School of Veterinary Medicine and Biomedical Sciences, University of Nebraska, Lincoln, Nebraska, United States of America; Nebraska Center for Virology, University of Nebraska, Lincoln, Nebraska, United States of America.
PLoS One. 2014 Jun 19;9(6):e100511. doi: 10.1371/journal.pone.0100511. eCollection 2014.
BAF (Barrier to Autointegration Factor) is a highly conserved DNA binding protein that senses poxviral DNA in the cytoplasm and tightly binds to the viral genome to interfere with DNA replication and transcription. To counteract BAF, a poxviral-encoded protein kinase phosphorylates BAF, which renders BAF unable to bind DNA and allows efficient viral replication to occur. Herein, we examined how BAF phosphorylation is affected by herpes simplex virus type 1 (HSV-1) infection and tested the ability of BAF to interfere with HSV-1 productive infection. Interestingly, we found that BAF phosphorylation decreases markedly following HSV-1 infection. To determine whether dephosphorylated BAF impacts HSV-1 productive infection, we employed cell lines stably expressing a constitutively unphosphorylated form of BAF (BAF-MAAAQ) and cells overexpressing wild type (wt) BAF for comparison. Although HSV-1 production in cells overexpressing wtBAF was similar to that in cells expressing no additional BAF, viral growth was reduced approximately 80% in the presence of BAF-MAAAQ. Experiments were also performed to determine the mechanism of the antiviral activity of BAF with the following results. BAF-MAAAQ was localized to the nucleus, whereas wtBAF was dispersed throughout cells prior to infection. Following infection, wtBAF becomes dephosphorylated and relocalized to the nucleus. Additionally, BAF was associated with the HSV-1 genome during infection, with BAF-MAAAQ associated to a greater extent than wtBAF. Importantly, unphosphorylated BAF inhibited both viral DNA replication and gene expression. For example, expression of two regulatory proteins, ICP0 and VP16, were substantially reduced in cells expressing BAF-MAAAQ. However, other viral genes were not dramatically affected suggesting that expression of certain viral genes can be differentially regulated by unphosphorylated BAF. Collectively, these results suggest that BAF can act in a phosphorylation-regulated manner to impair HSV-1 transcription and/or DNA replication, which is similar to the antiviral activity of BAF during vaccinia infection.
BAF（自动整合因子的屏障）是一种高度保守的 DNA 结合蛋白，它在细胞质中感知痘病毒 DNA，并紧密结合病毒基因组，干扰 DNA 复制和转录。为了对抗 BAF，痘病毒编码的蛋白激酶使 BAF 磷酸化，使 BAF 无法结合 DNA，并允许有效的病毒复制发生。在此，我们研究了 BAF 磷酸化如何受到单纯疱疹病毒 1（HSV-1）感染的影响，并测试了 BAF 干扰 HSV-1 有效感染的能力。有趣的是，我们发现 HSV-1 感染后 BAF 磷酸化明显降低。为了确定去磷酸化的 BAF 是否影响 HSV-1 的有效感染，我们使用稳定表达组成型非磷酸化形式 BAF（BAF-MAAAQ）的细胞系和过表达野生型（wt）BAF 的细胞进行比较。尽管过表达 wtBAF 的细胞中 HSV-1 的产生与未表达额外 BAF 的细胞相似，但在 BAF-MAAAQ 的存在下，病毒生长减少了约 80%。还进行了实验以确定 BAF 的抗病毒活性的机制，结果如下。BAF-MAAAQ 定位于细胞核，而 wtBAF 在感染前分布在细胞中。感染后，wtBAF 去磷酸化并重新定位到细胞核。此外，BAF 在感染过程中与 HSV-1 基因组结合，BAF-MAAAQ 的结合程度高于 wtBAF。重要的是，未磷酸化的 BAF 抑制病毒 DNA 复制和基因表达。例如，在表达 BAF-MAAAQ 的细胞中，两个调节蛋白 ICP0 和 VP16 的表达大大降低。然而，其他病毒基因没有受到明显影响，这表明某些病毒基因的表达可以被未磷酸化的 BAF 差异调节。总的来说，这些结果表明 BAF 可以以磷酸化调节的方式作用，损害 HSV-1 的转录和/或 DNA 复制，这与 BAF 在牛痘感染中的抗病毒活性相似。
