Institute of Virology, Free University Berlin, Berlin, Germany.
Department of Virology, Faculty of Veterinary Medicine, Cairo University, Giza, Egypt.
Cell Microbiol. 2021 Jun;23(6):e13322. doi: 10.1111/cmi.13322. Epub 2021 Mar 9.
Influenza A viruses contain two S-acylated proteins, the ion channel M2 and the glycoprotein hemagglutinin (HA). Acylation of the latter is essential for virus replication. Here we analysed the expression of each of the 23 members of the family of ZDHHC acyltransferases in human airway cells, the site of virus replication. RT-PCR revealed that every ZDHHC acyltransferase (except ZDHHC19) is expressed in A549 and Calu cells. Interestingly, expression of one ZDHHC, ZDHHC22, is upregulated in virus-infected cells; this effect is more pronounced after infection with an avian compared to a human virus strain. The viral protein NS1 triggers ZDHHC22 expression in transfected cells, whereas recombinant viruses lacking a functional NS1 gene did not cause ZDHHC22 upregulation. CRISPR/Cas9 technology was then used to knock-out the ZDHHC22 gene in A549 cells. However, acylation of M2 and HA was not reduced, as analysed for intracellular HA and M2 and the stoichiometry of S-acylation of HA incorporated into virus particles did not change according to MALDI-TOF mass spectrometry analysis. Comparative mass spectrometry of palmitoylated proteins in wt and ΔZDHHC22 cells identified 25 potential substrates of ZDHHC22 which might be involved in virus replication.
甲型流感病毒包含两种 S 酰化蛋白，即离子通道 M2 和糖蛋白血凝素（HA）。后者的酰化对于病毒复制是必不可少的。在这里，我们分析了家族中的 23 种 ZDHHC 酰基转移酶在人呼吸道细胞（病毒复制的部位）中的表达。RT-PCR 显示，除 ZDHHC19 外，每种 ZDHHC 酰基转移酶（ZDHHC19）在 A549 和 Calu 细胞中均有表达。有趣的是，一种 ZDHHC，即 ZDHHC22，在感染病毒的细胞中表达上调；与感染人类病毒株相比，感染禽流感病毒株时这种效应更为明显。病毒蛋白 NS1 可在转染细胞中触发 ZDHHC22 的表达，而缺乏功能性 NS1 基因的重组病毒则不会引起 ZDHHC22 的上调。然后，使用 CRISPR/Cas9 技术敲除 A549 细胞中的 ZDHHC22 基因。然而，如 MALDI-TOF 质谱分析所示，M2 和 HA 的酰化并未减少，细胞内 HA 和 M2 的分析以及整合到病毒颗粒中的 HA 的 S 酰化的化学计量也没有改变。wt 和 ΔZDHHC22 细胞中棕榈酰化蛋白的比较质谱鉴定出 25 种 ZDHHC22 的潜在底物，这些底物可能参与病毒复制。
