Chromatin and Coactivators in HSV-1 gene regulation

描述（由申请人提供）：真核生物基因组包装在染色质中，染色质是一种含有DNA、组蛋白和非组蛋白染色体蛋白的核蛋白复合物。因此提供的压实为需要访问模板DNA的复制和转录等细胞过程提供了重要的障碍。感染真核细胞的DNA病毒为研究染色质对基因调控和复制的影响提供了有益的实验对象。HSV-1的VP16蛋白能有效地刺激病毒即时早期基因的转录，从而在裂解感染周期中触发病毒基因的级联表达。在异源实验背景下，VP16可以招募染色质修饰共激活子到目标启动子。然而，先前的报道表明，在溶性感染期间，大的HSV-1病毒DNA基因组主要是非核小体的。这些观察结果揭示了一个明显的悖论：为什么像VP16这样的病毒激活因子会将染色质修饰辅助激活因子蛋白招募到缺乏核小体的DNA模板上？本项目的总体目标是确定组蛋白和染色质在HSV-1基因调控中的作用，包括感染期间病毒基因上染色质修饰共激活蛋白的募集和活性。迄今为止，我们的工作已经表明，在HSV感染期间，某些共激活因子确实以依赖于VP16激活域的方式与病毒DNA相关联。此外，组蛋白存在于病毒DNA的许多区域，但以vp16依赖的方式在IE启动子中缺失。在拟议的项目期间，将测试其他组蛋白和非组蛋白染色体蛋白在溶性感染过程中与HSV DNA的关联。我们将研究IE启动子上是否存在其他辅助激活蛋白，以及IE转录对这些辅助激活因子的依赖性。这项工作将促进对细胞调控机制的理解，以及利用HSV使这些机制适应病毒基因表达的策略。

DESCRIPTION (provided by applicant): Eukaryotic genomes are packaged in chromatin, a nucleoprotein complex containing DNA, histones, and nonhistone chromosomal proteins. The compaction thus provided presents a significant barrier to cellular processes such as replication and transcription that require access to template DNA. DNA viruses that infect eukaryotic cells have served as useful experimental subjects for studying effects of chromatin on gene regulation and replication. The VP16 protein of HSV-1 potently stimulates transcription of the viral immediate early genes and thus triggers the cascade of viral gene expression during the lytic infection cycle. In heterologous experimental contexts, VP16 can recruit chromatin modifying coactivators to target promoters. However, previous reports indicated that the large HSV-1 viral DNA genome remains predominantly non-nucleosomal during lytic infection. These observations reveal an apparent paradox: why would a viral activator such as VP16 recruit chromatin modifying coactivator proteins to a DNA template that lacks nucleosomes? The overall goal of this project is to define the roles of histones and chromatin in HSV-1 gene regulation, including the recruitment and activity of chromatin-modifying coactivator proteins on viral genes during infection. Our work to date has shown that certain coactivators do associate with viral DNA during HSV infection in a manner dependent on the VP16 activation domain. Moreover, histones are present at many regions of the viral DNA, but are absent from IE promoters in a VP16-dependent manner. In the proposed project period, the association of additional histones and nonhistone chromosomal proteins with HSV DNA throughout lytic infection will be tested. The presence of other coactivator proteins at IE promoters will be examined, as will the dependence of IE transcription on those coactivators. This work will advance the understanding of cellular regulatory mechanisms and the strategies used by HSV to adapt those mechanisms for viral -gene expression. Herpes simplex virus infections are widespread in the US population, and cause significant discomfort and occasional death. Early stages in virus infection seem to depend on certain enzymes from the host cell. This research will validate the roles of those enzymes in virus infection, which may lead to new antiviral drug targets.