition of additional rounds of translation acts in concert with known RQC pathways to prevent buildup of toxic proteins. Inability to block translation of faulty mRNAs, and subsequent accumulation of partially synthesized polypeptides, could explain the neurodevelopmental and neuropsychiatric disorders observed in mice and humans with compromised GIGYF2 function.

[1]  T. Inada,et al.  Identification of a novel trigger complex that facilitates ribosome-associated quality control in mammalian cells , 2020, Scientific Reports.

[2]  R. Parker,et al.  Coupling of translation quality control and mRNA targeting to stress granules , 2020, bioRxiv.

[3]  Thomas M. Norman,et al.  Titrating gene expression using libraries of systematically attenuated CRISPR guide RNAs , 2019, Nature Biotechnology.

[4]  Bo Huang,et al.  Deep profiling reveals substantial heterogeneity of integration outcomes in CRISPR knock-in experiments , 2019, bioRxiv.

[5]  Grant W. Brown,et al.  The endonuclease Cue2 cleaves mRNAs at stalled ribosomes during No Go Decay , 2019, bioRxiv.

[6]  S. Anders,et al.  Alanine Tails Signal Proteolysis in Bacterial Ribosome-Associated Quality Control , 2019, Cell.

[7]  J. Olsen,et al.  GIGYF1/2-Driven Cooperation between ZNF598 and TTP in Posttranscriptional Regulation of Inflammatory Signaling. , 2019, Cell reports.

[8]  R. Hegde,et al.  ZNF598 Is a Quality Control Sensor of Collided Ribosomes , 2018, Molecular cell.

[9]  Summer B. Thyme,et al.  Phenotypic Landscape of Schizophrenia-Associated Genes Defines Candidates and Their Shared Functions , 2018, Cell.

[10]  I. MacRae,et al.  Phase Transitions in the Assembly and Function of Human miRISC , 2018, Cell.

[11]  Nicolas Mandel,et al.  4EHP-independent repression of endogenous mRNAs by the RNA-binding protein GIGYF2 , 2018, Nucleic acids research.

[12]  Hani S. Zaher,et al.  Ribosome Collision Is Critical for Quality Control during No-Go Decay. , 2017, Molecular cell.

[13]  Nicholas T Ingolia,et al.  Transcriptome-wide measurement of translation by ribosome profiling. , 2017, Methods.

[14]  Nicholas T. Ingolia,et al.  Ubiquitination of stalled ribosome triggers ribosome-associated quality control , 2017, Nature Communications.

[15]  Jeffrey A. Hussmann,et al.  CAT-tailing as a fail-safe mechanism for efficient degradation of stalled nascent polypeptides , 2017, Science.

[16]  D. Weinberg,et al.  In vitro analysis of RQC activities provides insights into the mechanism and function of CAT tailing , 2017, eLife.

[17]  T. Tuschl,et al.  The E3 ubiquitin ligase and RNA-binding protein ZNF598 orchestrates ribosome quality control of premature polyadenylated mRNAs , 2017, Nature Communications.

[18]  E. Valkov,et al.  GIGYF1/2 proteins use auxiliary sequences to selectively bind to 4EHP and repress target mRNA expression , 2017, Genes & development.

[19]  Raymond H. Mak,et al.  ZNF598 and RACK1 Regulate Mammalian Ribosome-Associated Quality Control Function by Mediating Regulatory 40S Ribosomal Ubiquitylation. , 2017, Molecular cell.

[20]  R. Hegde,et al.  Initiation of Quality Control during Poly(A) Translation Requires Site-Specific Ribosome Ubiquitination , 2017, Molecular cell.

[21]  Max A. Horlbeck,et al.  Compact and highly active next-generation libraries for CRISPR-mediated gene repression and activation , 2016, eLife.

[22]  Max A. Horlbeck,et al.  Author response: Compact and highly active next-generation libraries for CRISPR-mediated gene repression and activation , 2016 .

[23]  F. Dautry,et al.  Post-transcriptional gene silencing activity of human GIGYF2. , 2016, Biochemical and biophysical research communications.

[24]  Marco Y. Hein,et al.  The Perseus computational platform for comprehensive analysis of (prote)omics data , 2016, Nature Methods.

[25]  Bo Huang,et al.  A scalable strategy for high-throughput GFP tagging of endogenous human proteins , 2016, Proceedings of the National Academy of Sciences.

[26]  J. Yates,et al.  The Rqc2/Tae2 subunit of the ribosome-associated quality control (RQC) complex marks ribosome-stalled nascent polypeptide chains for aggregation , 2016, eLife.

[27]  Kristofor J. Webb,et al.  Recruitment of the 4EHP-GYF2 cap-binding complex to tetraproline motifs of tristetraprolin promotes repression and degradation of mRNAs with AU-rich elements , 2016, RNA.

[28]  F. Hartl,et al.  Failure of RQC machinery causes protein aggregation and proteotoxic stress , 2016, Nature.

[29]  J. Yates,et al.  Author response: The Rqc2/Tae2 subunit of the ribosome-associated quality control (RQC) complex marks ribosome-stalled nascent polypeptide chains for aggregation , 2016 .

[30]  Kali T. Witherspoon,et al.  Excess of rare, inherited truncating mutations in autism , 2015, Nature Genetics.

[31]  Koichi Ito,et al.  Inhibiting K63 Polyubiquitination Abolishes No-Go Type Stalled Translation Surveillance in Saccharomyces cerevisiae , 2015, PLoS genetics.

[32]  Alan Brown,et al.  Structure and Assembly Pathway of the Ribosome Quality Control Complex , 2015, Molecular cell.

[33]  Max A. Horlbeck,et al.  Genome-Scale CRISPR-Mediated Control of Gene Repression and Activation , 2014, Cell.

[34]  R. Hegde,et al.  Reconstitution of a Minimal Ribosome-Associated Ubiquitination Pathway with Purified Factors , 2014, Molecular cell.

[35]  David A. Scott,et al.  Genome engineering using the CRISPR-Cas9 system , 2013, Nature Protocols.

[36]  E. Grayhack,et al.  Translation of CGA codon repeats in yeast involves quality control components and ribosomal protein L1 , 2013, RNA.

[37]  R. Hegde,et al.  Listerin-Dependent Nascent Protein Ubiquitination Relies on Ribosome Subunit Dissociation , 2013, Molecular cell.

[38]  M. Fromont-Racine,et al.  Cdc48-associated complex bound to 60S particles is required for the clearance of aberrant translation products , 2013, Proceedings of the National Academy of Sciences.

[39]  Luke A. Gilbert,et al.  Repurposing CRISPR as an RNA-Guided Platform for Sequence-Specific Control of Gene Expression , 2013, Cell.

[40]  R. Deshaies,et al.  Cdc48/p97 promotes degradation of aberrant nascent polypeptides bound to the ribosome , 2013, eLife.

[41]  Adam Frost,et al.  A Ribosome-Bound Quality Control Complex Triggers Degradation of Nascent Peptides and Signals Translation Stress , 2012, Cell.

[42]  Phillip A Sharp,et al.  A triple helix stabilizes the 3' ends of long noncoding RNAs that lack poly(A) tails. , 2012, Genes & development.

[43]  M. Ares.  Isolation of total RNA from yeast cell cultures. , 2012, Cold Spring Harbor protocols.

[44]  T. Inada,et al.  Roles of dom34:hbs1 in nonstop protein clearance from translocators for normal organelle protein influx. , 2012, Cell reports.

[45]  A. van Hoof,et al.  Degradation of mRNAs that lack a stop codon: a decade of nonstop progress , 2012, Wiley interdisciplinary reviews. RNA.

[46]  A. Gingras,et al.  A Novel 4EHP-GIGYF2 Translational Repressor Complex Is Essential for Mammalian Development , 2012, Molecular and Cellular Biology.

[47]  M. Mann,et al.  Andromeda: a peptide search engine integrated into the MaxQuant environment. , 2011, Journal of proteome research.

[48]  C. Joazeiro,et al.  Role of a ribosome-associated E3 ubiquitin ligase in protein quality control , 2010, Nature.

[49]  Michael Kofler,et al.  Conserved beta-hairpin recognition by the GYF domains of Smy2 and GIGYF2 in mRNA surveillance and vesicular transport complexes. , 2010, Structure.

[50]  G. Stoecklin,et al.  Human Pat1b Connects Deadenylation with mRNA Decapping and Controls the Assembly of Processing Bodies , 2010, Molecular and Cellular Biology.

[51]  Brian V. Jenkins,et al.  A mouse forward genetics screen identifies LISTERIN as an E3 ubiquitin ligase involved in neurodegeneration , 2009, Proceedings of the National Academy of Sciences.

[52]  M. Mann,et al.  MaxQuant enables high peptide identification rates, individualized p.p.b.-range mass accuracies and proteome-wide protein quantification , 2008, Nature Biotechnology.

[53]  J. Pelletier,et al.  Translation of nonSTOP mRNA is repressed post‐initiation in mammalian cells , 2007, The EMBO journal.

[54]  N. Sonenberg,et al.  Weak binding affinity of human 4EHP for mRNA cap analogs. , 2007, RNA.

[55]  T. Inada,et al.  Translation of the poly(A) tail plays crucial roles in nonstop mRNA surveillance via translation repression and protein destabilization by proteasome in yeast. , 2007, Genes & development.

[56]  R. Parker,et al.  Endonucleolytic cleavage of eukaryotic mRNAs with stalls in translation elongation , 2006, Nature.

[57]  T. Inada,et al.  Translation of aberrant mRNAs lacking a termination codon or with a shortened 3′‐UTR is repressed after initiation in yeast , 2005, The EMBO journal.

[58]  Kathryn A. O’Donnell,et al.  An mRNA Surveillance Mechanism That Eliminates Transcripts Lacking Termination Codons , 2002, Science.

[59]  R. Singer,et al.  Localization of ASH1 mRNA particles in living yeast. , 1998, Molecular cell.

[60]  A. Gingras,et al.  Cloning and Characterization of 4EHP, a Novel Mammalian eIF4E-related Cap-binding Protein* , 1998, The Journal of Biological Chemistry.

[61]  E. Myers,et al.  Basic local alignment search tool. , 1990, Journal of molecular biology.

[62]  T. Inada,et al.  Collided ribosomes form a unique structural interface to induce Hel2‐driven quality control pathways , 2019, The EMBO journal.

[63]  R. Rothstein.  Targeting, disruption, replacement, and allele rescue: integrative DNA transformation in yeast. , 1991, Methods in enzymology.