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Author Notes:

Corresponding Author: Dr. Jorge Arreola, 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, Email: arreola@dec1.ifisica.uaslp.mx, Fax: 52(444) 813-3874.

JJDeJP, SCR, AEES, NECF, ZQ: acquisition, analysis, and interpretation of data, drafting the work

PPC, HCH: conception or design of the work, drafting the work or revising it critically for important intellectual content.

JA: analysis, and interpretation of data, conception or design of the work, drafting the work or revising it critically for important intellectual content.

All authors: Approved the final version of the manuscript and agreed to be accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved.

Electrophysiological experiments were performed at JA lab

Inside out patches experiments were performed at HCH lab

ELISA and western blot experiments were performed at PPC lab

Super resolution microscopy experiments were performed at AMT lab

We wish to acknowledge the technical assistance of Carmen Hernández-Carballo, Yuan-Yuan Cui and Elsa Nydia Hernández Ríos.

We thank Dr. J. Riesgo for support on superresolution microscopy.

None of the authors have competing interests.

Subjects:

Research Funding:

This work was supported by grants 219949, and FDC 2016-01-1955 from CONACYT, México to J. Arreola and P. Pérez-Cornejo, by grants 220224 from CONACYT and IN206616 from PAPIIT-DGAPA, UNAM, Mexico to A. Martinez-Torres and by grants R01-EY0114852 and R01-AR067786 from the National Institutes of Health to H. Criss Hartzell.

J. J. De Jesús-Pérez was supported by Student Fellowship 234820 from CONACYT, México.

Silvia Cruz-Rangel was supported by Postdoctoral Fellowships 256034 and 290807 from CONACYT, México.

Keywords:

  • Science & Technology
  • Life Sciences & Biomedicine
  • Biochemistry & Molecular Biology
  • Biophysics
  • Cell Biology
  • Regulation
  • TMEM16A
  • Anoctamin
  • Lipids
  • Poly-unsaturated fatty acids
  • Patch clamp
  • Fluorescence microscopy
  • CA2+-ACTIVATED CL-CHANNEL
  • ANOCTAMIN 1
  • PLASMA-MEMBRANE
  • ION CHANNELS
  • LIPID RAFTS
  • PIP2
  • VOLTAGE
  • PROTEINS
  • CONTRIBUTES
  • SECRETION

Phosphatidylinositol 4,5-bisphosphate, cholesterol, and fatty acids modulate the calcium-activated chloride channel TMEM16A (ANO1)

Tools:

Journal Title:

Biochimica et Biophysica Acta Molecular and Cell Biology of Lipids

Volume:

Volume 1863, Number 3

Publisher:

, Pages 299-312

Type of Work:

Article | Post-print: After Peer Review

Abstract:

The TMEM16A-mediated Ca2+-activated Cl− current drives several important physiological functions. Membrane lipids regulate ion channels and transporters but their influence on members of the TMEM16 family is poorly understood. Here we have studied the regulation of TMEM16A by phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2), cholesterol, and fatty acids using patch clamp, biochemistry and fluorescence microscopy. We found that depletion of membrane PI(4,5)P2 causes a decline in TMEM16A current that is independent of cytoskeleton, but is partially prevented by removing intracellular Ca2+. On the other hand, supplying PI(4,5)P2 to inside-out patches attenuated channel rundown and/or partially rescued activity after channel rundown. Also, depletion (with methyl-β-cyclodextrin M-βCD) or restoration (with M-βCD + cholesterol) of membrane cholesterol slows down the current decay observed after reduction of PI(4,5)P2. Neither depletion nor restoration of cholesterol change PI(4,5)P2 content. However, M-βCD alone transiently increases TMEM16A activity and dampens rundown whereas M-βCD + cholesterol increases channel rundown. Thus, PI(4,5)P2 is required for TMEM16A function while cholesterol directly and indirectly via a PI(4,5)P2-independent mechanism regulate channel function. Stearic, arachidonic, oleic, docosahexaenoic, and eicosapentaenoic fatty acids as well as methyl stearate inhibit TMEM16A in a dose- and voltage-dependent manner. Phosphatidylserine, a phospholipid whose hydrocarbon tails contain stearic and oleic acids also inhibits TMEM16A. Finally, we show that TMEM16A remains in the plasma membrane after treatment with M-βCD, M-βCD + cholesterol, oleic, or docosahexaenoic acids. Thus, we propose that lipids and fatty acids regulate TMEM16A channels through a membrane-delimited protein-lipid interaction.

Copyright information:

© 2017 Elsevier B.V.

This is an Open Access work distributed under the terms of the Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License (http://creativecommons.org/licenses/by-nc-nd/4.0/).

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