PROJECT SUMMARY/ABSTRACT Increased multi-factorial stress resistance is a property widely shared by models of extended longevity across evolutionary boundaries. Growth hormone (GH) and insulin-like growth factor-1 (IGF-1) receptor deficiencies, for example, which extend lifespan in experimental rodents, also increase resistance to acute oxidative stressors such as paraquat. Dietary restriction, in addition to extending longevity in a wide range of experimental organisms, confers protection against numerous clinically relevant acute stressors, including ischemia reperfusion injury to brain, kidney and liver as well as protection against the toxic side- effects of chemotherapy. Using diet-induced protection from ischemic injury as a model system, we recently identified a novel role for endogenous hydrogen sulfide (H2S) produced by the transsulfuration pathway (TSP) in stress resistance and longevity regulation by dietary restriction. H2S is a gas produced by TSP enzymes CBS and CGL, whose primary role is to convert the essential amino acid methionine to cysteine. Exogenously added H2S can confer numerous benefits ranging from resistance to ischemic injury and suspended animation in experimental mammals, to extended longevity in flies and worms. However, endogenous H2S had not been previously linked to the benefits of dietary restriction. Here, we propose to test the hypothesis that increased endogenous H2S production by TSP enzymes underlies stress resistance and longevity benefits shared by long-lived models. In support of this hypothesis, TSP activity and H2S production are increased in a number of dietary restriction regimens across evolutionary boundaries including in yeast, worms and flies, and in multiple organs in mice upon fasting or dietary protein restriction. Our preliminary data indicate that H2S production by TSP enzymes is repressed by GH and mTOR signaling, two other pathways highly involved in regulation of longevity and stress resistance. Finally, pharmacological or genetic inhibition of CGL and H2S production prevented the benefits of short-term protein restriction against hepatic ischemic injury and protection of bone marrow stem cells from ionizing radiation. Together, these data warrant an investigation into the triggers of endogenous H2S production, the mechanisms by which it promotes oxidative stress resistance and stem cell regeneration, and its interaction with other longevity regulators such as the mitochondrial peptide humanin.