MOLECULAR ANALYSIS OF HSV-I REACTIVATION FROM LATENCY

HSV-I 潜伏期再激活的分子分析

• 批准号: 6488932
• 负责人: Nancy M. Sawtell
• 金额: $ 22.04万
• 依托单位: CINCINNATI CHILDRENS HOSP MED CTR
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-01-01 至 2002-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6488932
• 关键词: genetic transcription; herpes simplex virus 1; laboratory mouse; latent virus infection; molecular genetics; mutant; neurons; polymerase chain reaction; site directed mutagenesis; virus genetics; virus infection mechanism; virus replication

The long term goal of the proposed research is to define the molecular mechanisms involved in the transition from latent to lytic herpes simplex virus (HSV) gene transcription. They previously developed the hyperthermic stress (HS) reactivation model which is unique in that the production of infectious virus is detectable within 12-14 hours after the induction stimulus. Using this model, they have identified what is likely to be a key event in the regulation of reactivation, namely the up regulation of ICPO within 1 hour post HS. Insight into the molecular regulation of reactivation must ultimately be obtained from analysis of individual latently infected neurons. They have developed a new method, contextual expression analysis, CXA, to obtain quantitative information about the DNA and RNA in individual cells within solid tissues. In this proposal, the power of PCR and RT PCR will be harnessed through CXA to construct a molecular definition of latency and reactivation. Their ability to precisely quantify the number of latently infected neurons in the ganglia and examine the RNA and DNA content will allow them to meaningfully evaluate wild type and genetically engineered mutant strains to achieve the following specific aims: (1) Determine the impact of the number of latently infected neurons and/or the number of viral genome copies within individual latently infected neurons and/or the number of viral genome copies within individual latently infected neurons upon the initiation and progression of HS inducted reactivation in vivo; (2) Utilize CXA-RNA strategies to characterize viral transcription during latency and following HS induced reaction at the neuronal population and single cell level; (3) Determine the biological significance and biochemical basis of the rapid up regulation of the ICPO gene following HS induced reaction in vivo. Defining the regulatory mechanisms by which the "latent" repository of viral genetic information periodically give rise to infectious virus is central to understanding this important aspect of the viral life cycle. Insight into these viral functions could contribute significantly toward our ability to design effective vaccines, develop treatments for the prevention of recurrent disease, and efficiently transfer, maintain and regulate foreign genes in the human host.

拟议的研究的长期目标是定义分子 从潜在到裂解疱疹的过渡涉及的机制 病毒（HSV）基因转录。他们以前发展了高温 压力（HS）重新激活模型是独一无二的 感染后12-14小时可检测到感染性病毒 刺激。使用此模型，他们已经确定了可能是 重新激活调节的关键事件，即对 ICPO在HS后1小时内。 对重新激活分子调节的洞察力最终必须是 从分析单个延迟感染的神经元获得。他们有 开发了一种新方法，即上下文表达分析CXA，以获得 有关单个细胞中DNA和RNA的定量信息 固体组织。在此提案中，PCR和RT PCR的功能将是 通过CXA利用以构建潜伏和的分子定义 重新激活。他们精确量化潜在的数量的能力 神经节中感染的神经元并检查RNA和DNA含量将 让他们有意义地评估野生类型和基因工程 突变菌株以达到以下特定目的：（1）确定 潜在感染神经元和/或数量的影响 病毒基因组副本在单个潜在感染的神经元和/或 单个潜在感染神经元内的病毒基因组副本数量 在体内诱导重新激活HS的引发和进展之后； （2）利用CXA-RNA策略来表征病毒转录 延迟并在HS诱导的神经元种群中引起反应，并 单细胞水平； （3）确定生物学意义和 HS之后ICPO基因的快速调节的生化基础 体内诱导反应。定义法规机制 病毒遗传信息的“潜在”存储库会定期引起 感染性病毒是理解这一重要方面的核心 病毒生命周期。对这些病毒功能的洞察力可能有助于 显着朝着我们设计有效疫苗的能力，开发 预防复发性疾病的治疗方法有效 转移，维持和调节人类宿主中的外国基因。