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bioRxiv · 10.1101/2023.06.15.545201

Palmitoylation of the Hemagglutinin of Influenza B virus by ER-localized DHHC enzymes 1, 2, 4 and 6 is essential for virus replication

Meng, X.·Veit, M.
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Abstract

Covalent attachment of the fatty acids palmitate or stearate to the cytoplasmic domain of viral glycoproteins is often crucial for viral replication. This has previously been studied for the hemagglutinin (HA) of Influenza A virus and the responsible enzymes have been identified, but similar studies have not been performed with HA of Influenza B virus, which contains palmitate linked to two cysteines. We show here that the modification is essential for virus replication since exchange of both cysteines or the cysteine located at the end of the cytoplasmic tail prevented the generation of viable viruses. Viruses with an exchange of the membrane-distal cysteine rapidly reverted back to wild-type virus. Blocking exit of proteins from the endoplasmic reticulum (ER) revealed that palmitoylation of HA of Influenza B virus occurs in the ER, whereas acylation of HA of Influenza A virus also in the Golgi. Infecting cells deficient in DHHC palmitoyltransferases revealed that HA of Influenza B virus is acylated by the ER-localized DHHCs 1, 2, 4 and 6, which are thus different from the enzymes previously identified for acylation of HA of Influenza A virus. A comparison of predicted and experimentally determined protein structures suggests that the exclusive acylation of the HA of Influenza B virus with palmitate is not a function of the responsible DHHCs and that the transmembrane region may be critical for the acylation of HA of Influenza A and B virus by different DHHCs. ImportanceInfluenza viruses are a public health concern since they cause seasonal outbreaks and occasionally pandemics. Our study investigates the importance of a protein modification called "palmitoylation" in the replication of Influenza B virus. Palmitoylation involves attaching fatty acids to the viral protein hemagglutinin, and has previously been studied for Influenza A virus. We found that this modification is essential for the Influenza B virus to replicate, as mutating the sites where palmitate is attached prevented the virus from generating viable particles. Our experiments also showed that this modification occurs in the endoplasmic reticulum. We identified the specific enzymes responsible for this modification, which are different from those involved in palmitoylation of HA of Influenza A virus. Overall, our research illuminates the similarities and differences in fatty acid attachment to HA of Influenza A and B virus and identifies the responsible enzymes, which might be promising targets for antiviral therapy.

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Meng, X., Veit, M.. 2023-06-16. Palmitoylation of the Hemagglutinin of Influenza B virus by ER-localized DHHC enzymes 1, 2, 4 and 6 is essential for virus replication. https://doi.org/10.1101/2023.06.15.545201

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