Ebola Virus Glycoprotein Interacts with Cholesterol to Enhance Membrane Fusion and Cell Entry

Abstract

Cholesterol serves critical roles in enveloped virus fusion by modulating membrane biophysical properties. The glycoprotein (GP) of Ebola virus (EBOV) promotes fusion in the endosome after internalization of the virus by macropinocytosis. Fusion of EBOV requires the endosomal cholesterol transporter NPC1, but the role of cholesterol in EBOV fusion is unclear. Here we show that the membrane proximal external region and transmembrane domain (MPER/TM) of GP interacts with cholesterol, and that cell entry of virus-like particles (VLPs) and GP2-mediated fusion to supported membranes are enhanced with increasing cholesterol in EBOV membranes. NMR-based experiments revealed that several glycines, notably G660, in GP2 TM are critical for interaction with cholesterol. Moreover, G660L MPER/TM shows a more open angle between MPER and TM domains compared to WT. G660L particles show a higher probability of hemifusion (vs. full fusion), correlating with a lower efficiency of cell entry by G660L vs. WT VLPs. VLPs produced under cholesterol-lowering statin conditions show less entry than VLPs from mock-treated cells. We propose that cholesterol-TM interactions affect structural features of GP2 thereby facilitating fusion and cell entry and that statins may be used as part of EBOV treatments due to their ability to lower cholesterol in the viral membrane.