The Rqc2/Tae2 subunit of the ribosome-associated quality control (RQC) complex marks ribosome-stalled nascent polypeptide chains for aggregation

Abstract

Ribosome stalling during translation can potentially be harmful, and is surveyed by a conserved quality control pathway that targets the associated mRNA and nascent polypeptide chain (NC). In this pathway, the ribosome-associated quality control (RQC) complex promotes the ubiquitylation and degradation of NCs remaining stalled in the 60S subunit. NC stalling is recognized by the Rqc2/Tae2 RQC subunit, which also stabilizes binding of the E3 ligase, Listerin/Ltn1. Additionally, Rqc2 modifies stalled NCs with a carboxy-terminal, Ala- and Thr-containing extension—the 'CAT tail'. However, the function of CAT tails and fate of CAT tail-modified ('CATylated') NCs has remained unknown. Here we show that CATylation mediates formation of detergent-insoluble NC aggregates. CATylation and aggregation of NCs could be observed either by inactivating Ltn1 or by analyzing NCs with limited ubiquitylation potential, suggesting that inefficient targeting by Ltn1 favors the Rqc2-mediated reaction. These findings uncover a translational stalling-dependent protein aggregation mechanism, and provide evidence that proteins can become specifically marked for aggregation.DOI: http://dx.doi.org/10.7554/eLife.11794.001

Structure and function of the yeast listerin (Ltn1) conserved N-terminal domain in binding to stalled 60S ribosomal subunits

“…We note that, under these conditions, we have not observed a smear running immediately above the fGFP-K12 reporter band in either WT or Ltn1-deficient extracts, which might be attributed to Rqc2-mediated C-terminal extension with alanines and threonines ("CAT tails") (21). Likewise, we have not observed high molecular weight aggregates that could have been formed in a CAT tail-dependent manner (22,23) (e.g., see Fig. 1B).…”

Asc1, Hel2, and Slh1 couple translation arrest to nascent chain degradation

“…(A) Depletion of Asc1, Hel2, and Slh1 tested the requirement of these factors in detection of a higher molecular weight smear above the stalling reporter arrest product in ltn1Δ strains without (A) and with (B) RQC2 overexpression (OE). In panel B, the arrest product band in RQC2 OE ltn1Δ appears weak because a large proportion of the reporter is in detergent-insoluble aggregates in this strain (Yonashiro et al 2016). (C) IB of Rqc2-HA and Rps2 (a small ribosomal subunit marker) shows the distribution of Rqc2 in fractions of a 10%-30% sucrose polysome gradient with and without the presence of Asc1, Hel2, and Slh1.…”

“…Cells must therefore identify defective rounds of translation and take action before the nascent chain leaves the ribosome (Brandman and Hegde 2016). Degradation of the products of stalled translation is conserved throughout Eukarya (Passos et al 2009;Shao et al 2013Shao et al , 2015Shao and Hegde 2014;Ikeuchi et al 2016), and failures in this response have been linked to dysregulation of protein homeostasis (Choe et al 2016;Defenouillere et al 2016;Yang et al 2016;Yonashiro et al 2016) and neurodegenerative phenotypes (Chu et al 2009;Ishimura et al 2014).…”

In vivo aspects of protein folding and quality control

“…Chaperones have been shown to interfere at various steps of the aggregation cascade (144), including primary nucleation, fibril elongation and fragmentation, and secondary nucleation by the fibril surface (145,146). Toxic aggregates are also produced when faulty translation products accumulate (147-149)-for example, when polypeptide chains that stall during translation on the ribosome are not efficiently cleared by ribosome quality control (RQC) machinery and the UPS (149)(150)(151).…”