Abstract 1687 No-Go Decay Maintains the Fidelity of the Translating Pool of mRNA in Response to Physiologically Produced ROS

No-Go Decay (NGD) is a highly conserved quality control mechanism which allows for the degradation of damaged mRNA on which ribosomes have stalled during translation. The Dom34p/Hbs1p complex acts to remove stalled ribosomes and it promotes cleavage of damaged mRNA. The 5' fragment produced from this cleavage is degraded by the cytoplasmic exosome 3' to 5'. Whereas the 3' fragment is localized into P-bodies containing the exonuclease Xrn1p which is responsible for degrading mRNA in the 5' to 3' direction. Despite significant knowledge of NGD mechanistically, little is known as to why this quality control mechanism is conserved. Previous work showed that activation of NGD can be observed by using engineered mRNA substrates that elicit ribosome stalls, or by eliciting the presence of reactive oxygen species (ROS) using oxidants that are not physiological. Reactive oxygen species cause damage to mRNA resulting in the formation of 8-oxo(G) bases, which have been shown to cause ribosome stalls during translation and activation of NGD. Aerobic glucose metabolism is known to generate intracellular ROS which could cause 8-oxo(G) base formation in mRNA and an increase in NGD. By stressing yeast during diauxic shift or under glucose limiting conditions, where yeast perform aerobic respiration, we show that NGD is activated and there is a subsequent increase in P-bodies. Furthermore, general translation is attenuated. To combat damage brought on by ROS, eukaryotic cells express superoxide dismutase proteins. These proteins are highly conserved and act as scavengers that catalyze the breakdown of ROS into H2O2 and O2. Yeast express two conserved Sod proteins, Sod1p which localizes to the cytoplasm, and Sod2p which localizes to the mitochondrial matrix. In strains lacking Sod1p or Sod2p we also see activation of NGD, an increase in P-body assembly, and a decrease in global translation. Taken together, the data shows that increases in physiologic ROS result in upregulation of NGD. Our work suggests that NGD acts to maintain the fidelity of the translating pool in response to physiological ROS, and that could be why this mechanism is conserved. We thank the Winona State University Foundation, and Jim and Susan Anderson for research funding and the gift of a deconvolution microscope.