PROJECT SUMMARY/ABSTRACT This proposal outlines a five-year plan for the PI, Dr. Alexander Gitlin, to prepare him for an independent academic career as a physician-scientist. Dr. Gitlin received his M.D./Ph.D. degrees from the Weill Cornell/Rockefeller/Sloan-Kettering Tri-Institutional M.D.-Ph.D. program and is currently completing residency in Clinical Pathology at Stanford Hospital and Clinics. During residency, Dr. Gitlin developed a jointly mentored research program, co-advised by Drs. Bali Pulendran (Stanford University) and Vishva Dixit (Genentech, Inc.). Drs. Pulendran and Dixit are longtime colleagues and collaborators with highly complementary expertise. Dr. Pulendran is an expert in toll-like receptors, innate immunity, and dendritic cells; Dr. Dixit is an expert on inflammatory signaling, cell death, and ubiquitin biology. Both Drs. Pulendran and Dixit have served as mentors to numerous trainees, many of whom have become leading investigators in academia or industry. This joint mentorship program provides Dr. Gitlin with full access to the combined resources and expertise present at two neighboring, world-class research institutions, Stanford University and Genentech, which will provide outstanding environments for Dr. Gitlin to develop his own independent scientific career. In addition, Dr. Gitlin has assembled a K08 advisory committee composed of senior investigators whose scientific expertise and mentorship will greatly aid Dr. Gitlin’s career development. They include Drs. Stephen Galli, Denise Monack, Scott Boyd, and Andrey Shaw. Furthermore, Dr. Gitlin will have access to coursework, retreats, seminars and resources through Stanford University’s School of Medicine, The Stanford Office of Postdoctoral Affairs, and the Genentech Postdoctoral Program. Finally, the scientific resources available to Dr. Gitlin comprise the Pulendran/Dixit laboratories and core facilities at both Stanford University and Genentech, representing an extraordinary set of scientific resources. The research proposal detailed herein seeks to uncover the molecular mechanisms by which linear ubiquitin controls the magnitude of the innate immune response. Dysregulated inflammatory and cytokine responses are fundamental features of multiple chronic diseases, including autoimmunity, autoinflammatory syndromes, infectious diseases, immunodeficiencies and cancer. Yet, many of the complex signaling pathways that regulate inflammatory signaling are still being unraveled at a mechanistic level. During Dr. Gitlin’s short time as a co-mentee of Drs. Dixit and Pulendran, he has discovered that the NEDD4-binding protein 1 (N4BP1), a linear ubiquitin-binding protein, is a novel regulator of toll-like receptor (TLR) responses. In Aim 1, we will test the hypothesis that N4BP1 differentially regulates inflammatory cytokine production downstream of TLR signaling. In Aim 2, we will dissect the mechanistic basis of N4BP1 activity by inactivating its independent functional motifs....