PROJECT SUMMARY / ABSTRACT Acute kidney injury (AKI) is a common clinical disorder, with a total of more than 13 million people affected globally every year. It has a high associated morbidity and mortality, and there are currently no definitive treatments besides supportive care. One of the most common causes of AKI is ischemia reperfusion injury (IRI), characterized by acute tubular necrosis and intrarenal inflammation. Regulated cell death pathways have been increasingly implicated in the tubular injury and resulting inflammation of AKI, and necroptosis in particular plays a critical role during IRI. There are very few known host factors that regulate necroptosis. Preliminary data in this proposal has identified interferon-stimulated gene 15 (ISG15) as a novel regulator of necroptosis and inflammation. Mice lacking ISG15 display a complete susceptibility to AKI in an IRI model, which can be rescued by also eliminating the executioner of necroptosis, mixed-linage kinase domain like pseudokinase (MLKL). In addition, preliminary data suggest that ISG15 negatively regulates necroptosis in murine primary proximal tubule cells in vitro. The overall hypothesis is that ISG15 acts as a critical factor in the host response to renal IRI by regulating the magnitude and timing of necroptosis in proximal tubule cells and downstream inflammation in the kidney. In Aim 1, damage to the kidney epithelium and the magnitude and timing of necroptosis due to an 18-minute bilateral IRI in vivo will be determined, as well as the host inflammatory response in the kidney. In Aim 2, the cell type in which ISG15 is acting to limit injury will be identified, and the interaction of ISG15 with the key signaling complex of necroptosis will be determined. The overall objective of this proposal is to advance understanding of the role of cell death pathways during renal IRI and elucidate the novel role of ISG15 as a host protective factor regulating cell death. This information is critical to advancing our fundamental understanding of the pathophysiology of AKI and developing effective therapeutics. The proposed research and training plan will facilitate the applicant’s development of key knowledge and skills to become an independent physician-scientist. Dr. Deborah Lenschow, the sponsor of this work, has extensive experience studying disease pathogenesis and the innate immune response to tissue injury, and the co-sponsor Dr. Benjamin Humphreys has deep expertise in kidney biology, genomics, and injury. The institutional environment provides a rigorous and supportive intellectual atmosphere as well as collaborative experts in AKI and kidney injury pathogenesis. This fellowship will support the applicant in becoming an independent investigator and a practicing physician-scientist.