A 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 K(V)1.4 via activation of protein kinase A (PKA). Using mutant forms of K(V)1.4 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 K(V)1.4 localization following GIP stimulation demonstrated phosphorylation-dependent rapid endocytosis of K(V)1.4. Expression of K(V)1.4 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) K(V)1.4, or a T601A mutant form resistant to PKA phosphorylation, resulted in reduced glucose-stimulated insulin secretion; WT K(V)1.4 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 K(V)1.4 cell surface expression and modulation of A-type potassium currents, which is likely to be critically important for its insulinotropic action.
Kim
Su-Jin
SJ
Department of Cellular & Physiological Sciences, University of British Columbia, Vancouver, British Columbia V6T 1Z3, Canada.
Choi
Woo Sung
WS
Han
John Song Mou
JS
Warnock
Garth
G
Fedida
David
D
McIntosh
Christopher H S
CH
D000328:Adult; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D002460:Cell Line; D002478:Cells, Cultured; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D004705:Endocytosis / Q000187:drug effects; D005749:Gastric Inhibitory Polypeptide / Q000494:pharmacology*; D006801:Humans; D007328:Insulin / Q000378:metabolism; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000187:drug effects / Q000502:physiology; D007700:Kinetics; D051665:Kv1.4 Potassium Channel; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D010957:Plasmids; D026902:Potassium Channel Blockers / Q000494:pharmacology*; D024642:Potassium Channels, Voltage-Gated / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D011487:Protein Conformation; D051381:Rats; D011964:Receptors, Gastrointestinal Hormone / Q000235:genetics / Q000502:physiology*; D014162:Transfection
D007328:Insulin; C496977:KCNA4 protein, human; C496979:Kcna4 protein, rat; D051665:Kv1.4 Potassium Channel; D026902:Potassium Channel Blockers; D024642:Potassium Channels, Voltage-Gated; D011964:Receptors, Gastrointestinal Hormone; D005749:Gastric Inhibitory Polypeptide; C041086:gastric inhibitory polypeptide receptor; D017868:Cyclic AMP-Dependent Protein Kinases