Project Summary/Abstract Through this K22 Career Development Award proposal, I seek further mentored research training while developing career skills to facilitate a successful transition to research independence. Therefore, this proposed plan is tailored to extend beyond my skillset developed during my postdoctoral fellowship at the NHLBI beyond metabolic studies and into immunology. This proposal takes advantage of the outstanding scientific environment in my mentor’s lab and at the NHLBI to learn new techniques, including next-generation sequencing analysis, advanced flow cytometry, histopathology, and proteomics. To train in these methods and assist with the research aims, I have established collaborations with other labs and several NHLBI core facilities. Furthermore, a career development plan is included to ensure thorough preparation for an academic position by cultivating my oral and written communication, mentorship, and lab management. Advisory Committee has been assembled consisting of my primary mentor and several other prominent scientists, who not only have extensive scientific experience in fields related to the T cell biology and cellular metabolism, which are proposed in this grant, but have also committed to guiding presentations, job applications, and negotiation strategies. This training will help me to secure a tenure-track position at an extramural institution. The research goal of this proposal is to dissect the roles (Aim 1) and the detailed mechanisms (Aim 2) of mitochondrial fatty acid β-oxidation (FAO) in CD4 T cell differentiation and functions and to further evaluate the translational applications of altering FAO levels in T cells (Aim 3). T cells are central to cell-mediated immunity. The dysregulation of T cell differentiation and related functions leads to a variety of immune-related diseases, such as cancer, infections, and autoimmune and inflammatory diseases. The differentiation of T cell subsets is associated with metabolic rewiring. Metabolic rewiring is a bioenergetic adaptation to different conditions and a hallmark of T cell differentiation is FAO, a major catabolic process that degrades long-chain fatty acids. I have recently generated conditional knockout mice with an endothelial FAO deficiency, and have demonstrated that FAO is a critical regulator of endothelial cell fates both in vitro and in vivo. Another genetic mouse model with a T-cell-specific FAO deficiency has also been produced. Using this novel model, combined with cutting-edge biochemical techniques, I seek to fill the gap in our knowledge of the molecular basis of FAO in the regulation of T cell differentiation and related functions (Aims 1 and 2). Furthermore, the goal of Aim 3 is to evaluate the translational potential of altering FAO levels in T-cell-associated disease models. This proposal is of profound significance because, by answering the fundamental questions of how FAO controls T cell differentiation and functions, it will lay a s...