Two modes of Cue2-mediated mRNA cleavage with distinct substrate recognition initiate no-go decay

Abstract

Ribosome collisions are recognized by E3 ubiquitin ligase Hel2/ZNF598, leading to RQC (ribosome-associated quality control) and to endonucleolytic cleavage and degradation of the mRNA termed NGD (no-go decay). NGD in yeast requires the Cue2 endonuclease and occurs in two modes, either coupled to RQC (NGDRQC+) or RQC uncoupled (NGDRQC−). This is mediated by an unknown mechanism of substrate recognition by Cue2. Here, we show that the ubiquitin binding activity of Cue2 is required for NGDRQC− but not for NGDRQC+, and that it involves the first two N-terminal Cue domains. In contrast, Trp122 of Cue2 is crucial for NGDRQC+. Moreover, Mbf1 is required for quality controls by preventing +1 ribosome frameshifting induced by a rare codon staller. We propose that in Cue2-dependent cleavage upstream of the collided ribosomes (NGDRQC−), polyubiquitination of eS7 is recognized by two N-terminal Cue domains of Cue2. In contrast, for the cleavage within collided ribosomes (NGDRQC+), the UBA domain, Trp122 and the interaction between Mbf1 and uS3 are critical.

Proteostasis regulation through ribosome quality control and no‐go‐decay

“…Stabilization of the RQC‐disome is the signaling hub for quality control (Ikeuchi, Tesina, et al, 2019). Recent studies using mutants of Hel2 showed that NGD might also be independent of RQC or Hel2, and thus, the complete series of molecular events leading to NGD might be mRNA‐ and cell‐state‐dependent (Ikeuchi, Izawa, & Inada, 2019; Tomomatsu et al, 2023). We provide here an update on NGD factors and the sequence of events that ultimately direct the mRNA degradation, disassembly of the ribosomal subunits, and degradation of the truncated polypeptide chain (Figure 4 and Table 1).…”

“…Hel2 ubiquitinates the ribosomal proteins in its proximity; uS10 (Rps20), uS3 (Rps3), and eS7 (Rps7) in yeast (Ikeuchi, Tesina, et al, 2019; Matsuo et al, 2017; Sugiyama et al, 2019; Tomomatsu et al, 2023). In mammalian cells, ZNF598 ubiquitinates ribosomal proteins eS10 (Rps10) at K138/K139 and to a lesser extent uS10 at K4/K8 located in the mRNA entry channel of the 40S subunit (Juszkiewicz & Hegde, 2017; Sundaramoorthy et al, 2017).…”

“…In mammalian cells, ZNF598 ubiquitinates ribosomal proteins eS10 (Rps10) at K138/K139 and to a lesser extent uS10 at K4/K8 located in the mRNA entry channel of the 40S subunit (Juszkiewicz & Hegde, 2017; Sundaramoorthy et al, 2017). The ubiquitination of eS10 was not identified in this context in yeast (Matsuo et al, 2017; Tomomatsu et al, 2023). Hel2 was identified in a yeast genetic screen for factors mediating co‐translational stalling under heat stress conditions along with Asc1 and factors involved in nascent chain release and degradation, Ltn1, Rqc1, Rqc2/Tae2, and Cdc48p (Brandman et al, 2012).…”

“…In the NGD RQC‐ pathway, cleavage occurs upstream of the first disome unit. In both pathways, the endonuclease Cue2 recognizes stalling ribosomes and cleaves the mRNA (Tomomatsu et al, 2023).…”

“…Thus, ribosomes recognize errors in the mRNA during translation elongation, which halt their elongation and activate ribosome quality control (RQC) mechanisms (Brandman et al, 2012; Matsuo et al, 2017; Pisareva et al, 2011; Shoemaker et al, 2010; Shoemaker & Green, 2011). RQC encompasses mRNA surveillance and protein quality control to repress their translation and triggers the degradation of problematic mRNAs and truncated nascent peptides (Bengtson & Joazeiro, 2010; D'Orazio et al, 2019; Ito‐Harashima et al, 2007; Juszkiewicz & Hegde, 2017; Matsuo et al, 2017; Simms et al, 2017; Sitron et al, 2017; Tomomatsu et al, 2023; Veltri et al, 2022). Translation and ribosome biogenesis are energy‐expensive processes, as they require the coordination of rRNA transcription, modification of rRNAs, and translation of ribosomal RNAs to achieve ribosome assembly (Hershey et al, 2019; Holcik & Sonenberg, 2005; Li et al, 2009; Young & Guydosh, 2022).…”