Human CFTR Channel Function is Regulated by Cholesterol

Cholesterol, a major membrane lipid component, has been known to modulate the function of multiple ion channels by specific lipid-protein interactions, by physical property changes of the membrane, or by modification of regulatory proteins associated with the channels in signaling complexes. Previous studies show that epithelial cells expressing the most common mutant version of the Cystic Fibrosis Transmembrane conductance Regulator (CFTR), F508del-CFTR, exhibit increased cholesterol content at the plasma membrane compared to wild type control cells; however, neutrophils derived from CF patients show reduced cholesterol levels in the plasma membrane. But whether cholesterol directly modulates CFTR channel function remains unknown. To answer this question, we combine heterologous expression in oocytes with more physiologically relevant Ussing chamber recordings utilizing polarized Fischer Rat Thyroid (FRT) cells to determine the effects of changing plasma membrane cholesterol levels on CFTR channel function. Here, we report that cholesterol depletion with methyl-β-cyclodextrin (MβCD) or cholesterol oxidase (CO) has no macroscopic effect on the magnitude of CFTR-mediated whole-oocyte currents. However, depletion of cholesterol increased the effect of VX-770, a clinically-used potentiator of CFTR, when channels were activated at high PKA concentrations, but did not change the effect of VX-770 when channels were activated at low PKA concentrations. This change in efficacy of VX-770-mediated potentiation likely reflects the apparent shift in the sensitivity of WT-CFTR to PKA after depletion of membrane cholesterol. In FRT cells, P67L-CFTR also exhibited changes in phosphorylation-dependent activation after depletion of plasma membrane cholesterol. Sensitivity of WT-CFTR to block by GlyH-101 was significantly stronger after plasma membrane cholesterol depletion. These results demonstrate that changes in the cholesterol level of the plasma membrane significantly modulate multiple CFTR channel functions. (Support: CF Fdn. MCCART17G0, NIH T32 GM008602, NIH F31 HL143863-01)