A Novel Role for the Centrosomal Protein, Pericentrin, in Regulation of Insulin Secretory Vesicle Docking in Mouse Pancreatic β-cells

The centrosome is important for microtubule organization and cell cycle progression in animal cells. Recently, mutations in the centrosomal protein, pericentrin, have been linked to human microcephalic osteodysplastic primordial dwarfism (MOPD II), a rare genetic disease characterized by severe growth retardation and early onset of type 2 diabetes among other clinical manifestations. While the link between centrosomal and cell cycle defects may account for growth deficiencies, the mechanism linking pericentrin mutations with dysregulated glucose homeostasis and pre-pubertal onset of diabetes is unknown. In this report we observed abundant expression of pericentrin in quiescent pancreatic β-cells of normal animals which led us to hypothesize that pericentrin may have a critical function in β-cells distinct from its known role in regulating cell cycle progression. In addition to the typical centrosome localization, pericentrin was also enriched with secretory vesicles in the cytoplasm. Pericentrin overexpression in β-cells resulted in aggregation of insulin-containing secretory vesicles with cytoplasmic, but not centrosomal, pericentriolar material and an increase in total levels of intracellular insulin. RNAi- mediated silencing of pericentrin in secretory β-cells caused dysregulated secretory vesicle hypersecretion of insulin into the media. Together, these data suggest that pericentrin may regulate the intracellular distribution and secretion of insulin. Mice transplanted with pericentrin-depleted islets exhibited abnormal fasting hypoglycemia and inability to regulate blood glucose normally during a glucose challenge, which is consistent with our in vitro data. This previously unrecognized function for a centrosomal protein to mediate vesicle docking in secretory endocrine cells emphasizes the adaptability of these scaffolding proteins to regulate diverse cellular processes and identifies a novel target for modulating regulated protein secretion in disorders such as diabetes

PERICENTRIN KNOCKDOWN BY RNAi DISRUPTS INSULIN SECRETION IN CULTURED MURINE INSULINOMA CELLS

The centrosome functions in spindle formation, transport of cytoplasmic vesicles, and regulation of cell structure. Pericentrin, a centrosomal protein, was chosen to study for its role in microtubule nucleation and organization, cell cycle progression and ciliogenesis. This project's purpose was to determine if the amount of endogenous pericentrin would affect insulin storage or secretion in pancreatic beta cells. Immunofluorescent staining and electron microscopy revealed that pericentrin co-localizes to the granules of beta cells. After Pericentrin depletion with RNAi, a lower number of mature insulin granules was found, and a high amount of insulin was secreted into the media. This dys-regulation of insulin secretion was also verified by in vivo experiments