The Role of PIAS1 in EBV Biology

项目总结 爱泼斯坦-巴尔病毒(EBV)感染了95%的人类人口，并导致了1-2%的人类癌症。至 到目前为止，还没有专门的疗法来根除与这种病毒相关的癌症。成功者 开发EBV特异性抗癌策略需要对分子的深入了解 控制病毒潜伏和重新激活的机制。目前，我们在这方面存在着根本性的差距 了解细胞因素如何控制EBV潜伏期和重新激活。这项研究的中心目标是 该计划旨在阐明活化STAT1蛋白抑制因子(PIAS1)在EBV生命周期中的作用。 我们最近的研究表明，PIAS1是一种EBV限制因子，可以从潜伏期抑制病毒的重新激活。我们 已经证明PIAS1通过caspase被切割，并在裂解过程中被EBV蛋白激酶磷酸化 这是EBV复制所必需的。控制裂解性EBV复制的机制是 与PIAS1抑制病毒转录活性有关的关键细胞转录因子 用于裂解基因表达。此外，PIAS1还调节一组宿主基因的表达 胆固醇生物合成过程，EBV裂解复制的重要途径。这些结果导致了我们的中央 假设PIAS1通过抑制病毒和细胞转录因子来限制EBV裂解复制，并且 PIAS1被caspase依赖的切割和病毒激酶介导的磷酸化所拮抗，以促进裂解 复制。 对于这一假设，我们建议追求两个具体目标： 目的1：探讨PIAS1抑制EB病毒裂解复制的机制。我们将确定 并验证感染细胞中EBV和宿主基因组上的PIAS1结合位点。我们将确定PIAS1如何 通过抑制病毒和细胞转录因子来调节EBV裂解基因的表达。我们还将确定 PIAS1如何通过调节胆固醇生物合成途径部分抑制EBV的重新激活。 目的2：阐明PIAS1介导的抑制被拮抗为促进的机制 EBV裂解复制。PIAS1被caspase-3、-6和-8切割，并被EBV蛋白激酶磷酸化 在裂解复制过程中。我们将使用蛋白质组学和分子方法鉴定PIAS1的磷酸化位点。 我们将阐明促进EBV复制的磷酸化和裂解PIAS1之间的相互作用。 我们提出的研究将显著提高我们对PIAS1作为限制因素是如何 调节EBV感染，以及PIAS1在重新激活过程中如何受到病毒和细胞因子的调节。这些 研究将有可能阐明由一个中枢控制的EBV裂解再激活的分子基础 主机因素。

PROJECT SUMMARY Epstein-Barr virus (EBV) infects 95% of the human population and contributes to 1-2% of all human cancers. To date, there are no specific therapies to eradicate cancers associated with this virus. The successful development of EBV-specific anti-cancer strategies requires an in-depth understanding of the molecular mechanism controlling viral latency and reactivation. Currently, there are fundamental gaps in our understanding of how cellular factors control EBV latency and reactivation. The central goal of this research program is to elucidate the role of protein inhibitor of activated STAT1 (PIAS1) in the EBV life cycle. Our recent study revealed that PIAS1 is an EBV restriction factor that inhibits virus reactivation from latency. We have demonstrated that PIAS1 is cleaved via caspases and phosphorylated by an EBV protein kinase upon lytic induction and that this is required for EBV replication. The mechanism for controlling lytic EBV replication is related to the ability of PIAS1 to inhibit the transcriptional activity of viral and cellular transcription factors critical for lytic gene expression. In addition, PIAS1 also regulates the expression of a group host genes involved in the cholesterol biosynthesis process, a pathway important for EBV lytic replication. These results lead to our central hypothesis that PIAS1 restricts EBV lytic replication by inhibiting viral and cellular transcription factors, and that PIAS1 is antagonized by caspase-dependent cleavage and viral kinase-mediated phosphorylation to foster lytic replication. Towards this hypothesis, we propose to pursue two Specific Aims: AIM 1: To determine the mechanisms by which PIAS1 suppresses EBV lytic replication. We will identify and validate PIAS1 binding sites on the EBV and host genomes in infected cells. We will determine how PIAS1 regulates EBV lytic gene expression via inhibiting viral and cellular transcription factors. We will also determine how PIAS1 inhibits EBV reactivation partially through modulating the cholesterol biosynthesis pathway. Aim 2: To elucidate the mechanisms by which PIAS1-mediated suppression is antagonized to facilitate EBV lytic replication. PIAS1 is cleaved by caspase-3, -6 and -8, and phosphorylated by an EBV protein kinase during lytic replication. We will identify PIAS1 phosphorylation sites using proteomic and molecular approaches. We will elucidate the interplay between the phosphorylation and cleavage PIAS1 that facilitates EBV replication. Our proposed research will significantly improve our understanding of how PIAS1, as a restriction factor, regulates EBV infection and how PIAS1 is regulated by viral and cellular factors during reactivation. These studies will have the potential to illuminate the molecular basis of EBV lytic reactivation controlled by one central host factor.