Architecture of the Protein-Conducting Channel Associated with the Translating 80S Ribosome

Beckmann, Roland; Spahn, C.M.T.; Eswar, Narayanan; Helmers, Jürgen; Penczek, Pawel A.; Šali, Andrej; Frank, Joachim; Blobel, Günter

doi:10.1142/9789813234864_0026

Cited by 10 publications

(15 citation statements)

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“…Owing to their dynamic behavior, long ribosomal RNA expansion segments are typically not resolved in high-resolution cryo-EM reconstructions, with few exceptions in which their conformational landscape is confined by specific interactions with either the ribosomal core or ribosome-associated factors. For one of the longest eukaryotic rRNA ESs, ES27L, two distinct conformations were described 23,24 : ES27L either aligns with the ribosomal 40S/60S subunit interface ("L1-position"; "ES27L-L1") or projects toward the ribosomal tunnel exit ("exit-position"; "ES27L-exit") ( Supplementary Fig. 6).…”

Section: Resultsmentioning

confidence: 99%

MetAP-like Ebp1 occupies the human ribosomal tunnel exit and recruits flexible rRNA expansion segments

et al. 2020

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Human Ebp1 is a member of the proliferation-associated 2G4 (PA2G4) family and plays an important role in cancer regulation. Ebp1 shares the methionine aminopeptidase (MetAP) fold and binds to mature 80S ribosomes for translational control. Here, we present a cryo-EM single particle analysis reconstruction of Ebp1 bound to non-translating human 80S ribosomes at a resolution range from 3.3 to~8 Å. Ebp1 blocks the tunnel exit with major interactions to the general uL23/uL29 docking site for nascent chain-associated factors complemented by eukaryote-specific eL19 and rRNA helix H59. H59 is defined as dynamic adaptor undergoing significant remodeling upon Ebp1 binding. Ebp1 recruits rRNA expansion segment ES27L to the tunnel exit via specific interactions with rRNA consensus sequences. The Ebp1-ribosome complex serves as a template for MetAP binding and provides insights into the structural principles for spatial coordination of co-translational events and molecular triage at the ribosomal tunnel exit.

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Section: Resultsmentioning

confidence: 99%

MetAP-like Ebp1 occupies the human ribosomal tunnel exit and recruits flexible rRNA expansion segments

et al. 2020

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“…The interface involves mainly the small but to a lesser extent also the large subunit and involves many eukaryote-specific ribosomal proteins and rRNA expansion segments. ES27L is a flexible rRNA expansion segment capable of adopting two main conformations in the yeast 80S, one facing towards the peptide exit tunnel (ES27 exit) and the other one facing towards the flexible L1 stalk (ES27-L1, (Beckmann et al, 2001). Notably, major conformational changes occur in rRNA expansion segments (ES) upon disome formation in both ribosomes compared to the 80S reconstructions derived from the individual 80S ribosomes (Appendix Fig S4).…”

Section: Structural Analysis Of An Ngd and Rqc Substrate Disome Unitmentioning

confidence: 99%

Collided ribosomes form a unique structural interface to induce Hel2‐driven quality control pathways

et al. 2019

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Ribosome stalling triggers quality control pathways targeting the mRNA (NGD: no‐go decay) and the nascent polypeptide (RQC: ribosome‐associated quality control). RQC requires Hel2‐dependent uS10 ubiquitination and the RQT complex in yeast. Here, we report that Hel2‐dependent uS10 ubiquitination and Slh1/Rqt2 are crucial for RQC and NGD induction within a di‐ribosome (disome) unit, which consists of the leading stalled ribosome and the following colliding ribosome. Hel2 preferentially ubiquitinated a disome over a monosome on a quality control inducing reporter mRNA in an in vitro translation reaction. Cryo‐EM analysis of the disome unit revealed a distinct structural arrangement suitable for recognition and modification by Hel2. The absence of the RQT complex or uS10 ubiquitination resulted in the elimination of NGD within the disome unit. Instead, we observed Hel2‐mediated cleavages upstream of the disome, governed by initial Not4‐mediated monoubiquitination of eS7 and followed by Hel2‐mediated K63‐linked polyubiquitination. We propose that Hel2‐mediated ribosome ubiquitination is required both for canonical NGD (NGDRQC+) and RQC coupled to the disome and that RQC‐uncoupled NGD outside the disome (NGDRQC−) can occur in a Not4‐dependent manner.

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“…Next, both the translating ribosome and the nascent precuror polypeptide chain are handed‐over to the so‐called protein‐translocon in the ER membrane and translation is allowed to pick up speed. In the course of this hand‐over, the signal peptide – with the help of the ribosome – initiates insertion of the nascent precursor polypeptide into the ER membrane, or – more precisely – into the ER‐membrane resident and heterotrimeric Sec61 complex, which forms an aqueous polypeptide‐conducting channel in its fully open state (Table ) . Ideally, signal peptides thus facilitate not only ER targeting but also full Sec61‐channel opening.…”

Section: Protein Transport Components/complexes In Hela Cells Alternmentioning

confidence: 99%

ER import of small human presecretory proteins: components and mechanisms

et al. 2019

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Protein transport into the mammalian endoplasmic reticulum (ER) used to be seen as strictly cotranslational, that is temporarily and mechanistically coupled to protein synthesis. In the course of the last decades, however, several classes of precursors of soluble and membrane proteins were found to be post‐translationally imported into the ER, without any involvement of the ribosome. The first such class to be identified were the small presecretory proteins; tail‐anchored membrane proteins followed next. In both classes, the inherent address tag is released from the translating ribosome before the initiation of ER import, as part of the fully synthesized precursor. In small presecretory proteins, the information for ER targeting and ‐translocation via the polypeptide‐conducting Sec61‐channel is encoded by a classical N‐terminal signal peptide, which is released from the ribsosome before targeting due to the small size of the full‐length precursor. Here, we discuss the current state of research on targeting and translocation of small presecretory proteins into the mammalian ER. In closing, we present a unifying hypothesis for ER protein translocation in terms of an energy diagram for Sec61‐channel gating.

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Architecture of the Protein-Conducting Channel Associated with the Translating 80S Ribosome

Cited by 10 publications

References 0 publications

MetAP-like Ebp1 occupies the human ribosomal tunnel exit and recruits flexible rRNA expansion segments

MetAP-like Ebp1 occupies the human ribosomal tunnel exit and recruits flexible rRNA expansion segments

Collided ribosomes form a unique structural interface to induce Hel2‐driven quality control pathways

ER import of small human presecretory proteins: components and mechanisms

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