Role of viral and cellular recombination proteins in HSV DNA replication

病毒和细胞重组蛋白在 HSV DNA 复制中的作用

• 批准号: 8438424
• 负责人: SANDRA K WELLER
• 金额: $ 32.21万
• 依托单位: UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-01-15 至 2017-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8438424
• 关键词: Adult; Affect; Antiviral Therapy; Bacteriophage lambda; Biochemical; Biological Assay; Cancer Biology; Cell physiology; Cells; Complex; DNA; DNA Damage; DNA biosynthesis; Development; Double Strand Break Repair; Ensure; Environment; Excision; Eye Infections; G22P1 gene; Generations; Genetic Recombination; Genital system; Genome Stability; Goals; Growth; Head; Herpesvirus 1; Human; Human Herpesvirus 2; Immune system; Immunocompetent; In Vitro; Individual; Infection; Lead; Length; Life; Maintenance; Measures; Mediating; Morbidity - disease rate; Mutation; Nonhomologous DNA End Joining; Oral; Pathway interactions; Plasmids; Process; Production; Proteins; Recombinants; Recruitment Activity; Reporter; Resolution; Role; Signal Transduction; Simplexvirus; Structure; Surgical Flaps; Tail; Testing; Transfection; Viral; Viral Genes; Viral Genome; Viral Proteins; Virion; Virus; Virus Diseases; Virus Replication; Vision; base; improved; irradiation; mortality; mutant; novel; nuclease; pathogen; protein function; recombinase; recombinational repair; repaired; response; ubiquitin-protein ligase; viral DNA

DESCRIPTION (provided by applicant): Herpes Simplex Virus (HSV) is a major pathogen responsible for oral, genital and sight-threatening ocular infections which can be life threatening in immunocompetent adults and even more severe in individuals with compromised immune systems. Production of concatemeric DNA is an essential step for the generation of progeny virus as the packaging machinery must recognize longer-than-unit-length concatemers during encapsidation; however, the mechanism by which they are formed is very poorly understood. Although it has been proposed that the viral genome circularizes and rolling circle replication leads to the formation of concatemers, several lines of evidence suggest that HSV DNA replication is more complex and may involve recombination-dependent replication reminiscent of bacteriophages lambda and T4. We have previously shown that HSV encodes a two-subunit viral recombinase (UL12 and ICP8). Furthermore cellular DNA Damage Response (DDR) proteins have been shown by us and others to both positively and negatively influence the production of infectious progeny virus. The two subunit viral recombinase (UL12 and ICP8) interacts with several DDR proteins. It is now clear that DDR proteins function in at least four pathways, three that require some amount of homology (A-NHEJ, HR and SSA) and one that does not (C-NHEJ). We hypothesize that HSV navigates this complex environment to ensure the production of viral genomes that can be packaged into infectious virus. The central hypothesis is that HSV infection requires the activation of one or more of the homology dependent repair/recombination pathways and that viral proteins including the immediate early E3 ubiquitin ligase ICP0 and the viral recombinase (UL12 and ICP8) act to influence pathway choice. Furthermore we suggest that the C-NHEJ pathway is inactivated by viral proteins in order to promote end resection and the homology dependent pathways leading to the formation of concatemers that can be processed into infectious virus. In aim 1, we will determine which cellular pathways are activated and inactivated by HSV infection and the consequences of these actions. In Aim 2 we will test the hypothesis that UL12 affects DDR signaling and directs pathway choice. In aim 3 we will test the hypothesis that the structure of viral DNA depends on the repair pathway activated during infection and that in the absence of UL12, inappropriate pathway choice leads to the accumulation of structurally aberrant DNA. It is expected that this study will improve our understanding of the mechanism of viral DNA replication, provide information on the functions of cellular proteins important for genome stability and cancer biology and aid in the development of new antiviral therapies.

描述（由申请人提供）：单纯疱疹病毒（HSV）是负责口服，生殖器和视力危及眼感染的主要病原体，在免疫能力的成年人中可能会威胁生命，而在患有受损的免疫系统的患者中可能会威胁生命。产生串联DNA是产生后代病毒的重要步骤，因为包装机制必须在封装过程中识别超过单位长度的conpatemer。但是，它们形成的机制非常了解。尽管已经提出，病毒基因组循环和滚动圆的复制导致了辅导者的形成，但几条证据表明，HSV DNA复制更为复杂，可能涉及重组依赖性依赖性复制，让人联想到噬菌体lambda和t4。我们先前已经表明，HSV编码了两亚基病毒重组酶（UL12和ICP8）。此外，我们和其他人已经证明了细胞DNA损伤反应（DDR）蛋白，以积极和负面影响传染性后代病毒的产生。两个亚基病毒重组酶（UL12和ICP8）与多种DDR蛋白相互作用。现在很明显，DDR蛋白在至少四个途径中起作用，三种需要一定量的同源性（a-nhej，hr和ssa），而一种不具有（c-nhej）。我们假设HSV会导航这个复杂的环境，以确保可以包装成传染病的病毒基因组的产生。中心假设是HSV感染需要激活一个或多个同源性依赖性修复/重组途径，以及包括立即E3早期E3泛素连接酶ICP0和病毒重组酶（UL12和ICP8）的病毒蛋白，以影响途径。此外，我们建议C-NHEJ途径被病毒蛋白灭活，以促进最终切除术和同源性依赖性途径，导致形成了可以加工成感染性病毒的辅助者的形成。在AIM 1中，我们将确定哪些细胞途径被HSV感染激活并灭活以及这些作用的后果。在AIM 2中，我们将测试UL12影响DDR信号并指导途径选择的假设。在AIM 3中，我们将检验以下假设：病毒DNA的结构取决于感染过程中激活的修复途径，并且在没有UL12的情况下，不适当的途径选择会导致结构异常DNA的积累。预计这项研究将提高我们对病毒DNA复制机制的理解，提供有关细胞蛋白对基因组稳定性和癌症生物学重要的功能的信息，并有助于开发新的抗病毒疗法。