title: Competition of calcified calmodulin N lobe and PIP2 to an LQT mutation site in Kv7.1 channel.
abstract: Voltage-gated potassium 7.1 (Kv7.1) channel and KCNE1 protein coassembly forms the slow potassium current IKSthat repolarizes the cardiac action potential. The physiological importance of the IKSchannel is underscored by the existence of mutations in human Kv7.1 and KCNE1 genes, which cause cardiac arrhythmias, such as the long-QT syndrome (LQT) and atrial fibrillation. The proximal Kv7.1 C terminus (CT) binds calmodulin (CaM) and phosphatidylinositol-4,5-bisphosphate (PIP2), but the role of CaM in channel function is still unclear, and its possible interaction with PIP2is unknown. Our recent crystallographic study showed that CaM embraces helices A and B with the apo C lobe and calcified N lobe, respectively. Here, we reveal the competition of PIP2and the calcified CaM N lobe to a previously unidentified site in Kv7.1 helix B, also known to harbor an LQT mutation. Protein pulldown, molecular docking, molecular dynamics simulations, and patch-clamp recordings indicate that residues K526 and K527 in Kv7.1 helix B form a critical site where CaM competes with PIP2to stabilize the channel open state. Data indicate that both PIP2and Ca2+-CaM perform the same function on IKSchannel gating by producing a left shift in the voltage dependence of activation. The LQT mutant K526E revealed a severely impaired channel function with a right shift in the voltage dependence of activation, a reduced current density, and insensitivity to gating modulation by Ca2+-CaM. The results suggest that, after receptor-mediated PIP2depletion and increased cytosolic Ca2+, calcified CaM N lobe interacts with helix B in place of PIP2to limit excessive IKScurrent inhibition.
authors.affiliation: Department of Physiology and Pharmacology, Sackler Faculty of Medicine and Sagol School of Neurosciences, Tel Aviv University, Tel Aviv 69978, Israel.
authors.forename: William Sam
authors.initials: WS
authors.lastname: Tobelaim
authors.affiliation: Department of Physiology and Pharmacology, Sackler Faculty of Medicine and Sagol School of Neurosciences, Tel Aviv University, Tel Aviv 69978, Israel.
authors.forename: Meidan
authors.initials: M
authors.lastname: Dvir
authors.affiliation: Department of Biochemistry and Molecular Biology, George S. Wise Faculty of Life Sciences, Institute of Structural Biology, Tel Aviv University, Tel Aviv 69978, Israel.
authors.forename: Guy
authors.initials: G
authors.lastname: Lebel
authors.affiliation: Department of Physiology and Biophysics, Virginia Commonwealth University School of Medicine, Richmond, VA 23298.
authors.forename: Meng
authors.initials: M
authors.lastname: Cui
authors.affiliation: Department of Biochemistry and Molecular Biology, George S. Wise Faculty of Life Sciences, Institute of Structural Biology, Tel Aviv University, Tel Aviv 69978, Israel.
authors.forename: Tal
authors.initials: T
authors.lastname: Buki
authors.affiliation: Department of Physiology and Pharmacology, Sackler Faculty of Medicine and Sagol School of Neurosciences, Tel Aviv University, Tel Aviv 69978, Israel.
authors.forename: Asher
authors.initials: A
authors.lastname: Peretz
authors.affiliation: Laboratory of Structural Physiology, Department of Physiology and Pharmacology, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv 69978, Israel.
authors.forename: Milit
authors.initials: M
authors.lastname: Marom
authors.affiliation: Laboratory of Structural Physiology, Department of Physiology and Pharmacology, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv 69978, Israel.
authors.forename: Yoni
authors.initials: Y
authors.lastname: Haitin
authors.affiliation: Department of Physiology and Biophysics, Virginia Commonwealth University School of Medicine, Richmond, VA 23298.
authors.forename: Diomedes E
authors.initials: DE
authors.lastname: Logothetis
authors.affiliation: Department of Biochemistry and Molecular Biology, George S. Wise Faculty of Life Sciences, Institute of Structural Biology, Tel Aviv University, Tel Aviv 69978, Israel.
authors.forename: Joel Alan
authors.initials: JA
authors.lastname: Hirsch
authors.affiliation: Department of Physiology and Pharmacology, Sackler Faculty of Medicine and Sagol School of Neurosciences, Tel Aviv University, Tel Aviv 69978, Israel; battali@post.tau.ac.il.
authors.forename: Bernard
authors.initials: B
authors.lastname: Attali
mesh_terms: D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D016466:CHO Cells; D020013:Calcium Signaling; D002147:Calmodulin / Q000737:chemistry / Q000378:metabolism*; D006224:Cricetinae; D003412:Cricetulus; D006801:Humans; D055767:Immobilized Proteins; D008133:Long QT Syndrome / Q000235:genetics*; D008958:Models, Molecular; D062105:Molecular Docking Simulation; D056004:Molecular Dynamics Simulation; D009154:Mutation; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000378:metabolism*; D017354:Point Mutation; D011188:Potassium / Q000378:metabolism; D024642:Potassium Channels, Voltage-Gated / Q000378:metabolism; D011487:Protein Conformation; D000072417:Protein Domains; D011994:Recombinant Proteins / Q000378:metabolism; D051637:Shaker Superfamily of Potassium Channels / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013050:Spectrometry, Fluorescence
chemical_list: D002147:Calmodulin; D055767:Immobilized Proteins; C431140:KCNA7 protein, human; C489622:KCNE1 protein, human; D019269:Phosphatidylinositol 4,5-Diphosphate; D024642:Potassium Channels, Voltage-Gated; D011994:Recombinant Proteins; D051637:Shaker Superfamily of Potassium Channels; D011188:Potassium
keywords: KCNQ; LQT; PIP2; calmodulin; potassium channel