Nicotinic acetylcholine receptor and cholesterol: lipid rafts, membrane asymmetry, structural conformation and functionality.

FABIANI, C.; CORRADI, J.; ANTOLLINI, S.S.

Castellon

Congreso; 6th International Iberian Biophysics Congress and X Iberoamerican Congress of Biophysics, Castellón, España; 2018

The muscle nicotinic acetylcholine receptor (AChR) has a neurotransmitter-binding site extracellular domain and a channel-gating transmembrane domain which exhibits extensive contacts with the surrounding lipids. A correct allosteric coupling between both domains is crucial for AChR function. Also, the AChR is present in high-density clusters in the muscle cell membrane which localize in lipid-ordered domains (Lo) enriched in cholesterol and sphingolipids. We studied the relationship between the AChR and cholesterol both in T. californica AChR-rich membranes and in model membranes containing purified AChR. Depletion of cholesterol by methyl--cyclodextrin; enrichment of cholesterol or cholesterol-hemisuccinate (in this last case, asymmetric membranes were obtained); and oxidation of cholesterol using cholesterol oxidase were correlated with AChR structural conformation by crystal violet fluorescent probe; AChR functionality by electrophysiology; augmentation/diminution of Lo domains by GUVs formation and fluorescence microscopy; and AChR location at these domains by FRET or by detergent treatment and SDS-PAGE. Altogether, we observed that a change in the amount, distribution or oxidation of cholesterol impacts in the size and location of Lo domains and in the AChR preference for them but also in the AChR functionality and AChR structural conformation, and hence in the coupling between its agonist-binding and its channel-gating domains.