NIH Research Festival
G protein-coupled receptors (GPCRs) regulate many key functions of pancreatic beta-cells. Activation of beta-cell M3 muscarinic receptors (M3Rs) greatly facilitates insulin release, suggesting that this pathway represents a potential target for novel antidiabetic drugs. M3R activity, like signaling through other GPCRs, is regulated by phosphorylation by various kinases, including GPCR kinases (GRKs), CK1a, and CK2. An initial screen demonstrated that siRNA-mediated knockdown of CK2a expression in cultured beta-cells (MIN6 cells) endogenously expressing M3Rs greatly enhanced M3R-mediated calcium responses. In vitro studies also demonstrated that CK2 phosphorylates the M3R and that this effect is sensitive to blockade by the highly selective and potent CK2 inhibitor, CX4945. We found that muscarinic agonist induced-insulin release was greatly increased in both MIN6 cells and mouse pancreatic islets following treatment with CK2a siRNA or CX4945, respectively. This augmentation of insulin release was not seen with other insulin secretagogues, suggesting that CK2 inhibition selectively promotes M3R signaling in beta-cells, most likely due to reduced receptor phosphorylation. Consistent with this concept, we found that CX4945 treatment did not affect muscarinic agonist-induced signaling mediated by a phosphorylation-deficient mutant M3R. Finally, in vivo studies showed that treatment of mice with CX4945 greatly enhanced insulin release mediated by activation of beta-cell M3Rs and protected mice against diet-induced hyperglycemia and glucose intolerance. This is the first work demonstrating that CK2 phosphorylation of a GPCR is of physiological relevance. Our findings may lead to the development of novel strategies to augment M3R signaling in pancreatic beta-cells for therapeutic purposes.
Scientific Focus Area: Molecular Biology and Biochemistry
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