MX2 viral substrate breadth and inhibitory activity are regulated by protein phosphorylation
MX2 viral substrate breadth and inhibitory activity are regulated by protein phosphorylation
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MX2 病毒底物广度和抑制活性受蛋白质磷酸化调节
DOI：
10.1101/2022.01.24.477574
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发表时间：
2022
期刊：
影响因子：
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通讯作者：
Betancor G
中科院分区：
文献类型：
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作者：
Betancor G
人类黏液病毒抗性2 （MX2/MxB）能有效抑制人类免疫缺陷病毒1型（HIV-1）感染，这种病毒能与病毒衣壳结合并阻断病毒DNA的核输入。我们最近发现磷酸化是MX2抗病毒活性的关键调节因子，位点14、17和18的丝氨酸残基磷酸化抑制MX2的功能。在这里，我们扩展了MX2翻译后修饰的研究，并确定了MX2所有结构域的丝氨酸和苏氨酸磷酸化。通过用天冬氨酸或丙氨酸取代这些残基，从而分别模拟磷酸基团的存在或缺失，我们确定了控制MX2抗病毒活性的关键位置。天冬氨酸取代Ser306或Thr334残基和丙氨酸取代Thr343残基产生的蛋白质抗病毒活性大大降低，而在Ser28、Thr151或Thr343位置存在天冬氨酸导致活性增强：被称为高变形突变体。在某些情况下，这些高形态突变，特别是与其他MX2突变（如S28D/T151D或T151D/T343A）配对时，获得了抑制已知对野生型MX2不敏感的HIV-1衣壳突变的能力，如P90A或T210K，以及MX2抗性逆转录病毒，如马传染性贫血病毒（EIAV）和小鼠白血病病毒（MLV）。这项工作强调了MX2磷酸化在抗病毒活性调控和易感病毒底物选择中的复杂性和重要性。人类免疫缺陷病毒1型（HIV-1）的生产性感染需要将病毒复制复合体输入到感染细胞的细胞核中。黏液病毒抗性2 （MX2/MxB）阻断这一步骤，阻止病毒DNA的核积累和病毒复制。我们最近证明了MX2的氨基末端区域的三段丝氨酸的磷酸化如何抑制抗病毒活性。在这里，我们确定了MX2中磷酸化状态降低或增强抗病毒功能的其他位置（分别为半态和超态变体）。重要的是，超形态突变蛋白不仅增加了对野生型HIV-1的抑制活性，而且还能表现出对抗野生型MX2的HIV-1衣壳突变病毒的抗病毒能力。此外，其中一些蛋白还能抑制对MX2不敏感的逆转录病毒。因此，我们认为磷酸化是MX2调控和底物决定的主要因素。
Human immunodeficiency virus type-1 (HIV-1) infection is potently inhibited by human myxovirus resistance 2 (MX2/MxB), which binds to the viral capsid and blocks the nuclear import of viral DNA. We have recently shown that phosphorylation is a key regulator of MX2 antiviral activity, with phosphorylation of serine residues at positions 14, 17, and 18 repressing MX2 function. Here, we extend the study of MX2 posttranslational modifications and identify serine and threonine phosphorylation in all domains of MX2. By substituting these residues with aspartic acid or alanine, hence mimicking the presence or absence of a phosphate group, respectively, we identified key positions that control MX2 antiviral activity. Aspartic acid substitutions of residues Ser306 or Thr334 and alanine substitutions of Thr343 yielded proteins with substantially reduced antiviral activity, whereas the presence of aspartic acid at positions Ser28, Thr151, or Thr343 resulted in enhanced activity: referred to as hypermorphic mutants. In some cases, these hypermorphic mutations, particularly when paired with other MX2 mutations (e.g., S28D/T151D or T151D/T343A) acquired the capacity to inhibit HIV-1 capsid mutants known to be insensitive to wild-type MX2, such as P90A or T210K, as well as MX2-resistant retroviruses such as equine infectious anemia virus (EIAV) and murine leukemia virus (MLV). This work highlights the complexity and importance of MX2 phosphorylation in the regulation of antiviral activity and in the selection of susceptible viral substrates.IMPORTANCEProductive infection by human immunodeficiency virus type-1 (HIV-1) requires the import of viral replication complexes into the nuclei of infected cells. Myxovirus resistance 2 (MX2/MxB) blocks this step, halting nuclear accumulation of viral DNA and virus replication. We recently demonstrated how phosphorylation of a stretch of three serines in the amino-terminal domain of MX2 inhibits the antiviral activity. Here, we identify additional positions in MX2 whose phosphorylation status reduces or enhances antiviral function (hypomorphic and hypermorphic variants, respectively). Importantly, hypermorphic mutant proteins not only increased inhibitory activity against wild-type HIV-1 but can also exhibit antiviral capabilities against HIV-1 capsid mutant viruses that are resistant to wild-type MX2. Furthermore, some of these proteins were also able to inhibit retroviruses that are insensitive to MX2. Therefore, we propose that phosphorylation comprises a major element of MX2 regulation and substrate determination.