The Role of VP22 in Protein Synthesis and mRNA Accumulation During HSV-1 Infectio

VP22 在 HSV-1 感染过程中蛋白质合成和 mRNA 积累中的作用

• 批准号: 7712379
• 负责人: Carol Lynn Duffy
• 金额: $ 17.88万
• 依托单位: UNIVERSITY OF ALABAMA IN TUSCALOOSA
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-07-17 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7712379
• 关键词: Affect; Attenuated; Biological Assay; Blindness; Cell Culture Techniques; Cells; Complex; Cornea; Dactinomycin; Data; Defect; Development; Disease; Encephalitis; Foundations; Future; Genes; Genetic Transcription; Genital system; Goals; Herpes Labialis; Herpesvirus 1; Human; In Vitro; Individual; Infection; Lesion; Life; Mass Spectrum Analysis; Messenger RNA; Molecular; Mus; Mutation; Nuclear; Pathogenesis; Phenotype; Play; Production; Protein Biosynthesis; Proteins; Radiolabeled; Recurrence; Regulation; Research; Role; Running; Satellite Viruses; Simplexvirus; Solid; Techniques; Testing; Therapeutic Uses; Time; Translations; Two-Dimensional Gel Electrophoresis; VP 16; Viral; Viral Proteins; Viral Regulatory Proteins; Virus; Virus Diseases; Virus Replication; base; extracellular; genetic regulatory protein; improved; mRNA Export; mRNA Transcript Degradation; monolayer; novel; pathogen; protein function; public health relevance; radiotracer; repaired; translation assay

DESCRIPTION (provided by applicant): Herpes simplex virus type 1 (HSV-1) is a widespread human pathogen that causes a diverse array of diseases ranging from recurrent cold sores and genital lesions to blindness and encephalitis. The pathogenesis of HSV-1 depends both on the virus's ability to replicate and its ability to spread. The HSV-1 VP22 protein, encoded by the UL49 gene, has been shown to play an important role in both virus production and spread. Our studies show a UL49-null virus possesses an intriguing and unusual phenotype -- that of normal protein synthesis at early times in infection followed by a synchronous and nearly-global shutdown in protein synthesis at late times in infection. The defect is in protein synthesis and not protein stability. This same virus possesses a defect in the accumulation of a subset of viral mRNAs at early times in infection. The defects in protein synthesis and mRNA accumulation are distinct and separable with regards to both timing during infection and the genes affected. As these findings are novel, nothing is known regarding the mechanisms by which VP22 regulates protein synthesis at late times and mRNA accumulation at early times in infection. The data obtained in the proposed studies will identify those mechanisms and serve as a solid foundation for a future R01 application in which we will molecularly dissect VP22's roles in protein synthesis and mRNA accumulation. Our first aim is to identify the mechanism(s) responsible for the protein synthesis shutdown observed in UL49-null virus infected cells at late times in infection. We will determine whether decreased mRNA export, decreased translation, or both are responsible for the decreased levels of protein synthesis observed late in UL49-null virus infections. We will also identify proteins that undergo decreased synthesis late in UL49-null virus infections. Finally, we will determine whether VP16 and vhs are involved in VP22-dependent protein synthesis. VP22, VP16, and vhs form a complex and secondary mutations that alleviate the UL49-null virus-associated spread defect frequently occur in vhs. Thus, we hypothesize VP22 acts in concert with VP16 and/or vhs to regulate protein synthesis and will test this hypothesis through targeted mutational analyses. Our second aim is to identify the mechanism(s) responsible for the decreased steady-state mRNA levels observed at early times in UL49-null virus infections. We will determine whether increased mRNA degradation, decreased transcription, or both are responsible for the decreased steady-state mRNA levels observed early in UL49-null virus infections. We will perform HSV-1 microarray studies to identify mRNAs present in decreased levels early in UL49-null virus infections. Finally, we hypothesize that VP22 acts to increase mRNA accumulation early in infection by modulating the activity of vhs, either independently or in concert with VP16. We will test this hypothesis through targeted mutational analyses. PUBLIC HEALTH RELEVANCE: Herpes simplex virus type 1 (HSV-1) is a widespread human pathogen that causes a diverse array of diseases ranging from recurrent cold sores and genital lesions to blindness and encephalitis. The pathogenesis of HSV-1 depends both on the virus's ability to replicate itself and its ability to spread. The studies proposed herein will enable us to elucidate mechanisms utilized by HSV-1 for virus replication and spread as a prerequisite to the development of improved therapeutics for use in attenuating the above diseases.

描述(申请人提供)：单纯疱疹病毒1型(HSV-1)是一种广泛传播的人类病原体，可导致从复发性唇疱疹和生殖器损伤到失明和脑炎等多种疾病。HSV-1的发病机制既取决于病毒的复制能力，也取决于病毒的传播能力。HSV-1 VP22蛋白由UL49基因编码，已被证明在病毒生产和传播中发挥重要作用。我们的研究表明，UL49缺失病毒具有一种耐人寻味的不同寻常的表型--感染早期蛋白质合成正常，随后感染后期蛋白质合成同步且几乎全局性停止。缺陷在于蛋白质合成，而不是蛋白质稳定性。同样的病毒在感染早期的病毒mRNAs子集的积累方面存在缺陷。蛋白质合成和mRNA积累的缺陷在感染的时间和受影响的基因方面都是明显和可分离的。由于这些发现是新的，关于VP22在感染后期调节蛋白质合成和在感染早期调节信使核糖核酸积累的机制尚不清楚。这些研究中获得的数据将确定这些机制，并为未来R01的应用奠定坚实的基础，在这一应用中，我们将从分子上剖析VP22的S在蛋白质合成和基因积累中的作用。我们的第一个目标是确定导致UL49缺失病毒感染细胞在感染后期观察到的蛋白质合成停止的机制(S)。我们将确定在UL49缺失病毒感染后期观察到的蛋白质合成水平降低是否与信使核糖核酸输出减少、翻译减少或两者共同作用有关。我们还将确定在UL49缺失病毒感染后期经历合成减少的蛋白质。最后，我们将确定VP16和VHS是否参与VP22依赖的蛋白质合成。VP22、VP16和VHS形成复杂的继发性突变，缓解VHS中频繁发生的UL49缺失病毒相关传播缺陷。因此，我们假设VP22与VP16和/或VHS协同作用来调节蛋白质合成，并将通过有针对性的突变分析来检验这一假设。我们的第二个目标是确定在UL49缺失病毒感染早期观察到的稳态水平下降的机制(S)。我们将确定在UL49缺失病毒感染早期观察到的稳态mRNA水平降低是否与mRNA降解增加、转录减少或两者兼而有之有关。我们将进行HSV-1微阵列研究，以确定在UL49缺失病毒感染早期存在水平降低的mRNAs。最后，我们假设VP22通过独立或与VP16共同调节VHS的活性来增加感染早期的mRNA积累。我们将通过有针对性的突变分析来检验这一假设。公共卫生相关性：1型单纯疱疹病毒(HSV-1)是一种广泛传播的人类病原体，可导致从复发性唇疱疹和生殖器损伤到失明和脑炎等各种疾病。HSV-1的发病机制既取决于病毒自我复制的能力，也取决于病毒的传播能力。本文提出的研究将使我们能够阐明HSV-1用于病毒复制和传播的机制，作为开发用于减轻上述疾病的改进疗法的先决条件。