title: Dual-mode phospholipid regulation of human inward rectifying potassium channels.
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.
authors.affiliation: Department of Cell Biology and Physiology, Washington University School of Medicine, St. Louis, Missouri; Center for Investigation of Membrane Excitability Diseases, Washington University School of Medicine, St. Louis, Missouri.
authors.forename: Wayland W L
authors.initials: WWL
authors.lastname: Cheng
authors.affiliation: Department of Cell Biology and Physiology, Washington University School of Medicine, St. Louis, Missouri; Center for Investigation of Membrane Excitability Diseases, Washington University School of Medicine, St. Louis, Missouri.
authors.forename: Nazzareno
authors.initials: N
authors.lastname: D'Avanzo
authors.affiliation: School of Biochemistry and Immunology, Trinity College Dublin, Dublin, Ireland.
authors.forename: Declan A
authors.initials: DA
authors.lastname: Doyle
authors.affiliation: Department of Cell Biology and Physiology, Washington University School of Medicine, St. Louis, Missouri; Center for Investigation of Membrane Excitability Diseases, Washington University School of Medicine, St. Louis, Missouri. Electronic address: cnichols@wustl.edu.
authors.forename: Colin G
authors.initials: CG
authors.lastname: Nichols
mesh_terms: D000214:Acyl Coenzyme A / Q000494:pharmacology; D000596:Amino Acids / Q000378:metabolism; D000818:Animals; D000838:Anions; D002417:Cattle; D006801:Humans; D042541:Intracellular Space / Q000187:drug effects / Q000378:metabolism; D015640:Ion Channel Gating / Q000187:drug effects; D010713:Phosphatidylcholines / Q000494:pharmacology; D010715:Phosphatidylglycerols / Q000378:metabolism; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000378:metabolism; D010743:Phospholipids / Q000378:metabolism*; D024661:Potassium Channels, Inwardly Rectifying / Q000037:antagonists & inhibitors / Q000378:metabolism*
chemical_list: D000214:Acyl Coenzyme A; D000596:Amino Acids; D000838:Anions; C489457:KCNJ2 protein, human; C489463:Kir2.2 channel; D010713:Phosphatidylcholines; D010715:Phosphatidylglycerols; D019269:Phosphatidylinositol 4,5-Diphosphate; D010743:Phospholipids; D024661:Potassium Channels, Inwardly Rectifying; C017585:oleoyl-coenzyme A; C060037:1-palmitoyl-2-oleoylglycero-3-phosphoglycerol