Abstract This F31 proposal describes a comprehensive training and mentorship program for Morgan Gingerich, a Ph.D. candidate in the Cellular and Molecular Biology program at the University of Michigan. Ms. Gingerich will participate in a rigorous didactic and laboratory training curriculum, supervised by her co-mentors, Drs. Soleimanpour and Schnell, as well as support from a multi-disciplinary mentorship/thesis committee. Her training program will include supervised molecular, structural, and biophysical training from a team of scientific experts, as well as mentored opportunities to engage in scientific writing, presentations, and grant applications. The ultimate goal of this proposal is to best position Ms. Gingerich for an independent and productive scientific career. Diabetes is a global epidemic of increasing prevalence. All forms of diabetes stem from insufficient beta- cell function and/or mass to meet peripheral insulin demands. Mitochondrial bioenergetics are critical for beta- cell function, and are disrupted in all forms of diabetes. The long-term objective of my project is to clarify genetic, molecular, and structural mechanisms contributing to beta cell mitochondrial dysfunction in diabetes. Our lab has identified the T1D gene CLEC16A to encode an E3 ligase that regulates selective elimination of damaged/dysfunctional mitochondria via mitophagy. Despite our identification of Clec16a as an E3 ligase, there is no available information identifying conserved functional or structural domains. The overall goal of this project is to define structure-function relationships in Clec16a, through the study of a novel human Clec16a disease- associated isoform which lacks internal/C-terminal regions. Our preliminary data indicate that loss of the Clec16a C-terminus leads to glucose intolerance and beta cell mitochondrial function in vivo, and that both the internal and C-terminal domains affect Clec16a stability and assembly within a key mitophagy regulatory complex. I hypothesize that the Clec16a disease-variant lacks structural and functional regions critical for the maintenance of beta-cell mitophagy and insulin secretion. I will test this hypothesis in the following Specific Aims: Aim 1 will elucidate the role of the Clec16a disease variant in regulating beta-cell function and mitophagy using primary islets from a novel Clec16a C-terminal knockout mouse. I will also assess beta-cell function and mitophagy by re-expressing internal/C-terminal mutants and the disease variant in a Clec16a null beta-cell line. Aim 2 will determine the structural properties and molecular functions of the Clec16a internal and C- terminal regions, clarifying their biological role in disease susceptibility. We will use NMR and informatics to assess Clec16a structure. We will determine the mechanism by which regions lost in the Clec16a disease- associated variant alter Clec16a levels/stability and mitophagy complex assembly using mutagenesis and biochemical assays...