Specific recognition and ubiquitination of translating ribosomes by mammalian CCR4–NOT

Translation affects messenger RNA stability and, in yeast, this is mediated by the Ccr4–Not deadenylation complex. The details of this process in mammals remain unclear. Here, we use cryogenic electron microscopy (cryo-EM) and crosslinking mass spectrometry to show that mammalian CCR4–NOT specifical...

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Veröffentlicht in:	Nature structural & molecular biology Jg. 30; H. 9; S. 1314 - 1322
Hauptverfasser:	Absmeier, Eva, Chandrasekaran, Viswanathan, O’Reilly, Francis J., Stowell, James A. W., Rappsilber, Juri, Passmore, Lori A.
Format:	Journal Article
Sprache:	Englisch
Veröffentlicht:	New York Nature Publishing Group US 01.09.2023 Nature Publishing Group
Schlagworte:	631/337/1645 631/337/1645/2020 631/337/574 631/337/574/1789 631/535/1258/1259 Animals Biochemistry Biological Microscopy Biomedical and Life Sciences Conformation Crosslinking E coli Electron microscopy Elongation Humans Inserts Life Sciences Mammals Mass Spectrometry Mass spectroscopy Membrane Biology Molecular biology mRNA stability mRNA turnover Polypeptides Protein Structure Proteins Rabbits Receptors, CCR4 Ribonucleases Ribosomes Saccharomyces cerevisiae Saccharomyces cerevisiae Proteins Stability Stalling Sucrose Transcription Factors Transfer RNA Translation elongation Translations Ubiquitin Ubiquitination Yeast
ISSN:	1545-9993, 1545-9985, 1545-9985
Online-Zugang:	Volltext
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Zusammenfassung:	Translation affects messenger RNA stability and, in yeast, this is mediated by the Ccr4–Not deadenylation complex. The details of this process in mammals remain unclear. Here, we use cryogenic electron microscopy (cryo-EM) and crosslinking mass spectrometry to show that mammalian CCR4–NOT specifically recognizes ribosomes that are stalled during translation elongation in an in vitro reconstituted system with rabbit and human components. Similar to yeast, mammalian CCR4–NOT inserts a helical bundle of its CNOT3 subunit into the empty E site of the ribosome. Our cryo-EM structure shows that CNOT3 also locks the L1 stalk in an open conformation to inhibit further translation. CCR4–NOT is required for stable association of the nonconstitutive subunit CNOT4, which ubiquitinates the ribosome, likely to signal stalled translation elongation. Overall, our work shows that human CCR4–NOT not only detects but also enforces ribosomal stalling to couple translation and mRNA decay. Using cryo-EM, the authors show that the mammalian CCR4–NOT complex specifically recognizes stalled translating ribosomes similar to the yeast complex, locks them in a translation-incompetent state and coordinates their ubiquitylation, highlighting its central role in linking translation to mRNA stability.
Bibliographie:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 14 content type line 23
ISSN:	1545-9993 1545-9985 1545-9985
DOI:	10.1038/s41594-023-01075-8