GIGYF2 and 4EHP Inhibit Translation Initiation of Defective Messenger RNAs to Assist Ribosome-Associated Quality Control

Abstract

Ribosome-associated Quality Control (RQC) pathways protect cells from toxicity caused by incomplete protein products resulting from translation of damaged or problematic mRNAs.Extensive work in yeast has identified highly conserved mechanisms that lead to the degradation of the faulty mRNA and partially synthesized polypeptide. Here, we used CRISPR-Cas9-based screening to search for additional RQC strategies in mammals. We found that failed translation leads to specific silencing of translation initiation on that message. This negative feedback loop is mediated by two translation inhibitors, GIGYF2 and 4EHP. Their recruitment to defective messages can be mediated by different factors, including potentially the collision sensor ZNF598. Both model substrates and growth-based assays established that inhibition of additional rounds of translation acts in concert with known RQC pathways to prevent buildup of toxic proteins. Inability to block translation of faulty mRNAs, and subsequent accumulation of partially synthesized polypeptides, could explain the neurodevelopmental and neuropsychiatric disorders observed in mice and humans with compromised GIGYF2 function..

Ribosome states signal RNA quality control

“…Here, poly-A sequences have been used to promote internal ribosomal stalls, though as we will discuss, these may be less good mimics of ribosomes trapped upon damaged mRNA (Arthur et al, 2015;Chandrasekaran et al, 2019). One strong stalling reporter that has been used in mammalian cells is an mRNA with a highly structured 3 0 end (the MALAT sequence), though it is not clear that this 3 0 terminal sequence effectively mimics an ORF-internal stalling mRNA (Hickey et al, 2020). More recently, a known natural stalling sequence, XBP1u, has been used to induce ribosomal collisions in reporter mRNA and will likely continue to be a useful tool for studying mammalian ribosomal collisions (Han et al, 2020;Sinha et al, 2020).…”

Combinatorial optimization of mRNA structure, stability, and translation for RNA-based therapeutics

“…The data from the Bioanalyzer were analyzed using a custom script that performs the following analysis. We first converted elution times to nucleotides based on a ladder control (25,200, 500, 1000, 2000, and 4000 nts). We then estimated relative mRNA amounts based on peak areas at expected band lengths (in our case,~900 nucleotides for the mRNAs of interest and~265 nucleotides for the control).…”

Ribosome quality control activity potentiates vaccinia virus protein synthesis during infection

“…Our work indicates that the evolution of a viral response to the host ISR has exposed the virus to a conditional requirement for the RQC pathway in regulating translational reprogramming. While ribosome collisions are part of the regular translation cycle, feedback inhibition of translation initiation through the ISR normally curtails overt RQC activation(Hickey et al 2020; Juszkiewicz et al 2020a; Sinha et al 2020; Vind et al 2020; Wu et al 2020). When viruses interfere with this feedback inhibition, such as vaccinia virus antagonism of PKR-mediated eIF2α phosphorylation, non-productive translation cycles ensue and therefore require RQC based ribosomal rescue.…”

“…Thorough biochemical and genetic studies in multiple organisms have delineated a growing list of molecular constituents within the RQC pathway that act on collided ribosomes(Joazeiro 2019; Inada 2020). Recent studies have uncovered cellular signaling pathways that are induced by collisions that both mount a transcriptional response and result in translation initiation inhibition by multiple routes(Tollenaere et al 2019; Hickey et al 2020; Juszkiewicz et al 2020a; Meydan and Guydosh 2020; Sinha et al 2020; Vind et al 2020; Wu et al 2020). How viruses utilize various RQC components for viral propagation or are impacted by RQC dysfunction remains largely unexplored.…”