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Review
. 2019 Jun;20(6):368-383.
doi: 10.1038/s41580-019-0118-2.

Mechanisms and functions of ribosome-associated protein quality control

Claudio A P Joazeiro  1   2
Affiliations
Review

Mechanisms and functions of ribosome-associated protein quality control

Claudio A P Joazeiro. Nat Rev Mol Cell Biol. 2019 Jun.

Abstract

The stalling of ribosomes during protein synthesis results in the production of truncated polypeptides that can have deleterious effects on cells and therefore must be eliminated. In eukaryotes, this function is carried out by a dedicated surveillance mechanism known as ribosome-associated protein quality control (RQC). The E3 ubiquitin ligase Ltn1 (listerin in mammals) plays a key part in RQC by targeting the aberrant nascent polypeptides for proteasomal degradation. Consistent with having an important protein quality control function, mutations in listerin cause neurodegeneration in mice. Ltn1/listerin is part of the multisubunit RQC complex, and recent findings have revealed that the Rqc2 subunit of this complex catalyses the formation of carboxy-terminal alanine and threonine tails (CAT tails), which are extensions of nascent chains known to either facilitate substrate ubiquitylation and targeting for degradation or induce protein aggregation. RQC, originally described for quality control on ribosomes translating cytosolic proteins, is now known to also have a role on the surfaces of the endoplasmic reticulum and mitochondria. This Review describes our current knowledge on RQC mechanisms, highlighting key features of Ltn1/listerin action that provide a paradigm for understanding how E3 ligases operate in protein quality control in general, and discusses how defects in this pathway may compromise cellular function and lead to disease.

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Figures

Fig. 1:
Fig. 1:. The eukaryotic ribosome-associated quality control pathway
The ribosomal rescue and Ribosome-Associated Quality Control (RQC) pathways for protein surveillance consist of two sequential steps that each sense a unique defect. The first step (rescue) senses stalled ribosomes and mediates 80S subunit splitting; the second step (RQC) detects 60S subunits obstructed with peptidyl-tRNA—products of the rescue step—and promotes the resolution of this aberrant structure, to release free, translation-competent 60S ribosomal subunits and mediate proteolysis of the nascent chain. (Left) Ribosomal rescue. Ribosomes that are stalled at the mRNA 3’ end are sensed by Hbs1 in yeast (or HBS1L and GTPBP2 in mammals) and Dom34 (pelota (PELO) in mammals). The mechanisms for the recognition of ribosomes stalled on internal mRNA sequences are poorly understood and at least in some cases involve the E3 ubiquitin-protein ligase Hel2/ZNF598. Whether Dom34/PELO also acts downstream of Hel2/ZNF598 is not known, but these two factors can target the same mRNA molecule, as ribosome stalling elicits endonuclease cleavage of the transcript. Dom34/PELO recruits Rli1 (ATP-binding cassette protein ABCE1 in mammals), which separates the small (40S) and large (60S) ribosomal subunits (Box 1). The released truncated mRNA is degraded by the 5’−3’ exoribonuclease Xrn1 and the exosome complex, which prevents the mRNA from being translated again, as it may be defective. However, unlike eRF1, Dom34 lacks peptidyl-tRNA hydrolase activity, the peptidyl-tRNA remains bound to the 60S subunit. Although the peptidyl-tRNA−60S complex can potentially reassociate with free 40S subunits, the reassociation can be reversed by Dom34, Hbs1 and Rli1,,. (Right) RQC. Resolution of the peptidyl-tRNA−60S complex is initiated by binding of the Rqc2 (NEMF in mammals) subunit of the RQC complex, which recruits and stabilizes the binding of the E3 ligase Ltn1/Listerin. At this stage, Rqc2 may synthesize CAT tails to help expose Lys residues that are buried in the ribosomal exit tunnel, to be ubiquitylated by Ltn1/Listerin (see below). The ribosome binding mode of the Rqc1 subunit (TCF25 in mammals) of the RQC complex and its exact function remain unknown. The ubiquitin chain that is polymerised by Ltn1 on the nascent polypeptides signals recruitment of the AAA ATPase Cdc48 and its cofactors. Cdc48 extracts nascent polypeptides from the 60S ribosomal subunit after they have been released from the conjugated tRNA by Vms1 (ANKZF1 in mammals). Released polypeptides can then be degraded by the proteasome. The extracted nascent polypeptides may remain associated with RQC subunits in the form of a ‘light RQC’ complex before proteasomal degradation.
Fig. 2:
Fig. 2:. The function of Ltn1/Listerin in protein surveillance
(a) Structural model of the ribosomal 60S subunit obstructed with peptidyl-tRNA and bound to the ribosomal quality control (RQC) complex subunits Listerin and NEMF. Superposition of cryo-EM volume of the RQC (EMDB 2832), molecular models of NEMF and the Listerin C-terminal Domain (CTD) (PDB 3J92), and the crystal structure of the Ltn1 N-terminal Domain (NTD) (PDB 5FG1). The cartoon on the right is a representation of the 60S subunit bond to the peptidyl-tRNA, Listerin/Ltn1 and NEMF/Rqc2. The conserved N-terminal and C-terminal domains of Ltn1, separated by a variable middle linker containing HEAT/ARM repeats are indicated. The C-terminal Ltn1 RING domain recruits E2~Ub to ubiquitylate nascent chains. The dotted grey oval indicates the space that would be occupied by the 40S subunit if present, to indicate steric clashes that would occur with Rqc2/NEMF and the Ltn1/Listerin NTD. (b) Comparison of the RQC structures from yeast (EMDB 2797) and mammals (EMDB 2832). (Figures courtesy of H. Paternoga)
Fig. 3:
Fig. 3:. CAT tail synthesis and functions
(a) The ribosome-associated protein quality control (RQC) complex subunit Rqc2 recognizes ribosomal 60S subunits that are obstructed by a peptidyl-tRNA by simultaneously binding to components of the 60S subunit and the tRNA. When ubiquitylation of the nascent polypeptide by Ltn1 fails, Rqc2 can extend the trapped nascent polypeptide with a C-terminal tail composed of Ala and Thr residues (CAT tail). The reaction is mediated by the direct recruitment of charged tRNA-Ala and tRNA-Thr by Rqc2 and occurs without an mRNA template. The Vms1 protein can terminate CAT tail synthesis by releasing the P-site tRNA and presumably promoting Rqc2 dissociation from the complex. (b) Alternative fates of RQC substrates. The canonical RQC pathway of Ltn1-mediated ubiquitylation of nascent polypeptides is kinetically preferred, provided that Lys ubiquitylation sites on nascent chains are readily accessible (top). In this pathway, Rqc2 functions in recruiting and stabilizing Ltn1 in the complex. An alternative pathway takes place when ubiquitylation is compromised (middle). In this pathway, Rqc2 catalyses the elongation of a CAT tail, which can result in the exposure of Lys residues that would otherwise be hidden in the ribosomal exit tunnel. The increased accessibility to Lys residues enables ubiquitylation by Ltn1. However, when ubiquitylation fails altogether (bottom), CAT tail-modified nascent chains form aggregates, which can have different fates and effects in cellular function, including stress signalling.
Figure 4:
Figure 4:. RQC on the endoplasmic reticulum and mitochondrial membranes
Ribosomes can stall while translating proteins destined to different subcellular compartments, such as the cytosol, mitochondria and the endoplasmic reticulum (ER). (a) ER-RQC: hypothetical model for RQC at ER-associated ribosomes. On nascent chains obstructing the 60S ribosomal subunit on organellar surfaces, Ltn1 only has limited access to Lys residues, within a short segment of cytosolic-exposed polypeptide sequence immediately outside the exit tunnel. The nascent chain segments buried in the ribosomal exit tunnel or the translocon, or inside the organelle, are not accessible to ubiquitylation. However, Lys residues hidden in the ribosomal exit tunnel can be exposed to Ltn1 through Rqc2-mediated nascent chain CATylation. Cdc48 is recruited to extract the ubiquitylated nascent chain and deliver it to the proteasome for degradation in the cytosol, after Vms1 catalyses nascent chain release from the tRNA. (b) Mito-RQC: Example of Ltn1 dysfunction on the mitochondrial surface. In the absence of nascent chain ubiquitylation, nascent chains are CATylated by Rqc2 and released into the mitochondrial matrix by Vms1. The directional pulling force by a mitochondrial chaperone (in pink) is indicated. In the matrix, moderate levels of CAT tail-dependent aggregates can be handled by the protein homeostasis machinery. However, if both Ltn1- and Vms1-mediated processes fail, CATylation proceeds for an extended period, resulting in the accumulation of larger aggregates which mitochondria may have more difficulty in eliminating, and which can interfere with the function of the organelle.
Box 1
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