Depletion of Membrane Cholesterol Suppresses Drosophila Transient Receptor Potential-Like (TRPL) Channel Activity.

Abstract

Cholesterol is an essential compound of higher eukaryotic cell membranes and a known modulator of ion channel activity. Changes in phospholipids and cholesterol composition of cell membranes are known to alter the activity of ion channels. However, there is little knowledge on the effects of cholesterol on transient receptor potential (TRP) channels. In this study, we explore the effects of cholesterol depletion on the Drosophila photoreceptor channel TRP-like (TRPL), when expressed in tissue culture cells. Depletion of membrane cholesterol with methyl-β-cyclodextrin (MβCD) induced fast (<100s) suppression of spontaneous TRPL channel activity, a typical state of expressed TRPL channels in Drosophila S2 cells. An equally fast suppression of receptor-induced TRPL channel activity in HEK293 cells, downstream of phospholipase C (PLC) activation, was also induced by MβCD. Biochemical experiments showed binding of TRPL to immobilized cholesterol, suggesting direct binding of cholesterol to TRPL. Exploring the effects of several mutations in a putative cholesterol-binding site of TRPL was inconclusive as some did not render the channel insensitive to cholesterol depletion while others rendered the channel inactive. We conclude that (i) cholesterol is essential for TRPL channel activity, (ii) TRPL channels interact with cholesterol, and (iii) the binding site of cholesterol in TRPL differs from the putative binding site of TRPV1. Thus, the fast and strong effects of cholesterol depletion on the TRPL channel activity suggest that cholesterol is an important component of fly photoreceptor signaling membrane.