PROJECT SUMMARY Diabetes mellitus is the most common cause of chronic kidney disease. Approximately 40% of diabetic patients develop diabetic nephropathy (DN); and nearly half of all incident cases of end-stage renal disease in the United States are due to DN. Despite widespread use of targeted therapies to lower glucose and to antagonize the renin-angiotensin system, the prevalence of DN has remained stable. Thus, effective and disease-specific therapies for DN are needed. The immediate goal of the applicant is to study the mechanisms underlying the development of glomerular basement membrane (GBM) thickening in DN. The applicant's long- term goal is to understand the pathogenesis of diabetic kidney disease in sufficient detail to develop mechanism-based therapies for DN. A clear and focused career development plan has been designed by the applicant and the mentoring team to provide new and enhanced training in cell-extracellular matrix interactions in DN, insulin receptor signaling, communication and presentation skills, grant and manuscript writing, and responsible conduct in research. There is a plan for progressive increase in independence over the course of the CDA-2 Award. Timelines for the specific aims, laboratory space allocation, scheduled meetings between the applicant and the mentoring team, expectations of the applicant, and the specific role(s) of each mentor are addressed in detail in the career development plan. In addition, the applicant is expected to submit smaller grant proposals in the earlier years and an application for independent funding (VA Merit Award, NIH R01) in the last two years of the CDA-2 Award. Studies currently being conducted by the applicant have demonstrated that aminopeptidase-A (APA), a type II transmembrane protein expressed in visceral glomerular epithelial cells (podocytes), may play an important role in the pathogenesis of GBM thickening in DN. Initial results show that APA knockout mice have significantly thickened GBMs. Moreover, APA is down-regulated in patients with DN and in rat models of type 2 DN. Also, knockdown of the insulin receptor in cultured glomerular epithelial cells reduces the expression of APA. In this proposal, the applicant will study the role of APA downregulation in mediating the expression of genes involved in GBM remodeling. The overall hypothesis is that APA down-regulation in the diabetic state causes displacement of transcriptional factors (ZHX proteins) that migrate into the podocyte nucleus and induce changes in the expression of matrix- related genes, thereby leading to GBM thickening. The goal of the proposed studies is to increase our understanding of key molecular changes that result in the development of GBM thickening and DN. In Specific Aim 1, the effects of APA down-regulation on matrix-related gene expression will be studied. In Specific Aim 2, the importance of APA-ZHX interaction in the development of GBM thickening will be examined. In Specific Aim 3, the effects of insulin receptor deficiency on APA and ZHX proteins will be investigated.