Ebolavirus Requires Acid Sphingomyelinase Activity and Plasma Membrane Sphingomyelin for Infection

Acid sphingomyelinase (ASMase) converts the lipid sphingomyelin (SM) to phosphocholine and ceramide and has optimum activity at acidic pH. Normally, ASMase is located in lysosomes and endosomes, but membrane damage or the interaction with some bacterial and viral pathogens can trigger its recruitment to the plasma membrane. Rhinovirus and measles viruses each require ASMase activity during early stages of infection. Both sphingomyelin and ceramide are important components of lipid rafts and are potent signaling molecules. Each plays roles in mediating macropinocytosis, which has been shown to be important for ebolavirus (EBOV) infection. Here, we investigated the role of ASMase and its substrate, SM, in EBOV infection. The work was performed at biosafety level 4 with wild-type virus with specificity and mechanistic analysis performed using virus pseudotypes and virus-like particles. We found that virus particles strongly associate with the SM-rich regions of the cell membrane and depletion of SM reduces EBOV infection. ASM-specific drugs and multiple small interfering RNAs strongly inhibit the infection by EBOV and EBOV glycoprotein pseudotyped viruses but not by the pseudotypes bearing the glycoprotein of vesicular stomatitis virus. Interestingly, the binding of virus-like particles to cells is strongly associated with surface-localized ASMase as well as SM-enriched sites. Our work suggests that ASMase activity and SM presence are necessary for efficient infection of cells by EBOV. The inhibition of this pathway may provide new avenues for drug treatment.