Direct regulation of prokaryotic Kir channel by cholesterol.
Our earlier studies have shown that channel activity of Kir2 subfamily of inward rectifiers is strongly suppressed by the elevation of cellular cholesterol. The goal of this study is to determine whether cholesterol suppresses Kir channels directly. To achieve this goal, purified prokaryotic Kir (KirBac1.1) channels were incorporated into liposomes of defined lipid composition, and channel activity was assayed by (86)Rb(+) uptake. Our results show that (86)Rb(+) flux through KirBac1.1 is strongly inhibited by cholesterol. Incorporation of 5% (mass cholesterol/phospholipid) cholesterol into the liposome suppresses (86)Rb(+) flux by >50%, and activity is completely inhibited at 12-15%. However, epicholesterol, a stereoisomer of cholesterol with similar physical properties, has significantly less effect on KirBac-mediated (86)Rb(+) uptake than cholesterol. Furthermore, analysis of multiple sterols suggests that cholesterol-induced inhibition of KirBac1.1 channels is mediated by specific interactions rather than by changes in the physical properties of the lipid bilayer. In contrast to the inhibition of KirBac1.1 activity, cholesterol had no effect on the activity of reconstituted KscA channels (at up to 250 microg/mg of phospholipid). Taken together, these observations demonstrate that cholesterol suppresses Kir channels in a pure protein-lipid environment and suggest that the interaction is direct and specific.
Singh
Dev K
DK
Department of Medicine, University of Illinois, Chicago, Illinois 60612, USA. dksingh@uic.edu
Rosenhouse-Dantsker
Avia
A
Nichols
Colin G
CG
Enkvetchakul
Decha
D
Levitan
Irena
I
D001426:Bacterial Proteins / Q000037:antagonists & inhibitors*; D002784:Cholesterol / Q000502:physiology*; D008081:Liposomes; D021962:Membrane Microdomains; D024661:Potassium Channels, Inwardly Rectifying / Q000037:antagonists & inhibitors*; D011387:Prokaryotic Cells; D012414:Rubidium Radioisotopes / Q000493:pharmacokinetics
D001426:Bacterial Proteins; D008081:Liposomes; D024661:Potassium Channels, Inwardly Rectifying; D012414:Rubidium Radioisotopes; D002784:Cholesterol