Novel mechanism for the suppression of a voltage-gated potassium channel by glucose-dependent insulinotropic polypeptide - Protein kinase A-dependent endocytosis

The mechanisms involved in glucose regulation of insulin secretion by ATP-sensitive ( K-ATP) and calcium-activated ( K-CA) potassium channels have been extensively studied, but less is known about the role of voltage-gated ( K-V) potassium channels in pancreatic beta-cells. The incretin hormone, glucose-dependent insulinotropic polypeptide ( GIP) stimulates insulin secretion by potentiating events underlying membrane depolarization and exerting direct effects on exocytosis. In the present study, we identified a novel role for GIP in regulating K(V)1.4 channel endocytosis. In GIP receptor-expressing HEK293 cells, GIP reduced A-type peak ionic current amplitude of KV1.4 via activation of protein kinase A (PKA). Using mutant forms of KV1.4 with Ala-Ser/Thr substitutions in a potential PKA phosphorylation site, C-terminal phosphorylation was shown to be linked to GIP-mediated current amplitude decreases. Proteinase K digestion and immunocytochemical studies on mutant KV1.4 localization following GIP stimulation demonstrated phosphorylation-dependent rapid endocytosis of KV1.4. Expression of KV1.4 protein was also demonstrated in human beta-cells; GIP treatment resulting in similar decreases in A-type potassium current peak amplitude to those in HEK293 cells. Transient overexpression in INS-1 beta-cells ( clone 832/13) of wild-type ( WT) KV1.4, or a T601A mutant form resistant to PKA phosphorylation, resulted in reduced glucose- stimulated insulin secretion; WT KV1.4 overexpression potentiated GIP-induced insulin secretion, whereas this response was absent in T601A cells. These results strongly support an important novel role for GIP in regulating KV1.4 cell surface expression and modulation of A-type potassium currents, which is likely to be critically important for its insulinotropic action.