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Voltage- and calcium-dependent gating of TMEM16A/Ano1 chloride channels are physically coupled by the first intracellular loop.

ABSTRACT: Ca(2+)-activated Cl(-) channels (CaCCs) are exceptionally well adapted to subserve diverse physiological roles, from epithelial fluid transport to sensory transduction, because their gating is cooperatively controlled by the interplay between ionotropic and metabotropic signals. A molecular understanding of the dual regulation of CaCCs by voltage and Ca(2+) has recently become possible with the discovery that Ano1 (TMEM16a) is an essential subunit of CaCCs. Ano1 can be gated by Ca(2+) or by voltage in the absence of Ca(2+), but Ca(2+)- and voltage-dependent gating are very closely coupled. Here we identify a region in the first intracellular loop that is crucial for both Ca(2+) and voltage sensing. Deleting (448)EAVK in the first intracellular loop dramatically decreases apparent Ca(2+) affinity. In contrast, mutating the adjacent amino acids (444)EEEE abolishes intrinsic voltage dependence without altering the apparent Ca(2+)affinity. Voltage-dependent gating of Ano1 measured in the presence of intracellular Ca(2+) was facilitated by anions with high permeability or by an increase in [Cl(-)](e). Our data show that the transition between closed and open states is governed by Ca(2+) in a voltage-dependent manner and suggest that anions allosterically modulate Ca(2+)-binding affinity. This mechanism provides a unified explanation of CaCC channel gating by voltage and ligand that has long been enigmatic.

SUBMITTER: Xiao Q

PROVIDER: S-EPMC3102354 | biostudies-literature | 2011 May

REPOSITORIES: biostudies-literature

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Voltage- and calcium-dependent gating of TMEM16A/Ano1 chloride channels are physically coupled by the first intracellular loop.

Xiao Qinghuan Q Yu Kuai K Perez-Cornejo Patricia P Cui Yuanyuan Y Arreola Jorge J Hartzell H Criss HC

Proceedings of the National Academy of Sciences of the United States of America 20110509 21

Ca(2+)-activated Cl(-) channels (CaCCs) are exceptionally well adapted to subserve diverse physiological roles, from epithelial fluid transport to sensory transduction, because their gating is cooperatively controlled by the interplay between ionotropic and metabotropic signals. A molecular understanding of the dual regulation of CaCCs by voltage and Ca(2+) has recently become possible with the discovery that Ano1 (TMEM16a) is an essential subunit of CaCCs. Ano1 can be gated by Ca(2+) or by volt ...[more]

PMID: 21555582

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Project description:Pressure-induced arterial depolarization and constriction (the myogenic response) is a smooth muscle cell (myocyte)-specific mechanism that controls regional organ blood flow and systemic blood pressure. Several different nonselective cation channels contribute to pressure-induced depolarization, but signaling mechanisms involved are unclear. Similarly uncertain is the contribution of anion channels to the myogenic response and physiological functions and mechanisms of regulation of recently discovered transmembrane 16A (TMEM16A), also termed Anoctamin 1, chloride (Cl(-)) channels in arterial myocytes.To investigate the hypothesis that myocyte TMEM16A channels control membrane potential and contractility and contribute to the myogenic response in cerebral arteries.Cell swelling induced by hyposmotic bath solution stimulated Cl(-) currents in arterial myocytes that were blocked by TMEM16A channel inhibitory antibodies, RNAi-mediated selective TMEM16A channel knockdown, removal of extracellular calcium (Ca(2+)), replacement of intracellular EGTA with BAPTA, a fast Ca(2+) chelator, and Gd(3+) and SKF-96365, nonselective cation channel blockers. In contrast, nimodipine, a voltage-dependent Ca(2+) channel inhibitor, or thapsigargin, which depletes intracellular Ca(2+) stores, did not alter swelling-activated TMEM16A currents. Pressure-induced (-40 mm Hg) membrane stretch activated ion channels in arterial myocyte cell-attached patches that were inhibited by TMEM16A antibodies and were of similar amplitude to recombinant TMEM16A channels. TMEM16A knockdown reduced intravascular pressure-induced depolarization and vasoconstriction but did not alter depolarization-induced (60 mmol/L K(+)) vasoconstriction.Membrane stretch activates arterial myocyte TMEM16A channels, leading to membrane depolarization and vasoconstriction. Data also provide a mechanism by which a local Ca(2+) signal generated by nonselective cation channels stimulates TMEM16A channels to induce myogenic constriction.

Dataset Information

Voltage- and calcium-dependent gating of TMEM16A/Ano1 chloride channels are physically coupled by the first intracellular loop.

Publications

Voltage- and calcium-dependent gating of TMEM16A/Ano1 chloride channels are physically coupled by the first intracellular loop.

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