Anionic Phospholipids Bind to and Modulate the Activity of Human TRESK Background K+ Channel

Abstract

The background K+ channel TRESK regulates sensory neuron excitability and changes in its function/expression contribute to neuronal hyperexcitability after injury/inflammation, making it an attractive therapeutic target for pain-related disorders. Factors that change the plasma membrane bilayer composition/properties (including volatile anesthetics, chloroform, chlorpromazine, shear stress and cell swelling/shrinkage) modify TRESK current but despite the importance of anionic phospholipids (e.g. PIP2) in the regulation of many ion channels, it remains unknown if membrane lipids affect TRESK function. We describe that both human and rat TRESK contain potential anionic phospholipid binding sites (apbs) in the large cytoplasmic loop, but only the human channel is able to bind to multilamellar vesicles (MLVs), enriched with anionic phospholipids, suggesting an electrostatically-mediated interaction. We mapped the apbs to a short stretch of 14 amino acids in the loop, located at the membrane-cytosol interface. Disruption of electrostatic lipid-TRESK interactions inhibited hTRESK currents, whilst subsequent application of Folch Fraction MLVs or a PIP2 analog activated hTRESK, an effect that was absent in the rat ortholog. Strikingly, channel activation by anionic phospholipids was conferred to rTRESK by replacing the equivalent rat sequence with the human apbs. Finally, stimulation of a Gq/11-linked GPCR reduced hTRESK current when Ca2+/calcineurin is blocked, while in physiological conditions, the Ca2+-mediated stimulation is prominent. This novel regulation of hTRESK by anionic phospholipids is a characteristic of the human channel that is not present in rodent orthologs. This must be considered when extrapolating results from animal models and may open the door to the development of novel channel modulators as analgesics.