The influenza virus acquires its lipid envelope when it buds from the host cell’s plasma membrane. Intriguingly, the influenza viral envelope is enriched with cholesterol and sphingolipids, and depletion of either cellular cholesterol or sphingolipids is detrimental to influenza virus replication. These observations led to the long-standing hypothesis that the influenza viral proteins are recruited to ordered cholesterol- and sphingolipid-enriched domains in the plasma membrane for virus particle assembly and budding. Though this hypothesis is well supported by indirect evidence, whether cholesterol and sphingolipids are enriched at the site of influenza budding has not been decisively tested due to the difficulty of simultaneously imaging cholesterol and sphingolipids within the plasma membrane. We have overcome this obstacle by developing a secondary ion mass spectrometry (SIMS) approach to image metabolically incorporated, stable isotope-labeled lipids in the plasma membrane with a lateral resolution of ~100 nm. Here we have imaged metabolically incorporated <sup>15</sup>N-sphingolipids and <sup>18</sup>O-cholesterol as well as immunolabeled viral envelope proteins on the surfaces of influenza-infected MDCK cells, and assessed their co-localization. The results of this decisive test of the long-standing hypothesis that the influenza virus buds from cholesterol- and sphingolipid-enriched membrane domains will be presented

Chemical imaging of the lipid composition at the site of influenza budding

253rd National Meeting (2017)

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