CAT tails drive degradation of stalled polypeptides on and off the ribosome

Stalled translation produces incomplete, ribosome-tethered polypeptides that the Ribosome-associated Quality Control (RQC) pathway targets for degradation via the E3 ubiquitin ligase Ltn1. During this process, the protein Rqc2 and the large ribosomal subunit elongate stalled polypeptides with carboxy-terminal alanine and threonine residues (CAT tails). Failure to degrade CAT-tailed proteins disrupts global protein homeostasis, as CAT-tailed proteins can aggregate and sequester chaperones. Why cells employ such a potentially toxic process during RQC is unclear. Here, we developed quantitative techniques to assess how CAT tails affect stalled polypeptide degradation in Saccharomyces cerevisiae. We found that CAT tails enhance Ltn1’s efficiency in targeting structured polypeptides, which are otherwise poor Ltn1 substrates. If Ltn1 fails to ubiquitylate those stalled polypeptides or becomes limiting, CAT tails act as degrons, marking proteins for proteasomal degradation off the ribosome. Thus, CAT tails functionalize the carboxy-termini of stalled polypeptides to drive their degradation on and off the ribosome.