甲型流感病毒(IAV)对人类生命财产构成严重威胁。IAV基质蛋白2(M2)在病毒出芽中是重要的。越来越多的研究已经证明宿主因子在IAV复制中的重要作用。在这项研究中,免疫沉淀结合质谱显示宿主蛋白酪氨酸3-单加氧酶/色氨酸5-单加氧酶活化蛋白γ(YWHAG),属于14-3-3蛋白支架家族,与M2相互作用。通过共免疫沉淀(Co-IP)进一步证实了它们的相互作用,免疫荧光,和病毒感染的HeLa细胞的共聚焦显微镜。此外,我们构建了YWHAG-KO和YWHAG过表达细胞,发现YWHAG敲除显着增加病毒的产生,而其过表达降低了病毒后代的滴度。因此,YWHAG是IAV感染期间的负调节因子。Further,YWHAG敲除或过表达对结合没有影响,条目,或病毒的RNA复制在病毒生命周期的早期阶段。相反,如使用透射电子显微镜确定的,它损害了质膜上病毒体的释放,并抑制了M2介导的流感病毒的出芽。重要的是,发现YWHAG的H158F突变影响与M2的相互作用及其出芽。总的来说,我们的工作表明,YWHAG是一种新型的细胞调节因子,靶向并介导M2的相互作用和释放.
Influenza A virus (IAV) poses a serious threat to human life and property. The IAV matrix protein 2 (M2) is significant in viral budding. Increasing studies have proven the important roles of host factors in IAV replication. In this study, immunoprecipitation combined with mass spectrometry revealed that the host protein tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein gamma (
YWHAG), which belongs to the 14-3-3 protein scaffold family, interacts with M2. Their interactions were further confirmed by co-immunoprecipitation (Co-IP), immunofluorescence, and confocal microscopy of virus-infected HeLa cells. Moreover, we constructed
YWHAG-KO and
YWHAG-overexpressing cells and found that
YWHAG knockout significantly increased viral production, whereas its overexpression reduced the titer of virus progeny. Therefore,
YWHAG is a negative regulatory factor during IAV infection. Further,
YWHAG knockout or overexpression had no effect on the binding, entry, or viral RNA replication in the early stages of the virus life cycle. On the contrary, it impaired the release of virions at the plasma membrane as determined using transmission electron microscopy and suppressed the M2-mediated budding of the influenza virus. Importantly, the H158F mutation of
YWHAG was found to affect interaction with M2 and its budding. Collectively, our work demonstrates that YWHAG is a novel cellular regulator that targets and mediates the interaction and release of M2.