Cholesterol and ion channels

Frontiers in Physiology, 2014

Numerous studies demonstrated that membrane cholesterol is a major regulator of ion channel function. The goal of this review is to discuss significant advances that have been recently achieved in elucidating the mechanisms responsible for cholesterol regulation of ion channels. The first major insight that comes from growing number of studies that based on the sterol specificity of cholesterol effects, show that several types of ion channels (nAChR, Kir, BK, TRPV) are regulated by specific sterol-protein interactions. This conclusion is supported by demonstrating direct saturable binding of cholesterol to a bacterial Kir channel. The second major advance in the field is the identification of putative cholesterol binding sites in several types of ion channels. These include sites at locations associated with the well-known cholesterol binding motif CRAC and its reversed form CARC in nAChR, BK, and TRPV, as well as novel cholesterol binding regions in Kir channels. Notably, in the majority of these channels, cholesterol is suggested to interact mainly with hydrophobic residues in non-annular regions of the channels being embedded in between transmembrane protein helices. We also discuss how identification of putative cholesterol binding sites is an essential step to understand the mechanistic basis of cholesterol-induced channel regulation. Clearly, however, these are only the first few steps in obtaining a general understanding of cholesterol-ion channels interactions and their roles in cellular and organ functions.

Advances in Experimental Medicine and Biology, 2019

Biomembranes separate a live cell from its environment and keep it in an off-equilibrium, steady state. They contain both phospholipids and nonphospholipids, depending on whether there are phosphate groups in the headgroup regions. Cholesterol (CHOL) is one type of nonphospholipids, and one of the most abundant lipid molecules in humans. Its content in plasma membranes and intracellular membranes varies and is tightly regulated. Voltage-gated ion channels are universally present in every cell and are fairly diversified in the eukaryotic domain of life. Our lipid-dependent gating hypothesis postulates that the controlled switch of the voltage-sensor domains (VSDs) in a voltage-gated potassium (Kv) channel between the "down" and the "up" state (gating) is sensitive to the ratio of phospholipids : nonphospholipids in the annular layer around the channel. High CHOL content is found to exert strong inhibitory effects on Kv channels. Such effects have been observed in in vitro membranes, cultured cells and animal models for cholesterol metabolic defects. Thermodynamic analysis of the CHOL-dependent gating suggests that the inhibitory effects of CHOL result from collective interactions between annular CHOL molecules and the channel, which appear to be a more generic principle behind the CHOL-effects on other ion channels and transporters. We will review the recent progress in the CHOL-dependent gating of voltage-gated ion channels, discuss the current technical limitations, and then expand briefly the learned principles to other ion channels that are known to be sensitive to the CHOL-channel interactions.

Journal of Membrane Biology, 1995

The ubiquity of cholesterol in cell membranes and changes in its concentration during development, aging and in various diseases suggest that it plays an important role in modulating cell function. We examined this possibility by monitoring the effects of cholesterol on the activity of the calcium-activated potassium (BK) channel reconstituted into lipid bilayers from rat brain homogenates. Increasing the cholesterol concentration to 11% of total lipid weight resulted in a 70% reduction in channel mean open time and a reduction of the open probability of the channel by 80%. Channel conductance was reduced by 7%. Cholesterol is known to change the order state and the modulus of compressibility of bilayers. These physico-chemical changes may be translated into an overall increase in the structural stress in the bilayer, and this force may be transmitted to proteins residing therein. By examining the characteristics of the BK channel as a function of temperature, in the presence and absence of cholesterol, we were able to estimate the activation energy based on Arrhenius plots of channel kinetics. Cholesterol reduced the activation energy of the BK channel by 50% for the open to closed transition. This result is consistent with an increased stress energy in the bilayer and favors the channel moving into the closed state. Taken together, these data are consistent with a model in which cholesterol induces structural stress which enhances the transition from the open to the closed state of the channel. We suggest that this is an important mechanism for regulating the activity of membrane-integral proteins and therefore membrane function, and that the concept of structural stress may be relevant to understanding the modulation of ion channel activity in cell membranes.

Pflügers Archiv : European journal of physiology, 1989

The patch-clamp technique and fluorescence polarization analysis were used to study the dependence of Ca2(+)-dependent K+ channel kinetics and membrane fluidity on cholesterol (CHS) levels in the plasma membranes of cultured smooth muscle rabbit aortic cells. Mevinolin (MEV), a potent inhibitor of endogenous CHS biosynthesis was used to deplete the CHS content. Elevation of CHS concentration in the membrane was achieved using a CHS-enriching medium. Treatment of smooth muscle cells with MEV led to a nearly twofold increase in the rotational diffusion coefficient of DPH (D) and to about a ninefold elevation of probability of the channels being open (Po). The addition of CHS to the cells membrane resulted in a nearly twofold decrease in D and about a twofold decrease in Po. Elementary conductance of the channels did not change under these conditions. These data suggest that variations of the CHS content in the plasma membrane of smooth muscle cells affect the kinetic properties of Ca2...

F1000 - Post-publication peer review of the biomedical literature, 2012

Many types of ion channel localize to cholesterol and sphingolipid-enriched regions of the plasma membrane known as lipid microdomains or 'rafts'. The precise physiological role of these unique lipid microenvironments remains elusive due largely to difficulties associated with studying these potentially extremely small and dynamic domains. Nevertheless, increasing evidence suggests that membrane rafts regulate channel function in a number of different ways. Raft-enriched lipids such as cholesterol and sphingolipids exert effects on channel activity either through direct protein-lipid interactions or by influencing the physical properties of the bilayer. Rafts also appear to selectively recruit interacting signalling molecules to generate subcellular compartments that may be important for efficient and selective signal transduction. Direct interaction with raft-associated scaffold proteins such as caveolin can also influence channel function by altering gating kinetics or by affecting trafficking and surface expression. Selective association of ion channels with specific lipid microenvironments within the membrane is thus likely to be an important and fundamental regulatory aspect of channel physiology. This brief review highlights some of the existing evidence for raft modulation of channel function.

Journal of Biological Chemistry, 2011

Interaction of large conductance Ca 2؉ -and voltage-activated K ؉ (BK Ca ) channels with Na ؉ /K ؉ -ATPase, caveolin-1, and cholesterol was studied in human melanoma IGR39 cells. Functional BK Ca channels were enriched in caveolin-rich and detergent-resistant membranes, i.e. rafts, and blocking of the channels by a specific BK Ca blocker paxilline reduced proliferation of the cells. Disruption of rafts by selective depletion of cholesterol released BK Ca channels from these domains with a consequent increase in their activity. Consistently, cholesterol enrichment of the cells increased the proportion of BK Ca channels in rafts and decreased their activity. Immunocytochemical analysis showed that BK Ca channels co-localize with Na ؉ /K ؉ -ATPase in a cholesterol-dependent manner, thus suggesting their co-presence in rafts. Supporting this, ouabain, a specific blocker of Na ؉ /K ؉ -ATPase, inhibited BK Ca whole-cell current markedly in control cells but not in cholesterol-depleted ones. This inhibition required the presence of external Na ؉ . Collectively, these data indicate that the presence of Na ؉ /K ؉ -ATPase in rafts is essential for efficient functioning of BK Ca channels, presumably because the pump maintains a low intracellular Na ؉ proximal to the BK Ca channel. In conclusion, cholesterol could play an important role in cellular ion homeostasis and thus modulate many cellular functions and cell proliferation.

Journal of Biological Chemistry, 2012

Background: Cholesterol regulation of large conductance, Ca 2ϩ-and voltage-gated K ϩ (BK) channels has widespread pathophysiological consequences. Results: Cholesterol-channel recognition involves hydrophobic and hydrophilic interactions and several cholesterol recognition/interaction amino acid consensus motifs in the BK channel long C-end. Conclusion: Cholesterol regulation of BK channels involves specific channel protein-sterol recognition. Significance: we provide for the first time the structural basis of BK channel cholesterol sensitivity. Large conductance, Ca 2؉-and voltage-gated K ؉ (BK) channel proteins are ubiquitously expressed in cell membranes and control a wide variety of biological processes. Membrane cholesterol regulates the activity of membrane-associated proteins, including BK channels. Cholesterol modulation of BK channels alters action potential firing, colonic ion transport, smooth muscle contractility, endothelial function, and the channel alcohol response. The structural bases underlying cholesterol-BK channel interaction are unknown. Such interaction is determined by strict chemical requirements for the sterol molecule, suggesting cholesterol recognition by a protein surface. Here, we demonstrate that cholesterol action on BK channel-forming Cbv1 proteins is mediated by their cytosolic C tail domain, where we identified seven cholesterol recognition/interaction amino acid consensus motifs (CRAC4 to 10), a distinct feature of BK proteins. Cholesterol sensitivity is provided by the membrane-adjacent CRAC4, where Val-444, Tyr-450, and Lys-453 are required for cholesterol sensing, with hydrogen bonding and hydrophobic interactions participating in cholesterol location and recognition. However, cumulative truncations or Tyr-to-Phe substitutions in CRAC5 to 10 progressively blunt cholesterol sensitivity, documenting involvement of multiple CRACs in cholesterol-BK channel interaction. In conclusion, our study provides for the first time the structural bases of BK channel cholesterol sensitivity; the presence of membrane-adjacent CRAC4 and the long cytosolic C tail domain with several other CRAC motifs, which are not found in other members of the TM6 superfamily of ion channels, very likely explains the unique cholesterol sensitivity of BK channels. Cholesterol (CLR) 2 is a major constituent of plasma membranes in eukaryotes and crucial in membrane organization, sorting, dynamics, and function (1). In particular, CLR plays a critical role in regulating the activity of membrane-associated proteins, including ion channels (2-7). Large conductance, Ca 2ϩ-and voltage-gated K ϩ (BK) channels belong to the TM6 superfamily of ion channel proteins. BK channels are ubiquitously expressed in cell membranes and regulate a wide variety of processes, including neuronal excitability, neurotransmitter release, neurosecretion, tuning of cochlear hair cells, smooth muscle tone, immune responses, apoptosis, and brain tumor metastasis (7-10). Moreover, CLR modulation of BK currents has been linked to changes in neuroendocrine GH3 cell action potential firing rate (11), ion transport in colonic epithelial cells (12), membrane smooth muscle excitability and uterine contractility (13), endothelial function (14), vascular myocyte signaling (15), endothelium-dependent and-independent vasodilation (16), and BK channel responses to ethanol and eventual alcohol-induced cerebrovascular constriction (17). Despite the wide range of pathophysiological implications of CLR action on BK currents, which usually results in reduced ionic current (7), the mechanisms and structural bases underlying the CLR-BK channel interaction remain unknown. Functional BK channels result from the tetrameric association of channel-forming ␣ subunits (Fig. 1a), encoded by the * This work was supported, in whole or in part, by National Institutes of Health Grants 2R37 AA011560 and R01 HL104632 (to A. M. D.). This work was also supported by a University of Tennessee Health Science Center Neuroscience Institute postdoctoral fellowship (to A. K. S.). □ S This article contains supplemental Figs. S1 and S2 and Movies S1-S4.

2018

Pharmacology & Therapeutics, 2012

Cholesterol (CLR) is an essential component of eukaryotic plasma membranes. CLR regulates the membrane physical state, microdomain formation and the activity of membrane-spanning proteins, including ion channels. Large conductance, voltage-and Ca 2+-gated K + (BK) channels link membrane potential to cell Ca 2+ homeostasis. Thus, they control many physiological processes and participate in pathophysiological mechanisms leading to human disease. Because plasmalemma BK channels cluster in CLR-rich membrane microdomains, a major driving force for studying BK channel-CLR interactions is determining how membrane CLR controls the BK current phenotype, including its pharmacology, channel sorting, distribution, and role in cell physiology. Since both BK channels and CLR tissue levels play a pathophysiological role in human disease, identifying functional and structural aspects of the CLR-BK channel interaction may open new avenues for therapeutic intervention. Here, we review the studies documenting membrane CLR-BK channel interactions, dissecting out the many factors that determine the final BK current response to changes in membrane CLR content. We also summarize work in reductionist systems where recombinant BK protein is studied in artificial lipid bilayers, which documents a direct inhibition of BK channel activity by CLR and builds a strong case for a direct interaction between CLR and the BK channel-forming protein. Bilayer lipid-mediated mechanisms in CLR action are also discussed. Finally, we review studies of BK channel function during hypercholesterolemia, and underscore the many consequences that the CLR-BK channel interaction brings to cell physiology and human disease.

Circulation Research, 1992

To determine whether membrane free cholesterol affects calcium currents in vascular smooth muscle cells, whole-cell patch clamp recordings were made before and after cholesterol enrichment of cells by exposure to cholesterol-rich liposomes. Exposure to cholesterol-rich liposomes resulted in a gradual increase in the L-type current over 20 hours and a plateau (73 +7% increase over basal) between 20 and 32 hours. This effect was associated with a rightward shift in the inactivation potential and a decrease in the sensitivity to (-)-PN-202-791, a dihydropyridine antagonist. There was no change in the maximum L-type current stimulated by (+)-PN-202-791, a dihydropyridine agonist. Liposome exposure caused a small, transient increase in the T-type current (peak effect, 20 minutes). We conclude that membrane cholesterol has important effects on the L-type calcium current in vascular smooth muscle cells, which is most likely due to an alteration in channel functional state rather than an increase in channel expression.