Structure and Assembly Pathway of the Ribosome Quality Control Complex

Summary During ribosome-associated quality control, stalled ribosomes are split into subunits and the 60S-housed nascent polypeptides are poly-ubiquitinated by Listerin. How this low-abundance ubiquitin ligase targets rare stall-generated 60S among numerous empty 60S is unknown. Here, we show that Listerin specificity for nascent chain-60S complexes depends on nuclear export mediator factor (NEMF). The 3.6 Å cryo-EM structure of a nascent chain-containing 60S-Listerin-NEMF complex revealed that NEMF makes multiple simultaneous contacts with 60S and peptidyl-tRNA to sense nascent chain occupancy. Structural and mutational analyses showed that ribosome-bound NEMF recruits and stabilizes Listerin’s N-terminal domain, while Listerin’s C-terminal RWD domain directly contacts the ribosome to position the adjacent ligase domain near the nascent polypeptide exit tunnel. Thus, highly specific nascent chain targeting by Listerin is imparted by the avidity gained from a multivalent network of context-specific individually weak interactions, highlighting a new principle of client recognition during protein quality control.

[1]  K. Shirahige,et al.  Receptor for activated C kinase 1 stimulates nascent polypeptide‐dependent translation arrest , 2010, EMBO reports.

[2]  F. Netter,et al.  Supplemental References , 2002, We Came Naked and Barefoot.

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

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

[5]  Deborah A Hursh,et al.  Drosophila caliban, a nuclear export mediator, can function as a tumor suppressor in human lung cancer cells , 2005, Oncogene.

[6]  Wen Jiang,et al.  EMAN2: an extensible image processing suite for electron microscopy. , 2007, Journal of structural biology.

[7]  R. Hegde,et al.  Protein Targeting and Degradation are Coupled for Elimination of Mislocalized Proteins , 2011, Nature.

[8]  R. Green,et al.  Dom34:Hbs1 Promotes Subunit Dissociation and Peptidyl-tRNA Drop-Off to Initiate No-Go Decay , 2010, Science.

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

[10]  A. Sali,et al.  Architecture of the Protein-Conducting Channel Associated with the Translating 80S Ribosome , 2001, Cell.

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

[12]  R. Hegde,et al.  Design principles of protein biosynthesis-coupled quality control. , 2012, Developmental cell.

[13]  T. Inada,et al.  Nascent Peptide-dependent Translation Arrest Leads to Not4p-mediated Protein Degradation by the Proteasome* , 2009, Journal of Biological Chemistry.

[14]  Alan Brown,et al.  Tools for macromolecular model building and refinement into electron cryo-microscopy reconstructions , 2015, Acta crystallographica. Section D, Biological crystallography.

[15]  S. Scheres,et al.  Ribosome structures to near-atomic resolution from thirty thousand cryo-EM particles , 2013, eLife.

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

[17]  T. Liehr,et al.  Assignment1 of the serologically defined colon cancer antigen 1 gene (SDCCAG1) to human chromosome band 14q22 by in situ hybridization , 1999, Cytogenetic and Genome Research.

[18]  Yang Zhang,et al.  I-TASSER server for protein 3D structure prediction , 2008, BMC Bioinformatics.

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

[20]  E. Bennett,et al.  Protecting the proteome: Eukaryotic cotranslational quality control pathways , 2014, The Journal of cell biology.

[21]  C. Hellen,et al.  Dissociation by Pelota, Hbs1 and ABCE1 of mammalian vacant 80S ribosomes and stalled elongation complexes , 2011, The EMBO journal.

[22]  Conrad C. Huang,et al.  UCSF Chimera—A visualization system for exploratory research and analysis , 2004, J. Comput. Chem..

[23]  Israel S. Fernández,et al.  Structure of the Mammalian Ribosome-Sec61 Complex to 3.4 Å Resolution , 2014, Cell.

[24]  S. Rospert,et al.  Release Factor eRF3 Mediates Premature Translation Termination on Polylysine-Stalled Ribosomes in Saccharomyces cerevisiae , 2014, Molecular and Cellular Biology.

[25]  R. Hegde,et al.  In vitro dissection of protein translocation into the mammalian endoplasmic reticulum. , 2010, Methods in molecular biology.

[26]  Daniel N. Wilson,et al.  Structures of the human and Drosophila 80S ribosome , 2013, Nature.

[27]  J. Bonifacino,et al.  Deubiquitinases Sharpen Substrate Discrimination during Membrane Protein Degradation from the ER , 2013, Cell.

[28]  M. Wiedmann,et al.  Polypeptide‐binding proteins mediate completion of co‐translational protein translocation into the mammalian endoplasmic reticulum , 2003, EMBO reports.

[29]  M. Topf,et al.  Mechanism of eIF6-mediated Inhibition of Ribosomal Subunit Joining* , 2010, The Journal of Biological Chemistry.

[30]  M. Pool,et al.  Distinct Modes of Signal Recognition Particle Interaction with the Ribosome , 2002, Science.

[31]  Alan Brown,et al.  Structure of the Yeast Mitochondrial Large Ribosomal Subunit , 2014, Science.

[32]  Sjors H.W. Scheres,et al.  RELION: Implementation of a Bayesian approach to cryo-EM structure determination , 2012, Journal of structural biology.

[33]  J. Weissman,et al.  Differential Scales of Protein Quality Control , 2014, Cell.

[34]  R. Hegde,et al.  Quality and quantity control at the endoplasmic reticulum. , 2010, Current opinion in cell biology.

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

[36]  P. Emsley,et al.  Features and development of Coot , 2010, Acta crystallographica. Section D, Biological crystallography.

[37]  Roy Parker,et al.  Exosome-Mediated Recognition and Degradation of mRNAs Lacking a Termination Codon , 2002, Science.

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

[40]  R. Hegde,et al.  Identification of a Targeting Factor for Posttranslational Membrane Protein Insertion into the ER , 2007, Cell.

[41]  Vincent B. Chen,et al.  Correspondence e-mail: , 2000 .

[42]  Dmitry Lyumkis,et al.  Structural basis for translational surveillance by the large ribosomal subunit-associated protein quality control complex , 2014, Proceedings of the National Academy of Sciences.

[43]  L. Aravind,et al.  A highly conserved family of domains related to the DNA-glycosylase fold helps predict multiple novel pathways for RNA modifications , 2014, RNA biology.

[44]  R. Green,et al.  Translation drives mRNA quality control , 2012, Nature Structural &Molecular Biology.

[45]  N. Ban,et al.  L23 protein functions as a chaperone docking site on the ribosome , 2002, Nature.

[46]  Dmitry Lyumkis,et al.  Single-particle EM reveals extensive conformational variability of the Ltn1 E3 ligase , 2013, Proceedings of the National Academy of Sciences.