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Dual-mode phospholipid regulation of human inward rectifying potassium channels

Dual-mode phospholipid regulation of human inward rectifying potassium channels
Dual-mode phospholipid regulation of human inward rectifying potassium channels
The lipid bilayer is a critical determinant of ion channel activity; however, efforts to define the lipid dependence of channel function have generally been limited to cellular expression systems in which the membrane composition cannot be fully controlled. We reconstituted purified human Kir2.1 and Kir2.2 channels into liposomes of defined composition to study their phospholipid dependence of activity using (86)Rb(+) flux and patch-clamp assays. Our results demonstrate that Kir2.1 and Kir2.2 have two distinct lipid requirements for activity: a specific requirement for phosphatidylinositol 4,5-bisphosphate (PIP(2)) and a nonspecific requirement for anionic phospholipids. Whereas we previously showed that PIP(2) increases the channel open probability, in this work we find that activation by POPG increases both the open probability and unitary conductance. Oleoyl CoA potently inhibits Kir2.1 by antagonizing the specific requirement for PIP(2), and EPC appears to antagonize activation by the nonspecific anionic requirement. Phosphatidylinositol phosphates can act on both lipid requirements, yielding variable and even opposite effects on Kir2.1 activity depending on the lipid background. Mutagenesis experiments point to the role of intracellular residues in activation by both PIP(2) and anionic phospholipids. In conclusion, we utilized purified proteins in defined lipid membranes to quantitatively determine the phospholipid requirements for human Kir channel activity.
0006-3495
620-628
Cheng, Wayland W.L.
160e9e4c-b341-4da9-b4f2-3be8d456dd3e
D'Avanzo, Nazzareno
0af1c4a7-ffe0-44d9-90cc-caa53591750f
Doyle, Declan A.
f85f52c8-ce43-4f15-bd06-1df106f73b26
Nichols, Colin G.
042f1de5-f614-4b02-83fb-0b56b7f62e4d
Cheng, Wayland W.L.
160e9e4c-b341-4da9-b4f2-3be8d456dd3e
D'Avanzo, Nazzareno
0af1c4a7-ffe0-44d9-90cc-caa53591750f
Doyle, Declan A.
f85f52c8-ce43-4f15-bd06-1df106f73b26
Nichols, Colin G.
042f1de5-f614-4b02-83fb-0b56b7f62e4d

Cheng, Wayland W.L., D'Avanzo, Nazzareno, Doyle, Declan A. and Nichols, Colin G. (2011) Dual-mode phospholipid regulation of human inward rectifying potassium channels. Biophysical Journal, 100 (3), 620-628. (doi:10.1016/j.bpj.2010.12.3724). (PMID:21281576)

Record type: Article

Abstract

The lipid bilayer is a critical determinant of ion channel activity; however, efforts to define the lipid dependence of channel function have generally been limited to cellular expression systems in which the membrane composition cannot be fully controlled. We reconstituted purified human Kir2.1 and Kir2.2 channels into liposomes of defined composition to study their phospholipid dependence of activity using (86)Rb(+) flux and patch-clamp assays. Our results demonstrate that Kir2.1 and Kir2.2 have two distinct lipid requirements for activity: a specific requirement for phosphatidylinositol 4,5-bisphosphate (PIP(2)) and a nonspecific requirement for anionic phospholipids. Whereas we previously showed that PIP(2) increases the channel open probability, in this work we find that activation by POPG increases both the open probability and unitary conductance. Oleoyl CoA potently inhibits Kir2.1 by antagonizing the specific requirement for PIP(2), and EPC appears to antagonize activation by the nonspecific anionic requirement. Phosphatidylinositol phosphates can act on both lipid requirements, yielding variable and even opposite effects on Kir2.1 activity depending on the lipid background. Mutagenesis experiments point to the role of intracellular residues in activation by both PIP(2) and anionic phospholipids. In conclusion, we utilized purified proteins in defined lipid membranes to quantitatively determine the phospholipid requirements for human Kir channel activity.

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e-pub ahead of print date: 1 February 2011
Published date: 2 February 2011
Organisations: Centre for Biological Sciences

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Local EPrints ID: 342292
URI: http://eprints.soton.ac.uk/id/eprint/342292
ISSN: 0006-3495
PURE UUID: 8a31f02e-aec3-49bc-9e68-3c741407f7fe

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Date deposited: 20 Aug 2012 14:02
Last modified: 14 Sep 2021 19:54

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Contributors

Author: Wayland W.L. Cheng
Author: Nazzareno D'Avanzo
Author: Declan A. Doyle
Author: Colin G. Nichols

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