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This Article Full Text Full Text (PDF) All Versions of this Article: 571/2/303    most recent jphysiol.2005.100719v1 Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Ribalet, B. Articles by Weiss, J. N. Search for Related Content PubMed PubMed Citation Articles by Ribalet, B. Articles by Weiss, J. N. Related Collections Molecular and Genomic

¹ University of California Los Angeles Cardiovascular Research Laboratory
² Departments of Physiology
³ Medicine (Cardiology), David Geffen School of Medicine at University of California Los Angeles, Los Angeles, CA 90095, USA

ATP-sensitive K⁺ channels composed of the pore-forming protein Kir6.2 and the sulphonylurea receptor SUR1 are inhibited by ATP and activated by Phosphatidylinositol Bisphosphate (PIP₂). Residues involved in binding of these ligands to the Kir6.2 cytoplasmic domain have been identified, and it has been hypothesized that gating mechanisms involve conformational changes in the regions of the bundle crossing and/or the selectivity filter of Kir6.2. Regulation of Kir6.2 by SUR1, however, is not well-understood, even though this process is ATP and PIP₂ dependent. In this study, we investigated the relationship between channel regulation by SUR1 and PIP₂ by comparing a number of single and double mutants known to affect open probability (P_o), PIP₂ affinity, and sulphonylurea and MgADP sensitivity. When coexpressed with SUR1, the Kir6.2 mutant C166A, which is characterized by a P_o value close to 0.8, exhibits no sulphonylurea or MgADP sensitivity. However, when P_o was reduced by combining mutations at the PIP₂-sensitive residues R176 and R177 with C166A, sulphonylurea and MgADP sensitivities were restored. These effects correlated with a dramatic decrease in PIP₂ affinity, as assessed by PIP₂-induced channel reactivation and inhibition by neomycin, an antagonist of PIP₂ binding. Based on macroscopic and single-channel data, we propose a model in which entry into the high-P_o bursting state by the C166A mutation or by SUR1 depends on the interaction of PIP₂ with R176 and R177 and, to a lesser extent, R54. In conjunction with this PIP₂-dependent process, SUR1 also regulates channel activity via a PIP₂-independent, but MgADP-dependent process.

(Received 25 October 2005; accepted after revision 16 December 2005; first published online 22 December 2005)
Corresponding author B. Ribalet: Department of Physiology, School of Medicine, University of California Los Angeles, Los Angeles, CA 90095, USA. Email: bribalet{at}mednet.ucla.edu

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