Calcium/calmodulin-dependent serine protein kinase is involved in exendin-4-induced insulin secretion in INS-1 cells

Abstract

Objective: Exendin-4 (Ex-4) is an anti-diabetic drug that is a potent agonist of the glucagon-like peptide-1 (GLP-1) receptor. It has already been approved for the treatment of type 2 diabetes mellitus, but its underlying mechanisms of action are not fully understood. Calcium/calmodulin-dependent serine protein kinase (CASK), which plays a vital role in the transport and release of neurotransmitters in neurons, is expressed in pancreatic islet cells and β-cells. This study aimed to investigate whether CASK is involved in the insulin secretagogue action induced by Ex-4 in INS-1 cells.

Material/methods: A glucose-stimulated insulin secretion (GSIS) assay was performed with or without siRNA treatment against CASK. The expression level and location of CASK were evaluated by real-time PCR, western blotting and immunofluorescence. With the use of a protein kinase A (PKA) inhibitor or an exchange protein directly activated by cAMP-2 (Epac2) agonist, immunoblotting was performed to establish the signaling pathway through which Ex-4 alters CASK expression.

Results: Knock-down of CASK significantly attenuated the Ex-4-enhanced insulin release, and we showed that Ex-4 could increase transcription of CASK mRNA and expression of CASK protein but did not change the cellular location of CASK. A PKA inhibitor reduced the ability of Ex-4 to stimulate CASK expression, but an Epac2 agonist had no effect suggesting that regulation was mediated by the cAMP/PKA pathway.

Conclusion: Our study suggests that the stimulation of β-cell insulin secretion by Ex-4 is mediated, at least in part, by CASK via a novel signaling mechanism.

Keywords: 8-pCPT-2′-O-Me-cAMP-AM; CASK; ESCA-AM; Epac2; Ex-4; GLP-1; GSIS; H-89; N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinolinesulfonamide; PKA; cAMP; calcium/calmodulin-dependent serine protein kinase; cyclic adenosine monophosphate; exchange protein directly activated by cAMP-2; exendin-4; glucagon-like peptide-1; glucose-stimulated insulin secretion; protein kinase A..