Cholesterol Inhibition of Slo1 Channels Is Calcium-Dependent and Can Be Mediated by Either High-Affinity Calcium-Sensing Site in the Slo1 Cytosolic Tail.

Autor:	North KC; Department of Pharmacology, Addiction Science, and Toxicology, College of Medicine, University of Tennessee Health Science Center, Memphis, Tennessee., Zhang M; Department of Pharmacology, Addiction Science, and Toxicology, College of Medicine, University of Tennessee Health Science Center, Memphis, Tennessee., Singh AK; Department of Pharmacology, Addiction Science, and Toxicology, College of Medicine, University of Tennessee Health Science Center, Memphis, Tennessee., Zaytseva D; Department of Pharmacology, Addiction Science, and Toxicology, College of Medicine, University of Tennessee Health Science Center, Memphis, Tennessee., Slayden AV; Department of Pharmacology, Addiction Science, and Toxicology, College of Medicine, University of Tennessee Health Science Center, Memphis, Tennessee., Bukiya AN; Department of Pharmacology, Addiction Science, and Toxicology, College of Medicine, University of Tennessee Health Science Center, Memphis, Tennessee., Dopico AM; Department of Pharmacology, Addiction Science, and Toxicology, College of Medicine, University of Tennessee Health Science Center, Memphis, Tennessee adopico@uthsc.edu.
Jazyk:	angličtina
Zdroj:	Molecular pharmacology [Mol Pharmacol] 2022 Mar; Vol. 101 (3), pp. 132-143. Date of Electronic Publication: 2021 Dec 30.
DOI:	10.1124/molpharm.121.000392
Abstrakt:	Calcium- and voltage-gated K + channels of large conductance (BKs) are expressed in the cell membranes of all excitable tissues. Currents mediated by BK channel-forming slo1 homotetramers are consistently inhibited by increases in membrane cholesterol (CLR). The molecular mechanisms leading to this CLR action, however, remain unknown. Slo1 channels are activated by increases in calcium (Ca 2+ ) nearby Ca 2+ -recognition sites in the slo1 cytosolic tail: one high-affinity and one low-affinity site locate to the regulator of conductance for K + (RCK) 1 domain, whereas another high-affinity site locates within the RCK2 domain. Here, we first evaluated the crosstalking between Ca 2+ and CLR on the function of slo1 (cbv1 isoform) channels reconstituted into planar lipid bilayers. CLR robustly reduced channel open probability while barely decreasing unitary current amplitude, with CLR maximal effects being observed at 10-30 µ M internal Ca 2+ CLR actions were not only modulated by internal Ca 2+ levels but also disappeared in absence of this divalent. Moreover, in absence of Ca 2+ , BK channel-activating concentrations of magnesium (10 mM) did not support CLR action. Next, we evaluated CLR actions on channels where the different Ca 2+ -sensing sites present in the slo1 cytosolic domain became nonfunctional via mutagenesis. CLR still reduced the activity of low-affinity Ca 2+ (RCK1:E379A, E404A) mutants. In contrast, CLR became inefficacious when both high-affinity Ca 2+ sites were mutated (RCK1:D367A,D372A and RCK2:D899N,D900N,D901N,D902N,D903N), yet still was able to decrease the activity of each high-affinity site mutant. Therefore, BK channel inhibition by CLR selectively requires optimal levels of Ca 2+ being recognized by either of the slo1 high-affinity Ca 2+ -sensing sites. SIGNIFICANCE STATEMENT: Results reveal that inhibition of calcium/voltage-gated K + channel of large conductance (BK) (slo1) channels by membrane cholesterol requires a physiologically range of internal calcium (Ca 2+ ) and is selectively linked to the two high-affinity Ca 2+ -sensing sites located in the cytosolic tail domain, which underscores that Ca 2+ and cholesterol actions are allosterically coupled to the channel gate. Cholesterol modification of BK channel activity likely contributes to disruption of normal physiology by common health conditions that are triggered by disruption of cholesterol homeostasis. (Copyright © 2022 by The American Society for Pharmacology and Experimental Therapeutics.)
Databáze:	MEDLINE
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