DOC2b Enhances β-Cell Function Via A Novel Tyrosine Phosphorylation-Dependent Mechanism

Double C2 Domain Β (DOC2b) protein is required for glucose-stimulated insulin secretion (GSIS) in β-cells, the underlying mechanism of which remains unresolved. Our biochemical analysis using primary human islets, human and rodent clonal β-cells revealed that DOC2b is tyrosine phosphorylated within 2 minutes of glucose stimulation, and Src family kinase member YES is required for this process. Biochemical and functional analysis using DOC2b^Y301 mutants revealed the requirement of Y301 phosphorylation for the interaction of DOC2b with YES kinase and increased content of VAMP2, a protein on insulin secretory granules, at the plasma membrane (PM), concomitant with DOC2b-mediated enhancement of GSIS in β-cells. Co-immunoprecipitation studies demonstrated an increased association of DOC2b with ERM family proteins in β-cells following glucose stimulation or pervanadate-treatment. Y301 phosphorylation-competent DOC2b was required to increase ERM protein activation, and ERM protein knockdown impaired DOC2b-mediated boosting of GSIS, suggesting that tyrosine-phosphorylated DOC2b regulates GSIS via ERM-mediated granule localization to the PM. Taken together, these results demonstrate the glucose-induced post-translational modification of DOC2b in β-cells, pinpointing the kinase, site of action, and downstream signaling events revealing a regulatory role of YES kinase at various steps in GSIS. This work will enhance the development of novel therapeutic strategies to restore glucose homeostasis in diabetes.