The development of glucose-responsive, insulin-secreting cells for beta cell replacement therapy will require a better understanding of the molecular determinants of pancreatic islet function and beta-cell differentiation. Increased knowledge of the mechanisms that mediate insulin secretion is also necessary for the identification of new pharmaceutical targets for the treatment of type 2 diabetes. Guiding this proposal is the long-term goal of determining the role of neuronal synaptic proteins and neurotransmitter mechanisms in insulin secretion, in -cell paracrine signaling, and in the coordination of islet activity. The objective of the present application is to identify the role in beta-cell function of a distinct subset of synaptic cell-surface adhesion molecules: those capable of inducing synapse formation and assembly of the machinery necessary for inhibitory or excitatory neurotransmission. Guided by novel preliminary data, this application focuses on two key families of these adhesion proteins, the neuroligins and their major binding partners, the neurexins. The central hypothesis is that extracellular interactions between neuroligins and neurexins help drive -cell functional maturation, including assembly of the exocytic machinery, and are responsible for the enhancement of insulin secretion induced by -cell- cell contact. To attain the objective of this application, three specific aims are pursued: 1) Identify the neuroligin and neurexin isoforms that are necessary for normal beta-cell secretory function and their role in insulin exocytosis; 2) Identify the mechanisms whereby the neuroligins and neurexins influence beta-cell function; and 3) Test the role of changes in neuroligin and neurexin expression in type 2 diabetes-associated beta-cell dysfunction. Existing neuroligin knockout mice and RNA interference will be used to assess the effect on insulin secretion of reduced expression of neuroligin and neurexin isoforms. The role of neuroligins and neuroligin-neurexin interactions in -cell function will be analyzed using cocultures of INS-1 and dissociated rat-islet cells with neuroligin-expressing HEK293 cells, by immunohistochemical and immunoprecipitation experiments, and by use islet perifusion and monitoring of intracellular calcium. INS-1 cells cultured under glucotoxic conditions and islets from Zucker diabetes fatty rats will be used as models of beta cell dysfunction in type 2 diabetes. The proposed work is innovative because it stems from the novel observation that neuroligins and neurexins are coexpressed on the cell surface and influence insulin secretion and because it will be carried out in collaboration with a laboratory that is a leader in the field of synaptic function. The proposed research is significant because it is expected to advance and expand our understanding of the role of synaptic cell-surface molecules and extracellular protein interactions in insulin secretion and -cell functional maturation. PUBLIC HEALTH RELEVANCE: The proposed work is expected to enable the development of strategies to use beta cell surface proteins as pharmacological targets for the treatment of diabetes-associated beta-cell dysfunction and as cellular components to enhance the physiological resemblance of engineered cells to true cells. It may also assist in the identification of useful cell-surface markers of functional beta cells. It is thus expected to be of direct significance for the development of therapies for type 1 and type 2 diabetes and is responsive to PA-07-056.