Electrochemical measurement of membrane cholesterol correlates with CFTR function and is HDAC6-dependent

Background: Previous studies have demonstrated that CF epithelial cells exhibit increased cholesterol content at the plasma membrane compared to wild type controls as measured by electrochemical methods. Microtubule dysregulation that impacts intracellular transport has also been identified in CF cells and is reversible with histone deacetylase 6 (HDAC6) inhibition, a regulator of tubulin acetylation. The hypothesis of this study is that increased membrane cholesterol content in CF cells is dependent on HDAC6 regulation. Methods: Electrochemical measurement of membrane cholesterol in mouse trachea and in primary human CF bronchial epithelial cells is used to monitor CFTR correction and manipulation of cholesterol processing by HDAC6 inhibition. Results: Data demonstrate that induction of Cftr expression in an inducible CF mouse model restores tubulin acetylation levels and normalizes membrane cholesterol content. To test the relationship between tubulin acetylation, membrane cholesterol levels were measured in a CF mouse model depleted of Hdac6 expression (CF/HDA). CF/HDA mouse trachea have WT membrane cholesterol levels while CF mice have approximately two-fold increase in membrane cholesterol compared to WT consistent with previous studies. Pharmacological inhibition of HDAC6 in primary human CF bronchial epithelial cells also reduces membrane cholesterol levels. Conclusions: This study demonstrates that elevated membrane cholesterol in CF epithelium is regulated by HDAC6 function and that the electrochemical measure of membrane cholesterol correlates with both genetic and pharmacological CFTR correction.