Project Summary/Abstract Peroxiredoxin 6 (Prdx6) is a multi-functional enzyme that expresses glutathione peroxidase, phospholipase A2 (PLA2), and lysophosphatidylcholine acyltransferase (LPCAT) activities in separate catalytic sites. Prdx6 can reduce phospholipid hydroperoxides and hydrolyze and re-acylate phospholipid fatty acyl bonds. Prdx6 is, therefore, a complete enzyme for the repair of peroxidized cell membranes. Prdx6 has been implicated in several pathophysiological conditions, including acute lung injury, inflammation, carcinogenesis, various chronic central nervous system diseases, retinal disease, type 2 diabetes, muscle atrophy, and male infertility, but basic questions about the biology of this unique enzyme remain unanswered. Our preliminary data strongly suggest that Prdx6 suppresses ferroptosis, an iron-dependent form of regulated cell death driven by the accumulation of phospholipid hydroperoxides. Emerging evidence implicates ferroptosis in several degenerative diseases, carcinogenesis, stroke, traumatic brain injury, and ischemia/reperfusion injury, among others. Hence, establishing the mechanisms and physiological relevance of ferroptosis regulation by Prdx6 is crucial. My lab is interested in studying the role of the glutathione peroxidase, PLA2, and LPCAT activities of Prdx6 on the regulation of ferroptosis induced by inhibition of cystine uptake, hypoxia/reoxygenation, and oxygen toxicity. We will use mice and cells with single point mutations that inactivate each of the activities of Prdx6 without affecting the others, along with state-of-the-art analytical tools to dissect the role of this enzyme on the regulation of ferroptosis. In a second project, I propose to study the role of Prdx6 in the maintenance of mitochondrial function. Our preliminary data show that Prdx6 deficiency alters transcriptional signatures of mitochondrial metabolism and reduces mitochondrial respiration. We will study the effects of the catalytic activities of Prdx6 on mitochondrial morphology, dynamics, and function using extracellular flux assays and three-dimensional imaging techniques. The results of this proposal will contribute to our understanding of one of the critical mediators of cellular redox balance. These results will also provide essential information for future translational strategies for the prevention and treatment of diseases associated with dysregulated redox homeostasis.