Aggregation of CAT tails blocks their degradation and causes proteotoxicity in S. cerevisiae

Abstract

The Ribosome-associated Quality Control (RQC) pathway co-translationally marks incomplete polypeptides from stalled translation with two signals that trigger their proteasomemediated degradation. The E3 ligase Ltn1 adds ubiquitin and Rqc2 directs the large ribosomal subunit to append carboxy-terminal alanine and threonine residues (CAT tails). When excessive amounts of incomplete polypeptides evade Ltn1, CAT-tailed proteins accumulate and can self-associate into aggregates. CAT tail aggregation has been hypothesized to either protect cells by sequestering potentially toxic incomplete polypeptides or harm cells by disrupting protein homeostasis. To distinguish between these possibilities, we modulated CAT tail aggregation in Saccharomyces cerevisiae with genetic and chemical tools to analyze CAT tails in aggregated and un-aggregated states. We found that enhancing CAT tail aggregation induces proteotoxic stress and antagonizes degradation of CAT-tailed proteins, while inhibiting aggregation reverses these effects. Our findings suggest that CAT tail aggregation harms RQC-compromised cells and that preventing aggregation can mitigate this toxicity. OPEN ACCESS Citation: Sitron CS, Park JH, Giafaglione JM, Brandman O (2020) Aggregation of CAT tails blocks their degradation and causes proteotoxicity in S. cerevisiae. PLoS ONE 15(1): e0227841.

The discovery and consequences of the central role of the nervous system in the control of protein homeostasis

“…These aborted polypeptides are recognized by the cellular surveillance machinery and modified by the addition of carboxyterminal alanine and threonine residues (CAT tails) (Brandman et al, 2012; Kostova et al, 2017; Sitron, Park, & Brandman, 2017; Sitron, Park, Giafaglione, & Brandman, 2020; Sitron & Brandman, 2019). This, in turn, leads to their aggregation and targeted degradation through the sequestration of a group of specialized proteins called molecular chaperones that recognize the aberrant conformation of the CAT-tailed polypeptides (Sitron et al, 2017(Sitron et al, , 2020Sitron & Brandman, 2019). The sequestration of molecular chaperones is also thought to activate a conserved transcription factor to increase the amounts of molecular chaperone proteins available to surveil and prevent the build-up of aberrant polypeptides.…”

The discovery and consequences of the central role of the nervous system in the control of protein homeostasis

“…For instance, ribosome stalling that can occur due to stochastic perturbations of translation, mutations, or a number of biotic or abiotic fluctuations in the environment will lead to the aborted synthesis of a nascent polypeptide chains. These aborted polypeptides are recognized by the cellular surveillance machinery and modified by the addition of carboxy-terminal alanine and threonine residues (CAT tails) (Brandman et al, 2012;Kostova et al, 2017;Sitron, Park, & Brandman, 2017;Sitron, Park, Giafaglione, & Brandman, 2020;Sitron & Brandman, 2019). This, in turn, leads to their aggregation and targeted degradation through the sequestration of a group of specialized proteins called molecular chaperones that recognize the aberrant conformation of the CAT-tailed polypeptides (Sitron et al, 2017(Sitron et al, , 2020Sitron & Brandman, 2019).…”

“…These aborted polypeptides are recognized by the cellular surveillance machinery and modified by the addition of carboxy-terminal alanine and threonine residues (CAT tails) (Brandman et al, 2012;Kostova et al, 2017;Sitron, Park, & Brandman, 2017;Sitron, Park, Giafaglione, & Brandman, 2020;Sitron & Brandman, 2019). This, in turn, leads to their aggregation and targeted degradation through the sequestration of a group of specialized proteins called molecular chaperones that recognize the aberrant conformation of the CAT-tailed polypeptides (Sitron et al, 2017(Sitron et al, , 2020Sitron & Brandman, 2019). The sequestration of molecular chaperones is also thought to activate a conserved transcription factor to increase the amounts of molecular chaperone proteins available to surveil and prevent the build-up of aberrant polypeptides.…”

Cytosolic Quality Control of Mitochondrial Protein Precursors—The Early Stages of the Organelle Biogenesis

“…The 60S subunit with stalled peptide is then recognized by ribosome quality control complex subunit 2 (Rqc2 in yeast, NEMF in human), which helps recruit E3 ubiquitin ligase Ltn1 (Listerin). In turn, Ltn1/Listerin ubiquitinate stalled substrates, providing the signal for extraction and degradation [ 219 , 220 ]. In addition to Ltn1 recruitment, Rqc2 runs a non-canonical synthesis reaction that extends the C-terminus of the stalled peptide by adding alanine and threonine residues—a process called CAT-tailing.…”

“…In addition to Ltn1 recruitment, Rqc2 runs a non-canonical synthesis reaction that extends the C-terminus of the stalled peptide by adding alanine and threonine residues—a process called CAT-tailing. Incorporating CAT-tails can further facilitate the ubiquitination of stalled substrates by Ltn1/Listerin [ 219 , 220 ]. The ubiquitinated substrates are removed from the 60S ribosome by Cdc48 and directed for proteasomal degradation.…”