Influenza virus hemagglutinin (H3 subtype) requires palmitoylation of its cytoplasmic tail for assembly: M1 proteins of two subtypes differ in their ability to support assembly
Influenza virus hemagglutinin (H3 subtype) requires palmitoylation of its cytoplasmic tail for assembly: M1 proteins of two subtypes differ in their ability to support assembly
复制标题
DOI：
10.1128/jvi.79.21.13673-13684.2005
复制
发表时间：
2005-11-01
影响因子：
5.4
通讯作者：
Lamb, RA
中科院分区：
文献类型：
--
作者：
Chen, BJ;Takeda, M;Lamb, RA
甲型流感病毒血凝素（HA）跨膜结构域边界区和细胞质尾部含有3个半胱氨酸（H3 HA亚型的残基555、562和565），它们在16个HA亚型中高度保守，每一个都被棕榈酸的共价添加修饰。先前对这些保守半胱氨酸残基作用的分析得出了不同的数据，表明HA棕榈酰化要么没有作用，要么有重要作用。为了重新研究这些残基在流感病毒生命周期中的作用，利用高效的反向遗传系统将一系列半胱氨酸-丝氨酸突变引入流感病毒a /Udorn/72(Ud) （H3N2）的HA基因。含有HA- c562s和HA- c565s突变的突变病毒在MDCK细胞中生长减少，不能形成斑块，但在表达野生型HA的MDCK细胞系中形成野生型样斑块。在细胞-细胞融合实验中，非棕榈酰化的H3 HA在dna转染和病毒感染的细胞中都能完全胜任HA介导的膜融合。当检测HA细胞质尾部半胱氨酸突变体的脂质筏关联时，使用Triton X-100不溶性作为标准，筏关联的丧失与病毒复制的减少没有直接关系。然而，突变病毒的组装随着病毒复制的减少而减少。此外，菌株a /WSN/33 （WSN）的重组体含有突变C555S、C562S和C565S的Ud HA基因，产生的病毒可以在常规MDCK细胞上形成斑块，并且复制仅适度减少，这表明Ud和WSN HA与内部病毒蛋白之间的相互作用存在差异。对M1突变体中含有6个不同于Ud和WSN M1蛋白的残基替换的分析表明，在M1蛋白支持病毒与非棕榈酰化H3 HA组装的能力方面，一系列残基导致了M1蛋白之间的差异。
The influenza A virus hemagglutinin (HA) transmembrane domain boundary region and the cytoplasmic tail contain three cysteines (residues 555, 562, and 565 for the H3 HA subtype) that are highly conserved among the 16 HA subtypes and which are each modified by the covalent addition of palmitic acid. Previous analysis of the role of these conserved cysteine residues led to differing data, suggesting either no role for HA palmitoylation or an important role for HA palmitoylation. To reexamine the role of these residues in the influenza virus life cycle, a series of cysteine-to-serine mutations were introduced into the HA gene of influenza virus A/Udorn/72(Ud) (H3N2) by using a highly efficient reverse genetics system. Mutant viruses containing HA-C562S and HA-C565S mutations had reduced growth and failed to form plaques in MDCK cells but formed wild-type-like plaques in an MDCK cell line expressing wild-type HA. In cell-cell fusion assays, nonpalmitoylated H3 HA, in both cDNA-transfected and virus-infected cells, was fully competent for HA-mediated membrane fusion. When the HA cytoplasmic tail cysteine mutants were examined for lipid raft association, using as the criterion Triton X-100 insolubility, loss of raft association did not show a direct correlation with a reduction in virus replication. However, mutant virus assembly was reduced in parallel with reduced virus replication. Additionally, a reassortant of strain A/WSN/33 (WSN), containing the Ud HA gene with mutations C555S, C562S, and C565S, produced virus that could form plaques on regular MDCK cells and had only moderately decreased replication, suggesting differences in the interactions between Ud and WSN HA and internal viral proteins. Analysis of M1 mutants containing substitutions in the six residues that differ between the Ud and WSN M1 proteins indicated that a constellation of residues are responsible for the difference between the M1 proteins in their ability to support virus assembly with nonpalmitoylated H3 HA.