The ASC-1 Complex Disassembles Collided Ribosomes

Abstract

Summary Translating ribosomes that slow excessively incur collisions with trailing ribosomes. Persistent collisions are detected by ZNF598, a ubiquitin ligase that ubiquitinates sites on the ribosomal 40S subunit to initiate pathways of mRNA and protein quality control. The collided ribosome complex must be disassembled to initiate downstream quality control, but the mechanistic basis of disassembly is unclear. Here, we reconstitute the disassembly of a collided polysome in a mammalian cell-free system. The widely conserved ASC-1 complex (ASCC) containing the ASCC3 helicase disassembles the leading ribosome in an ATP-dependent reaction. Disassembly, but not ribosome association, requires 40S ubiquitination by ZNF598, but not GTP-dependent factors, including the Pelo-Hbs1L ribosome rescue complex. Trailing ribosomes can elongate once the roadblock has been removed and only become targets if they subsequently stall and incur collisions. These findings define the specific role of ASCC during ribosome-associated quality control and identify the molecular target of its activity.

The interaction of DNA repair factors ASCC2 and ASCC3 is affected by somatic cancer mutations

“…The helicase regions of ASCC3 and Brr2 seem to be organized in an analogous fashion, comprising an active Nterminal and an inactive, or at least largely dispensable, Cterminal helicase cassette. Consistent with this notion, an ASCC3 variant bearing an ATPase-disrupting substitution in the Cterminal helicase cassette, but not in the N-terminal cassette, can rescue a ribosome poly-A readthrough phenotype elicited by ASCC3 knockdown 25 . Likewise, in the yeast ASCC3 homolog, Slh1p, residue substitutions in the N-terminal cassette can abrogate ribosome quality control function in vivo 27 .…”

“…Finally, a genome-wide CRISPRi screen showed that ASCC2 and ASCC3 are the two most potent modifiers of cell fitness in the presence of a translation inhibitor, suggesting that the proteins affect stalled ribosomes 22 . Indeed, ASCC3 has been suggested to resolve stalled ribosomes on poly-A sequences in a ribosome quality control pathway 23 – 25 . Likewise, its yeast homolog, Slh1p, has been implicated in ribosome-associated quality control to prevent the accumulation of aberrant proteins 23 , 26 , 27 and in non-functional ribosomal RNA decay 28 .…”

EDF1 coordinates cellular responses to ribosome collisions

“…CUE domains are also present in Cue3 (ASCC2 in mammals) which is a part of the trimeric RQT complex (along with yKR023W and Slh1; known as TRIP4 and ASCC3 respectively in mammals) that promotes dissociation of collided ribosomes (Matsuo et al, 2020(Matsuo et al, , 2017. While Ltn1-mediated nascent peptide degradation is compromised in yeast carrying ubiquitin binding mutants of Cue3 (Matsuo et al, 2017), whether the recruitment of the RQT complex is dependent on Hel2-mediated ubiquitination events is currently unclear (Juszkiewicz et al, 2020). Finally, in mammals, ZNF598 was previously shown to associate with the cap-dependent translational initiation repressor EIF4E2 (also called 4EHP) through its binding to the Grb10-interacting GYF protein 2 (GIGYF2) (Garzia et al, 2017;Tollenaere et al, 2019).…”

“…Currently, we do not have evidence for ZNF598-mediated recruitment of GIGYF2•EIF4E2 to these same stall-inducing mRNAs. However, it remains possible that there are semi-redundant mechanisms for recruiting GIGYF2 and EIF4E2 in other situations, or under different environmental perturbations(Tollenaere et al, 2019).Recent studies have established that terminally stalled ribosomes on problematic mRNAs are disassembled by the RQT complex(D'Orazio et al, 2019;Juszkiewicz et al, 2020;Matsuo et al, 2020). If stalled ribosomes are cleared by the ribosomal rescue machinery, why and when are translational repression complexes recruited to problematic mRNAs?…”

Ribosomal stress-surveillance: three pathways is a magic number

“…Irreversible ribosome stalling induces active splitting and rescue of ribosome subunits, and degradation of the incomplete protein product (Figure 3 ). Under these conditions, the ASC-1 helicase complex (Slh1/Rqt2 in yeast) or the recycling factors Pelota, HBS1L (or GTPBP2) and ABCE1 (Dom34, Hbs1 and Rli1 in yeast) split the stalled ribosome into its constituent 60S and 40S subunits ( 101 , 102 ). Subsequently, the released mRNA is degraded by the 5′-3′ exoribonuclease Xrn1 and the exosome complex to prevent the aberrant mRNA from recruiting a new ribosome.…”