SAD-A Promotes Glucose-Stimulated Insulin Secretion Through Phosphorylation and Inhibition of GDIα in Male Islet β Cells

Rho GDP-dissociation inhibitor (GDI alpha) inhibits glucose-stimulated insulin secretion (GSIS) in part by locking Rho GTPases in an inactive GDP-bound form. The onset of GSIS causes phosphorylation of GDI alpha at Ser174, a critical inhibitory site for GDI alpha, leading to the release of Rho GTPases and their subsequent activation. However, the kinase regulator(s) that catalyzes the phosphorylation of GDI alpha in islet beta cells remains elusive. We propose that SAD-A, a member of AMP-activated protein kinase-related kinases that promotes GSIS as an effector kinase for incretin signaling, interacts with and inhibits GDI alpha through phosphorylation of Ser174 during the onset GSIS from islet beta cells. Coimmunoprecipitation and phosphorylation analyses were carried out to identify the physical interaction and phosphorylation site of GDI alpha by SAD-A in the context of GSIS from INS-1 beta cells and primary islets. We identified GDI alpha directly binds to SAD-A kinase domain and phosphorylated by SAD-A on Ser174, leading to dissociation of Rho GTPases from GDI alpha complexes. Accordingly, overexpression of SAD-A significantly stimulated GDI alpha phosphorylation at Ser174 in response to GSIS, which is dramatically potentiated by glucagonlike peptide-1, an incretin hormone. Conversely, SAD-A deficiency, which is mediated by short hairpin RNA transfection in INS-1 cells, significantly attenuated endogenous GDI alpha phosphorylation at Ser174. Consequently, coexpression of SAD-A completely prevented the inhibitory effect of GDI alpha on insulin secretion in islets. In summary, glucose and incretin stimulate insulin secretion through the phosphorylation of GDI alpha at Ser174 by SAD-A, which leads to the activation of Rho GTPases, culminating in insulin exocytosis.