Involvement of Lipids in Different Steps of the Flavivirus Fusion Mechanism

Flavivirus membrane fusion is triggered by acidic pH and mediated by the major envelope protein E. A structurally very similar fusion protein is found in alphaviruses, and these molecules are designated class II viral fusion proteins. In contrast to that of flaviviruses, however, alphavirus fusion has been shown to be absolutely dependent on the presence of cholesterol and sphingomyelin in the target membrane, suggesting significant differences in the fusion protein-membrane interactions that lead to fusion. With the flavivirus tick-borne encephalitis virus (TBEV), we have therefore conducted a study on the lipid requirements of viral fusion with liposomes and on the processes preceding fusion, specifically, the membrane-binding step and the fusion-associated oligomeric switch from E protein dimers to trimers. As with alphaviruses, cholesterol had a strong promoting effect on membrane binding and trimerization of the fusion protein, and-as shown by the use of cholesterol analogs-the underlying interactions involve the 3beta-hydroxyl group at C-3 in both viral systems. In contrast to alphaviruses, however, these effects are much less pronounced with respect to the overall fusion of TBEV and can only be demonstrated when fusion is slowed down by lowering the temperature. The data presented thus suggest the existence of structurally related interactions of the flavivirus and alphavirus fusion proteins with cholesterol in the molecular processes required for fusion but, at the same time, point to significant differences between the class II fusion machineries of these viruses.