The Role of Cholesterol in Viral Spike Glycoprotein-Mediated Membrane Fusion

Abstract

Viral spike glycoproteins such as influenza HA, HIV gp120/gp41, and Ebola virus (EBOV) GP1/GP2 undergo major conformational transitions to facilitate cell entry of these viruses by membrane fusion. Moderately hydrophobic fusion peptides [or fusion loops (FL) in the case of EBOV] are protected by these proteins in their pre-fusion states, but become exposed and insert into cellular target membranes during fusion. We have solved the structures of the EBOV FL, membrane proximal external region (MPER) and transmembrane (TM) domains in membrane environments by NMR and studied their molecular interactions by NMR and fluorescence. A structural model of how these domains cooperate to open a fusion pore emerges from these studies. Viral membrane fusion is modulated by cholesterol in the viral and target cell membranes. Interestingly, in the case of HIV gp41-mediated fusion, membranes that consist of co-existing liquid-ordered (Lo) and liquid-disordered (Ld) bilayer domains are more fusogenic than either pure Ld or Lo lipid bilayers and lipid bilayer phase co-existence in both the viral and the target membrane synergistically helps fusion. Single particle experiments on supported bilayers and giant unilamellar vesicles show that the boundaries between the Lo and Ld regions are the sites of virosome and pseudovirus docking and fusion. It appears that line-tension that characterizes lipid phase boundaries is a major driving force of membrane fusion in cell entry of HIV and perhaps other enveloped viruses. Although these studies were performed using model systems with optically resolved lipid phase separations, it is plausible and likely that similar mechanisms operate in cell membranes that are also heterogeneous in lipid and protein composition and therefore also contain domain boundaries that are more frequent, finer grained, and more transient than in our model systems.