Palmitoylation of Influenza virus hemagglutinin: search for DHHC-acyltransferases in human airway cells as potential targets for new antiviral drugs.
Veterinary Medical Science
Final Report Abstract
The research project yielded significant insights into the fatty acid attachment to membrane proteins of Influenza and Coronaviruses and the host DHHC enzymes involved in this process. Comparative analyses across virus genera showed conservation of acylation sites (cysteine residues at the beginning of the cytoplasmic tails) in hemagglutinin (HA) of Influenza A and B viruses and in the Spike of coronaviruses, confirming their fundamental role in viral replication. Experimental findings revealed that DHHC22 is upregulated in influenza A virus-infected cells, with this upregulation mediated by the viral protein NS1. Interestingly, knockout of DHHC22 did not affect the acylation of HA. However, we identified 25 potential cellular substrates of ZDHHC22, suggesting that its role in viral replication may be indirect. In our systematic effort to identify the specific DHHC enzymes responsible for HA acylation in influenza A, we found that DHHC2, DHHC8, DHHC15, and DHHC20 were key players in this process. These enzymes were also shown to acylate the M2 protein, although they exhibited slight differences in substrate preferences. Molecular dynamics simulations of DHHC20 indicated that its hydrophobic cavity accommodates C16 acyl chains optimally, while a mutation that enlarges the cavity shifted its preference towards C18 acyl chains, corroborating predictions based on its crystal structure. In terms of influenza B virus, we established that HA acylation is crucial for viral replication and occurs in the endoplasmic reticulum (ER), contrasting with the mainly Golgi-based process observed in influenza A. We identified ER-localized DHHCs 1, 2, 4, and 6 as responsible for HA acylation in influenza B. The differences in acylation mechanisms between influenza A and B viruses illustrate virus-specific adaptations. Next, we analysed how the DHHC enzyme 20 recognize their substrates. The acylation of M2 is significantly reduced, but not completely abolished if its amphiphilic helix that carries the fatty acid is altered. Molecular dynamics simulations revealed interactions between amino acids of the helix and the catalytic motifs of DHHC20, which might facilitate fatty acid transfer. Similar studies with the spike of SARS-CoV-2 revealed that specific transmembrane residues are required for efficient acylation and DHHC20 binding. In addition, conserved serine and threonine residues within the acylated cysteine cluster affect acylation and they might bind to a hydrophilic ridge in one transmembrane helix of DHHC20. Overall, our research highlights the importance of specific DHHC enzymes in viral protein acylation. These findings present these DHHC enzymes as potential targets for the development of pan-antiviral drugs, which could be effective against a broad range of viruses that rely on S-acylation for their life cycle.
Publications
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Toward the identification of ZDHHC enzymes required for palmitoylation of viral protein as potential drug targets. Expert Opinion on Drug Discovery, 15(2), 159-177.
Gadalla, Mohamed Rasheed & Veit, Michael
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Hemagglutinin of Influenza A, but not of Influenza B and C viruses is acylated by ZDHHC2, 8, 15 and 20. Biochemical Journal, 477(1), 285-303.
Gadalla, Mohamed Rasheed; Abrami, Laurence; van der Goot, F. Gisou & Veit, Michael
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NS1 ‐mediated upregulation of ZDHHC22 acyltransferase in influenza a virus infected cells. Cellular Microbiology, 23(6).
Gadalla, Mohamed Rasheed; Morrison, Eliot; Serebryakova, Marina V.; Han, Xueijiao; Wolff, Thorsten; Freund, Christian; Kordyukova, Larisa & Veit, Michael
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S-Acylation of Proteins of Coronavirus and Influenza Virus: Conservation of Acylation Sites in Animal Viruses and DHHC Acyltransferases in Their Animal Reservoirs. Pathogens, 10(6), 669.
Abdulrahman, Dina A.; Meng, Xiaorong & Veit, Michael
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Molecular Dynamics of DHHC20 Acyltransferase Suggests Principles of Lipid and Protein Substrate Selectivity. International Journal of Molecular Sciences, 23(9), 5091.
Panina, Irina; Krylov, Nikolay; Gadalla, Mohamed Rasheed; Aliper, Elena; Kordyukova, Larisa; Veit, Michael; Chugunov, Anton & Efremov, Roman
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Palmitoylation of the hemagglutinin of influenza B virus by ER-localized DHHC enzymes 1, 2, 4, and 6 is required for efficient virus replication. Journal of Virology, 97(10).
Meng, Xiaorong & Veit, Michael
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The role of an amphiphilic helix and transmembrane region in the efficient acylation of the M2 protein from influenza virus. Scientific Reports, 13(1).
Meng, Xiaorong; Templeton, Clark; Clementi, Cecilia & Veit, Michael
