Parallel ZNF598 and GIGYF2 pathways mediate conserved collision-dependent mRNA decay with distinct endonucleolytic features

SUMMARY Quality control of aberrant mRNAs is essential for maintaining cellular homeostasis. Ribosome collisions caused by translational stalling trigger mRNA surveillance pathways that promote degradation of faulty transcripts. In yeast, two major collision-responsive pathways have been proposed: the Hel2–Cue2 pathway and the Syh1–Xrn1 pathway, yet their functional relationship and conservation in mammals have remained unclear. Here, we demonstrate that these two pathways act redundantly to degrade collision-inducing mRNAs in both yeast and mammalian cells. In yeast, we dissect the contribution of Cue2-mediated endonucleolytic cleavage and show that eS7 polyubiquitination–dependent cleavage plays a dominant role in Hel2–Cue2 pathway. In mammals, although the proposed Cue2 homolog N4BP2 is dispensable, we provide the first direct evidence of collision-induced endonucleolytic mRNA cleavage. Together, our findings reveal both conserved and divergent features of ribosome collision–coupled mRNA quality control and expand our understanding of mRNA surveillance and potential therapeutic mRNA implications.