PDF(Pubmed)
Abstract：
Influenza A virus (IAV) can cause severe and life-threatening illness in humans and animals. Therefore, it is important to search for host antiviral proteins and elucidate their antiviral mechanisms for the development of potential treatments. As a part of human innate immunity, host restriction factors can inhibit the replication of viruses, among which SAM and HD domain containing deoxynucleoside triphosphate triphosphohydrolase 1 (SAMHD1) can restrict the replication of viruses, such as HIV and enterovirus EV71. Viruses also developed countermeasures in the arms race with their hosts. There are few reports about whether SAMHD1 has a restriction effect on IAV.
To investigate the impact of IAV infection on SAMHD1 expression in A549 cells, we infected A549 cells with a varying multiplicity of infection (MOI) of IAV and collected cell samples at different time points for WB and RT-qPCR analysis to detect viral protein and SAMHD1 levels. The virus replication level in the cell culture supernatant was determined using TCID50 assay. Luciferase assay was used to reveal that H5N1 virus polymerase acidic protein (PA) affected the activity of the SAMHD1 promoter. To assess the antiviral capacity of SAMHD1, we generated a knockdown and overexpressed cell line for detecting H5N1 replication.
In this study, we observed that SAMHD1 can restrict the intracellular replication of H5N1 and that the H5N1 viral protein PA can downregulate the expression of SAMHD1 by affecting SAMHD1 transcriptional promoter activity. We also found that SAMHD1\'s ability to restrict H5N1 is related to phosphorylation at 592-tyrosine.
In conclusion, we found that SAMHD1 may affect the replication of IAVs as a host restriction factor and be countered by PA. Furthermore, SAMHD1 may be a potential target for developing antiviral drugs.
To investigate the impact of IAV infection on SAMHD1 expression in A549 cells, we infected A549 cells with a varying multiplicity of infection (MOI) of IAV and collected cell samples at different time points for WB and RT-qPCR analysis to detect viral protein and SAMHD1 levels. The virus replication level in the cell culture supernatant was determined using TCID50 assay. Luciferase assay was used to reveal that H5N1 virus polymerase acidic protein (PA) affected the activity of the SAMHD1 promoter. To assess the antiviral capacity of SAMHD1, we generated a knockdown and overexpressed cell line for detecting H5N1 replication.
In this study, we observed that SAMHD1 can restrict the intracellular replication of H5N1 and that the H5N1 viral protein PA can downregulate the expression of SAMHD1 by affecting SAMHD1 transcriptional promoter activity. We also found that SAMHD1\'s ability to restrict H5N1 is related to phosphorylation at 592-tyrosine.
In conclusion, we found that SAMHD1 may affect the replication of IAVs as a host restriction factor and be countered by PA. Furthermore, SAMHD1 may be a potential target for developing antiviral drugs.
摘要：
背景：甲型流感病毒（IAV）可在人类和动物中引起严重且危及生命的疾病。因此,寻找宿主抗病毒蛋白并阐明其抗病毒机制对于开发潜在的治疗方法非常重要。作为人类先天免疫的一部分，宿主限制因子可以抑制病毒的复制，其中含有SAM和HD结构域的脱氧核苷三磷酸三磷酸水解酶1（SAMHD1）可以限制病毒的复制，如HIV和肠道病毒EV71。病毒还在与其宿主的军备竞赛中发展了对策。关于SAMHD1是否对IAV有限制作用的报道很少。
方法：为了研究IAV感染对A549细胞SAMHD1表达的影响，我们用不同的IAV感染复数(MOI)感染A549细胞,并在不同时间点收集细胞样品进行WB和RT-qPCR分析,以检测病毒蛋白和SAMHD1水平.使用TCID50测定法测定细胞培养上清液中的病毒复制水平。荧光素酶测定用于揭示H5N1病毒聚合酶酸性蛋白（PA）影响SAMHD1启动子的活性。为了评估SAMHD1的抗病毒能力,我们产生了用于检测H5N1复制的敲低和过表达的细胞系。
结果：在这项研究中，我们观察到SAMHD1可以限制H5N1的细胞内复制，并且H5N1病毒蛋白PA可以通过影响SAMHD1转录启动子活性来下调SAMHD1的表达。我们还发现SAMHD1限制H5N1的能力与592-酪氨酸的磷酸化有关。
结论：结论：我们发现SAMHD1可能作为宿主限制因子影响IAV的复制，并被PA抵消。此外，SAMHD1可能是开发抗病毒药物的潜在靶标。
方法：为了研究IAV感染对A549细胞SAMHD1表达的影响，我们用不同的IAV感染复数(MOI)感染A549细胞,并在不同时间点收集细胞样品进行WB和RT-qPCR分析,以检测病毒蛋白和SAMHD1水平.使用TCID50测定法测定细胞培养上清液中的病毒复制水平。荧光素酶测定用于揭示H5N1病毒聚合酶酸性蛋白（PA）影响SAMHD1启动子的活性。为了评估SAMHD1的抗病毒能力,我们产生了用于检测H5N1复制的敲低和过表达的细胞系。
结果：在这项研究中，我们观察到SAMHD1可以限制H5N1的细胞内复制，并且H5N1病毒蛋白PA可以通过影响SAMHD1转录启动子活性来下调SAMHD1的表达。我们还发现SAMHD1限制H5N1的能力与592-酪氨酸的磷酸化有关。
结论：结论：我们发现SAMHD1可能作为宿主限制因子影响IAV的复制，并被PA抵消。此外，SAMHD1可能是开发抗病毒药物的潜在靶标。
