The lipidomes of vesicular stomatitis virus, semliki forest virus, and the host plasma membrane analyzed by quantitative shotgun mass spectrometry

Abstract

Although enveloped virus assembly in the host cell is a crucial step in the virus life cycle, it remains poorly understood. One issue is how viruses include lipids in their membranes during budding from infected host cells. To analyze this issue, we took advantage of the fact that baby hamster kidney cells can be infected by two different viruses, namely, vesicular stomatitis virus and Semliki Forest virus, from the Rhabdoviridae and Togaviridae families, respectively. We purified the host plasma membrane and the two different viruses after exit from the host cells and analyzed the lipid compositions of the membranes by quantitative shotgun mass spectrometry. We observed that the lipid compositions of these otherwise structurally different viruses are virtually indistinguishable, and only slight differences were detected between the viral lipid composition and that of the plasma membrane. Taken together, the facts that the lipid compositions of the two viruses are so similar and that they strongly resemble the composition of the plasma membrane suggest that these viruses exert little selection in including lipids in their envelopes.

FIG. 1.

SDS-PAGE analysis of purified viruses. Approximately 1 μg of final virus preparation was loaded onto a NuPAGE 4 to 12% BisTris gel, run in morpholinepropanesulfonic acid (MOPS) buffer, and stained with silver nitrate.

FIG. 2.

Western blot analysis of total cells and purified PM. About 500 ng of protein (total cells; lanes T) and 5% purified PM (lanes PM) were loaded onto 4 to 12% BisTris NuPAGE gels, run in MOPS buffer, and blotted onto a nitrocellulose membrane. The membrane was cut along the marker (lane M), and the halves were developed separately, using polyclonal anti-calnexin (αCNX) antibody produced in rabbits with HRP-conjugated anti-rabbit antibody and HRP-conjugated ExtrAvidin biotin-binding protein.

FIG. 3.

Lipid class compositions of BHK cells, PM, VSV, and SFV. Lipid class average compositions were determined by adding the abundances of individual lipid species from each class (Fig. 5; see Fig. S2 to S8 in the supplemental material). Error bars are the correspondent standard deviations (n = 4 for each sample).

FIG. 4.

(A) Enrichment plot of PM versus BHK cells. (B) Enrichment plot of VSV versus PM and SFV versus PM. Enrichment is defined as the ratio of the average quantity of each lipid class in a given sample to the average quantity of each lipid class in another sample. Only the statistically significant (by ANOVA plus Tuckes test) changes are shown. *, P < 0.05; **, P < 0.01; ***, P < 0.001.

FIG. 5.

Molecular composition of major lipid classes. The abundance of each lipid class is represented in mol% (absolute amount of lipid species/absolute amount of total identified lipid species).

FIG. 6.

Profiling of glycerophospholipid chain length (A) and saturation (B). The relative abundances of each chain length and unsaturation state were normalized to the total glycerophospholipid in each sample.