Mechanisms of Early Growth Response Factor 1 (Egr-1) Induction by HSV-1 Lytic Inf

HSV-1 Lytic Inf 诱导早期生长反应因子 1 (Egr-1) 的机制

• 批准号: 7939557
• 负责人: Shaochung Victor Hsia
• 金额: $ 0.38万
• 依托单位: UNIVERSITY OF LOUISIANA AT MONROE
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-05-01 至 2010-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7939557
• 关键词: Address; Affect; Apoptosis; Be++ element; Beryllium; Binding; Bioinformatics; Biological Assay; Biological Process; Blindness; Catalytic DNA; Cell Line; Cell Proliferation; Cells; Cicatrix; Cornea; Cyclic AMP-Responsive DNA-Binding Protein; DNA purification; Data; Disease; Disease Progression; Elements; Encephalitis; Eye; Gene Expression; Genetic Transcription; Goals; Growth; Herpes Labialis; Herpesvirus 1; Hour; Immune response; Infection; Inflammation; Introns; Keratitis; Laboratories; Lesion; Light; Literature; Louisiana; Lytic; Lytic Phase; MEKs; Mediating; Messenger RNA; Mission; Molecular; Orthopoxvirus; Oryctolagus cuniculus; Pathway interactions; Plaque Assay; Play; Production; Proteins; Publishing; Recruitment Activity; Recurrence; Regulation; Research; Reverse Transcriptase Polymerase Chain Reaction; Role; Signal Transduction; Simplexvirus; Students; Testing; Time; Translations; Treatment Protocols; Universities; Viral; Viral Genes; Virus; Virus Diseases; Western Blotting; chromatin immunoprecipitation; lytic replication; novel therapeutics; overexpression; prevent; programs; promoter; public health relevance; recombinant virus; research study; response; transcription factor; virology

DESCRIPTION (provided by applicant): Herpes simplex virus type-1 (HSV-1) lytic infection causes diseases ranging from simple cold sores to dangerous keratitis and lethal encephalitis. The interaction between virus and host cells is being investigated extensively by many laboratories. Our data demonstrated that HSV-1 can rapidly induce the expression of multi-functional transcriptional factor early growth response-1 (Egr-1) during lytic infection in corneal cells. Egr-1 functions as a convergence point for many signaling cascades and is known to play an important role in regulating inflammation, cell proliferation and apoptosis. Western blot analyses showed that Egr-1 was absent in Vero and rabbit corneal cell line SIRC but the protein was detected starting from 24 hours post infection (hpi) and was directly proportional to the amount of virus used for infection. Infection with recombinant virus expressing EGFP followed by immunofluorescent studies revealed that Egr1 was expressed only within the infected cells. Reverse transcriptase polymerase chain reaction (RT-PCR) analyses indicated that Egr-1 mRNA was transcribed as early as 1 hpi and did not require de novo viral gene expression since UV inactivated virus initiated the Egr1 transcription but failed to produce protein. Chromatin Immunoprecipitation (ChIP) assays demonstrated that NFkB and cAMP response element binding protein (CREB) was induced by HSV-1 infection and recruited to the Egr-1 promoter upon infection. Collectively, these results suggest that Egr-1 is efficiently induced upon HSV-1 lytic infection and may play a key role in the viral replication and the disease progression in eyes. In this project, we will further identify the mechanisms that induced the Egr1 expression by HSV-1 infection and investigate the roles of Egr1 on HSV-1 replication through the following Specific Aims. Specific Aim 1. To understand how viral infection induced Egr1 expression. In this Aim, we will examine if viral binding of the cells, viral gene expression, viral replication, or all of the above were required to initiate Egr1 transcription/translation. Specific Aim 2. To determine the mechanism that controlled the transcription of Egr1. In this Aim, we will investigate the pathways that regulated Egr1 transcription. Specific Aim 3. To evaluate the effect of Egr1 on HSV-1 gene expression and replication. Our published data showed that overexpression of Egr1 bound to ICP22 intron and inhibited transcription of ICP4 and ICP22. In this aim, we will use DNAzyme to eliminate Egr1 expression by infection and check if the elimination can affect HSV-1 gene expression and replication. Our immediate goal is to establish an active research program at the University of Louisiana Monroe (ULM) so the students can study Virology since our laboratory is the only research program at ULM and Northeast Louisiana dedicating to virus research. The long-term mission is to identify the regulatory mechanisms controlling viral gene expression and replication to develop novel therapeutic protocols for treatment of this devastating and severe viral disease. PUBLIC HEALTH RELEVANCE: Herpes Simplex Virus -1 (HSV-1) primary and recurrent infection may result in scarring of the cornea and is the leading cause of blindness in US and the developed world. We showed that HSV-1 infection rapidly induced the expression of an important multifunctional protein Egr-1 in corneal cells. The completion of this proposal will shed light on the molecular mechanism of HSV-1 mediated Egr1 expression and assist to develop new strategy to prevent viral replication and blindness caused by HSV-1.

描述（由申请人提供）：单纯疱疹病毒类型1（HSV-1）裂解感染引起的疾病，范围从简单的唇疱疹到危险的角膜炎和致死性脑炎。许多实验室对病毒与宿主细胞之间的相互作用进行了广泛的研究。我们的数据表明，HSV-1可以迅速诱导角膜细胞裂解感染期间多功能转录因子早期生长反应1（EGR-1）的表达。 EGR-1充当许多信号级联反应的收敛点，众所周知在调节炎症，细胞增殖和凋亡方面起着重要作用。蛋白质印迹分析表明，在Vero和兔角膜细胞系SIRC中不存在EGR-1，​​但是从感染后24小时开始检测到该蛋白质，并且与用于感染的病毒量直接成正比。表达EGFP的重组病毒感染，然后进行免疫荧光研究表明，EGR1仅在感染细胞中表达。逆转录酶聚合酶链反应（RT-PCR）分析表明，EGR-1 mRNA早在1 HPI时被转录，并且不需要从头病毒基因表达，因为紫外线灭活病毒启动了EGR1转录，但未能产生蛋白质。染色质免疫沉淀（CHIP）测定表明，HSV-1感染诱导NFKB和CAMP反应元件结合蛋白（CREB），并在感染后招募到EGR-1启动子。总的来说，这些结果表明，在HSV-1裂解感染中有效诱导EGR-1，​​并且可能在病毒复制和眼睛中疾病进展中起关键作用。在该项目中，我们将进一步确定通过HSV-1感染诱导EGR1表达的机制，并通过以下特定目的研究EGR1在HSV-1复制中的作用。特定目的1。了解病毒感染如何诱导EGR1表达。在此目标中，我们将检查细胞的病毒结合，病毒基因表达，病毒复制或上述所有内容是否启动EGR1转录/翻译。具体目的2。确定控制EGR1转录的机制。在此目标中，我们将研究调节EGR1转录的途径。具体目的3。评估EGR1对HSV-1基因表达和复制的影响。我们已发布的数据表明，与ICP22内含子结合的EGR1的过表达并抑制了ICP4和ICP22的转录。在此目标中，我们将使用DNAZyme通过感染消除EGR1表达，并检查消除是否会影响HSV-1基因的表达和复制。我们的近期目标是在路易斯安那大学（ULM）（ULM）建立一个积极的研究计划，以便学生可以学习病毒学，因为我们的实验室是ULM和东北路易斯安那州唯一致力于病毒研究的研究计划。长期任务是确定控制病毒基因表达和复制的调节机制，以开发新的治疗方案来治疗这种毁灭性和严重的病毒疾病。 公共卫生相关性：单纯疱疹病毒-1（HSV -1）原发性感染和反复感染可能会导致角膜疤痕，并且是美国和发达国家失明的主要原因。我们表明，HSV-1感染迅速诱导了角膜细胞中重要的多功能蛋白EGR-1的表达。该提案的完成将阐明HSV-1介导的EGR1表达的分子机制，并有助于制定新的策略，以防止HSV-1引起的病毒复制和失明。