Insulin secretion stimulated by L-arginine and its metabolite L-ornithine depends on Gα(i2).

Bordetella pertussis toxin (PTx), also known as islet-activating protein, induces insulin secretion by ADP-ribosylation of inhibitory G proteins. PTx-induced insulin secretion may result either from inactivation of Gα(o) proteins or from combined inactivation of Gα(o), Gα(i1), Gα(i2), and Gα(i3) isoforms. However, the specific role of Gα(i2) in pancreatic ß-cells still remains unknown. In global (Gα(i2)(-/-)) and ß-cell-specific (Gα(i2)(ßcko)) gene-targeted Gα(i2) mouse models, we studied glucose homeostasis and islet functions. Insulin secretion experiments and intracellular Ca²âº measurements were used to characterize Gα(i2) function in vitro. Gα(i2)(-/-) and Gα(i2)(ßcko) mice showed an unexpected metabolic phenotype, i.e., significantly lower plasma insulin levels upon intraperitoneal glucose challenge in Gα(i2)(-/-) and Gα(i2)(ßcko) mice, whereas plasma glucose concentrations were unchanged in Gα(i2)(-/-) but significantly increased in Gα(i2)(ßcko) mice. These findings indicate a novel albeit unexpected role for Gα(i2) in the expression, turnover, and/or release of insulin from islets. Detection of insulin secretion in isolated islets did not show differences in response to high (16 mM) glucose concentrations between control and ß-cell-specific Gα(i2)-deficient mice. In contrast, the two- to threefold increase in insulin secretion evoked by L-arginine or L-ornithine (in the presence of 16 mM glucose) was significantly reduced in islets lacking Gα(i2). In accord with a reduced level of insulin secretion, intracellular calcium concentrations induced by the agonistic amino acid L-arginine did not reach control levels in ß-cells. The presented analysis of gene-targeted mice provides novel insights in the role of ß-cell Gα(i2) showing that amino acid-induced insulin-release depends on Gα(i2).

ER-resident Gi2 protein controls sar1 translocation onto the ER during budding of transport vesicles.

In our previous study, fluoride ([AlF(4) ](-) ) disturbed ER-to-Golgi transport through the activation of ER-resident heterotrimeric G protein (ER-G protein). Therefore, ER-G protein may be implicated in ER-to-Golgi transport at the early stage prior to coat protein assembly. Sar1 translocation onto the endoplasmic reticulum (ER) membrane is suppressed by non-selective protein kinase inhibitor H89, suggesting the participation of H89-sensitive kinase in this process. To investigate the involvement of ER-G protein in ER-to-Golgi transport, the effect of G(i) protein activator (mastoparan 7) was examined on Sar1 translocation onto the ER in a cell-free system consisting of microsome membrane and cytosol. Sar1 translocation onto the microsome membrane was induced by addition of GTPγS in the cell-free system. Translocation of Sar1 by GTPγS was suppressed significantly by both H89 and mastoparan 7. Mastoparan 7 suppressed the translocation of Sar1 onto the microsome membrane with dosage dependency, but mastoparan 17, the inactive analog of mastoparan 7, had no effect on Sar1 translocation. The suppressive effect of mastoparan 7 was recovered by treatment with pertussis toxin (IAP). Moreover, G(i2) protein was detected on the microsome membrane by western blotting for heterotrimeric G(i) proteins. These results indicate that ER-G(i2) protein modulated Sar1 translocation onto the ER, suggesting that ER-resident G(i2) protein is an important negative regulator of vesicular transport at the early stage of vesicle formation before coat protein assembly on the ER.