Obesity-linked type 2 diabetes is reaching epidemic proportions in the US, and is caused by an inadequate functional pancreatic ss-cell mass that no longer compensates for the increased metabolic demand. Key to this dysfunction is deficient insulin production that fails to keep pace with the increased secretory demand, and as such, endogenous ss-cell insulin stores deplete and secretory capacity is compromised. Thus, regulation of proinsulin biosynthesis is critically important to ss-cell function. Proinsulin biosynthesis is specifically regulated above that of the majority of ss-cell proteins by many factors, of which glucose is the most physiologically relevant. Unusually, the major control of glucose-induced proinsulin biosynthesis is specifically mediated at the translational level. This translational regulation is guided by conserved cis-elements that reside in the untranslated regions (UTRs) of preproinsulin mRNA (PPI mRNA). The 3'-UTR of PPI mRNA has a highly conserved 'UUGAA cis-element' that assists this translational control mechanism by prolonging PPI mRNA stability in a glucose-regulated fashion. The 5'-UTR of PPI mRNA contains an element (named 'ppIGE', for PPI mRNA glucose element) that is essential for glucose-induced translational control of proinsulin biosynthesis. However, the molecular mechanism behind translational regulation of proinsulin biosynthesis via these key PPI mRNA UTR cis-elements is only partly understood. There are ss-cell cytosolic trans-acting proteins that associate with these 5'-/3'-UTR cis-elements in response to glucose, but these require further functional characterization. We have identified an RNA-binding protein, vigilin, that specifically interacts with the UUGAA-element in the PPI mRNA 3'UTR in a glucose-dependent manner, and more recently identified the glucose-regulated PPI mRNA 5'UTR ppIGE-binding protein (ppIGE- BP) as 'heterogeneous nuclear ribonuclear protein-K' (hnRNP-K). A better characterization of these glucose- influenced RNA-protein interactions will give unique novel insight into the specific translational control mechanism for proinsulin biosynthesis. Moreover, novel reagents generated in the course of these studies will be used to connect back to the secondary signals emanating from glucose metabolism that control proinsulin biosynthesis translation. Somewhat surprisingly, dysfunctional proinsulin biosynthesis has not been directly measured in an obese/type 2 diabetes setting to any extent. Here, we will take advantage of the knowledge and reagents we have developed to gain insight into normal translational control of proinsulin biosynthesis, to also examine the pathological mechanism(s) behind inadequate insulin production in obesity/type 2 diabetes in both rodent models and human islets. It is anticipated that this course of proposed experiments will guide future novel therapeutic strategies to restore adequate insulin production and regain sufficient insulin secretory capacity of ss-cells, which alleviates symptoms of type 2 diabetes. Indeed, in the spirit of translational medicine, the feasibility of such a strategy will also be examined in preclinical models in this proposal. PUBLIC HEALTH RELEVANCE: Obesity-linked type-2 diabetes is caused by decreased in functional pancreatic ss-cell mass that is no longer able to compensate for the peripheral insulin resistance, of which a major part is inadequate insulin production. The overall goal of this application is to gain a much better idea of how insulin biosynthesis is specifically controlled at the molecular level via specific regulatory elements in the insulin mRNA, under normal, obese and type-2 diabetes conditions. This might eventually lead to a novel therapeutic means to adequately restore insulin production and secretory capacity to ss-cells in obesity that alleviates symptoms and delays, perhaps indefinitely, the onset of type 2 diabetes.