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Author Notes:

Corresponding Author: Dr. Jorge Arreola, Physics Institute, Universidad Autónoma de San Luis Potosí, Avenida Dr. Manuel Nava #6, Zona Universitaria, San Luis Potosí, SLP 78290, México, Tel. 52(444)826 2363 ext 136, arreola@dec1.ifisica.uaslp.mx, Fax: 52(444)813-3874

The authors thank C.Y. Hernandez-Carballo and Yuan Yuan Cui for excellent technical assistance and Kuai Yu for providing ANO1 patch clamp records for analysis.

Subjects:

Research Funding:

This work was supported by grants 219949 (CONACyT, Mexico) to JA. HCH is supported by grants from the National Institutes of Health GM60448, AR067786, and EY11482. JAC-V and JJ De J-P are recipients of a Graduate Student Fellowship from CONACyT, Mexico (229968 and 234820).

SCR is a recipient of a Postdoctoral Fellowship from CONACyT, Mexico (256034-290807).

Keywords:

  • Science & Technology
  • Life Sciences & Biomedicine
  • Physiology
  • Chloride channel
  • Kinetics
  • Patch clamp
  • Permeation
  • Gating
  • Mathematical modelling
  • CA2+-ACTIVATED CL-CHANNEL
  • GLAND ACINAR-CELLS
  • VOLTAGE-DEPENDENCE
  • EPITHELIAL-CELLS
  • ANION PERMEATION
  • XENOPUS OOCYTES
  • SINGLE-CHANNEL
  • CALCIUM
  • INACTIVATION
  • GATE

Revealing the activation pathway for TMEM16A chloride channels from macroscopic currents and kinetic models

Tools:

Journal Title:

Pflügers Archiv European Journal of Physiology

Volume:

Volume 468, Number 7

Publisher:

, Pages 1241-1257

Type of Work:

Article | Post-print: After Peer Review

Abstract:

TMEM16A (ANO1), the pore-forming subunit of calcium-activated chloride channels, regulates several physiological and pathophysiological processes such as smooth muscle contraction, cardiac and neuronal excitability, salivary secretion, tumour growth and cancer progression. Gating of TMEM16A is complex because it involves the interplay between increases in intracellular calcium concentration ([Ca 2+ ] i ), membrane depolarization, extracellular Cl − or permeant anions and intracellular protons. Our goal here was to understand how these variables regulate TMEM16A gating and to explain four observations. (a) TMEM16A is activated by voltage in the absence of intracellular Ca 2+ . (b) The Cl − conductance is decreased after reducing extracellular Cl − concentration ([Cl − ] o ). (c) I Cl is regulated by physiological concentrations of [Cl − ] o . (d) In cells dialyzed with 0.2 μM [Ca 2+ ] i , Cl − has a bimodal effect: at [Cl − ] o < 30 mM TMEM16A current activates with a monoexponential time course, but above 30 mM, [Cl − ] o I Cl activation displays fast and slow kinetics. To explain the contribution of V m , Ca 2+ and Cl − to gating, we developed a 12-state Markov chain model. This model explains TMEM16A activation as a sequential, direct, and V m -dependent binding of two Ca 2+ ions coupled to a V m -dependent binding of an external Cl − ion, with V m -dependent transitions between states. Our model predicts that extracellular Cl − does not alter the apparent Ca 2+ affinity of TMEM16A, which we corroborated experimentally. Rather, extracellular Cl − acts by stabilizing the open configuration induced by Ca 2+ and by contributing to the V m dependence of activation.

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© 2016, Springer-Verlag Berlin Heidelberg.

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