Glucose activates the carboxyl methylation of gamma subunits of trimeric GTP-binding proteins in pancreatic beta cells. Modulation in vivo by calcium, GTP, and pertussis toxin.

Abstract

The ␥ subunits of trimeric G-proteins ( ␥ 1 , ␥ 2 , ␥ 5 , and ␥ 7 isoforms) were found to be methylated at their carboxyl termini in normal rat islets, human islets and pure ␤ [HIT-T15] cells. Of these, GTP ␥ S significantly stimulated the carboxyl methylation selectively of ␥ 2 and ␥ 5 isoforms. Exposure of intact HIT cells to either of two receptor-independent agonists-a stimulatory concentration of glucose or a depolarizing concentration of K ϩ -resulted in a rapid (within 30 s) and sustained (at least up to 60 min) stimulation of ␥ subunit carboxyl methylation. Mastoparan, which directly activates G-proteins (and insulin secretion from ␤ cells), also stimulated the carboxyl methylation of ␥ subunits in intact HIT cells. Stimulatory effects of glucose or K ϩ were not demonstrable after removal of extracellular Ca 2 ϩ or depletion of intracellular GTP, implying regulatory roles for calcium fluxes and GTP; however, the methyl transferase itself was not directly activated by either. The stimulatory effects of mastoparan were resistant to removal of extracellular Ca 2 ϩ , implying a mechanism of action that is different from glucose or K ϩ but also suggesting that dissociation of the ␣␤␥ trimer is conducive to ␥ subunit carboxyl methylation. Indeed, pertussis toxin also markedly attenuated the stimulatory effects of glucose, K ϩ or mastoparan without altering the rise in intracellular calcium induced by glucose or K ϩ . Glucose-induced carboxyl methylation of ␥ 2 and ␥

Dominant-Negative α-Subunit of Farnesyl- and Geranyltransferase Inhibits Glucose-Stimulated, but Not KCl-Stimulated, Insulin Secretion in INS 832/13 Cells

“…Some of these include the ␥-subunits of trimeric G-proteins and nuclear lamins. Along these lines, we have provided evidence (43) for the direct activation, by glucose, of the carboxylmethylation of ␥-subunits of trimeric G-proteins in islet ␤-cells. It should be noted that the carboxylmethylation is not expected to take place until the candidate ␥-subunit is prenylated.…”

Bridging the gap between protein carboxyl methylation and phospholipid methylation to understand glucose-stimulated insulin secretion from the pancreatic β cell

“…Using normal rat islets and clonal β cell preparations, we have been able to demonstrate transient stimulation, by glucose, of the CML of small G-proteins [e.g., Cdc42 or Rap1] and the γ subunits of trimeric G-proteins [19]. Further, we have demonstrated that these proteins are rapidly demethylated in isolated β cell preparations [17]. We have also characterized protein prenyl methyl transferase activity in the islet β cell [20].…”

“…It utilizes the intracellular SAM as the methyl donor. In addition, a methylesterase activity that specifically cleaves the methylated residue has also been identified in the islet β cell [17]. Previously, we have identified several carboxylmethylated proteins in the pancreatic β cell.…”

“…In contrast, the CML step is subject to acute regulation at the level of the "on" step [addition of methyl groups] and/or the "off" step [removal of methyl groups]. Earlier studies have demonstrated relevance of post-translational modifications of these proteins in physiological insulin secretion [2,4,5,[16][17][18][19]. Of contextual importance to this current commentary are potential contributory roles for the protein CML steps in GSIS; these aspects are discussed below.…”

“…Follow-up studies from our laboratory have utilized similar experimental approaches and probes to decipher the roles of the CML of Gγ subunits in GSIS [17]. The γ subunits of trimeric G-proteins [γ 1 , γ 2 , γ 5 , and γ 7 isoforms] were found to be carboxylmethylated in normal rat islets, human islets and clonal β cell preparations.…”