IKs response to protein kinase A-dependent KCNQ1 phosphorylation requires direct interaction with microtubules

• Published in: Cardiovascular research Vol. 79; no. 3; pp. 427 - 435
• Main Authors: Nicolas, Céline S., Park, Kyu-Ho, El Harchi, Aziza, Camonis, Jacques, Kass, Robert S., Escande, Denis, Mérot, Jean, Loussouarn, Gildas, Le Bouffant, Françoise, Baró, Isabelle
• Format: Journal Article
• Language: English
• Published: Oxford Oxford University Press 01-08-2008
• Subjects: A Kinase Anchor Proteins - metabolism; Action Potentials; Animals; Biological and medical sciences; Cardiology. Vascular system; Cercopithecus aethiops; COS Cells; Cyclic AMP-Dependent Protein Kinases - metabolism; Guinea Pigs; K+ channel; KCNQ1; KCNQ1 Potassium Channel - genetics; KCNQ1 Potassium Channel - metabolism; Kinetics; Male; Medical sciences; Mice; Microtubules - drug effects; Microtubules - metabolism; Myocytes; Myocytes, Cardiac - drug effects; Myocytes, Cardiac - enzymology; Original; Osmotic Pressure; Phosphorylation; PKA; Protein Binding; Protein Structure, Tertiary; Signal transduction; Transfection; Tubulin - genetics; Tubulin - metabolism; Tubulin Modulators - pharmacology; β-tubulin; Phosphorylation; PKA; Ionic channel; Myocytes; cAMP-dependent protein kinase; Phlebology; β-tubulin; Myocyte; Response; K+ channel; Signal transduction; KCNQ1; Circulatory system; Microtubule; Cardiology
• ISSN: 0008-6363; 1755-3245
• DOI: 10.1093/cvr/cvn085

Aims KCNQ1 (alias KvLQT1 or Kv7.1) and KCNE1 (alias IsK or minK) co-assemble to form the voltage-activated K+ channel responsible for IKs—a major repolarizing current in the human heart—and their dysfunction promotes cardiac arrhythmias. The channel is a component of larger macromolecular complexes containing known and undefined regulatory proteins. Thus, identification of proteins that modulate its biosynthesis, localization, activity, and/or degradation is of great interest from both a physiological and pathological point of view. Methods and results Using a yeast two-hybrid screening, we detected a direct interaction between β-tubulin and the KCNQ1 N-terminus. The interaction was confirmed by co-immunoprecipitation of β-tubulin and KCNQ1 in transfected COS-7 cells and in guinea pig cardiomyocytes. Using immunocytochemistry, we also found that they co-localized in cardiomyocytes. We tested the effects of microtubule-disrupting and -stabilizing agents (colchicine and taxol, respectively) on the KCNQ1–KCNE1 channel activity in COS-7 cells by means of the permeabilized-patch configuration of the patch-clamp technique. None of these agents altered IKs. In addition, colchicine did not modify the current response to osmotic challenge. On the other hand, the IKs response to protein kinase A (PKA)-mediated stimulation depended on microtubule polymerization in COS-7 cells and in cardiomyocytes. Strikingly, KCNQ1 channel and Yotiao phosphorylation by PKA—detected by phospho-specific antibodies—was maintained, as was the association of the two partners. Conclusion We propose that the KCNQ1–KCNE1 channel directly interacts with microtubules and that this interaction plays a major role in coupling PKA-dependent phosphorylation of KCNQ1 with IKs activation.