title: Differential protein kinase C-dependent modulation of Kv7.4 and Kv7.5 subunits of vascular Kv7 channels.
abstract: The Kv7 family (Kv7.1-7.5) of voltage-activated potassium channels contributes to the maintenance of resting membrane potential in excitable cells. Previously, we provided pharmacological and electrophysiological evidence that Kv7.4 and Kv7.5 form predominantly heteromeric channels and that Kv7 activity is regulated by protein kinase C (PKC) in response to vasoconstrictors in vascular smooth muscle cells. Direct evidence for Kv7.4/7.5 heteromer formation, however, is lacking. Furthermore, it remains to be determined whether both subunits are regulated by PKC. Utilizing proximity ligation assays to visualize single molecule interactions, we now show that Kv7.4/Kv.7.5 heteromers are endogenously expressed in vascular smooth muscle cells. Introduction of dominant-negative Kv7.4 and Kv7.5 subunits in mesenteric artery myocytes reduced endogenous Kv7 currents by 84 and 76%, respectively. Expression of an inducible protein kinase Cα (PKCα) translocation system revealed that PKCα activation is sufficient to suppress endogenous Kv7 currents in A7r5 rat aortic and mesenteric artery smooth muscle cells. Arginine vasopressin (100 and 500 pm) and the PKC activator phorbol 12-myristate 13-acetate (1 nm) each inhibited human (h) Kv7.5 and hKv7.4/7.5, but not hKv7.4 channels expressed in A7r5 cells. A decrease in hKv7.5 and hKv7.4/7.5 current densities was associated with an increase in PKC-dependent phosphorylation of the channel proteins. These findings provide further evidence for a differential regulation of Kv7.4 and Kv7.5 channel subunits by PKC-dependent phosphorylation and new mechanistic insights into the role of heteromeric subunit assembly for regulation of vascular Kv7 channels.
authors.affiliation: From the Departments of Molecular Pharmacology and Therapeutics.
authors.forename: Lioubov I
authors.initials: LI
authors.lastname: Brueggemann
authors.forename: Alexander R
authors.initials: AR
authors.lastname: Mackie
authors.forename: Leanne L
authors.initials: LL
authors.lastname: Cribbs
authors.forename: Jessica
authors.initials: J
authors.lastname: Freda
authors.forename: Abhishek
authors.initials: A
authors.lastname: Tripathi
authors.forename: Matthias
authors.initials: M
authors.lastname: Majetschak
authors.forename: Kenneth L
authors.initials: KL
authors.lastname: Byron
mesh_terms: D000818:Animals; D001011:Aorta / Q000166:cytology / Q000378:metabolism; D001127:Arginine Vasopressin / Q000494:pharmacology; D002273:Carcinogens / Q000494:pharmacology; D002460:Cell Line; D006801:Humans; D051656:KCNQ Potassium Channels / Q000235:genetics / Q000378:metabolism*; D008297:Male; D008638:Mesenteric Arteries / Q000166:cytology / Q000378:metabolism; D020125:Mutation, Missense; D032389:Myocytes, Smooth Muscle / Q000166:cytology / Q000378:metabolism*; D051571:Protein Kinase C-alpha / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014662:Vasoconstrictor Agents / Q000494:pharmacology
chemical_list: D002273:Carcinogens; D051656:KCNQ Potassium Channels; C117367:KCNQ4 protein, human; C412231:KCNQ5 protein, human; C492166:Kcnq4 protein, rat; C501028:Kcnq5 protein, rat; D014662:Vasoconstrictor Agents; D001127:Arginine Vasopressin; C496628:PRKCA protein, human; D051571:Protein Kinase C-alpha; D013755:Tetradecanoylphorbol Acetate
keywords: KCNQ Channel; Mesenteric Artery; Patch Clamp Electrophysiology; Potassium Channels; Protein Phosphorylation; Signal Transduction; Vascular Smooth Muscle Cells; Vasopressin