TRICHINELLOSIS AND REGULATION OF HOST GENE EXPRESSION

旋毛虫病和宿主基因表达的调节

• 批准号: 2624115
• 负责人: DOUGLAS P JASMER
• 金额: $ 21.17万
• 依托单位: WASHINGTON STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-01-01 至 2001-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2624115
• 关键词: DNA replication; Trichinella; cell differentiation; cell growth regulation; disease reservoirs; gene expression; genetic library; genetic regulation; host organism interaction; immunoaffinity chromatography; immunofluorescence technique; immunoprecipitation; intracellular parasitism; laboratory mouse; laboratory rat; molecular cloning; muscle cells; proliferating cell nuclear antigen; striated muscles; trichinosis; western blottings; yeast two hybrid system

Trichinella spiralis is an intracellular nematode parasite of mammalian skeletal muscle cells that causes severe myositis and, sometimes, death in humans. The muscle infection lasts months to years, and domestic and wild mammals represent a continual environmental reservoir for human infection. When infected, terminally differentiated skeletal muscle cells are induced to re-enter the cell cycle, replicate DNA, then become terminally suspended in G2/M and cease muscle gene expression. The nuclei of these cells also become abnormally large (14-17 mu m), suggesting high transcriptional activity. These changes are thought to promote persistence of the intracellular parasite. We have identified parasite antigens that localize to host nuclei and likely regulate some of these changes. Methods were developed to extract these nuclear antigens (NA). It was further shown that mebendazole treatment of mice depletes NA from host nuclei and simultaneously leads to diminution in size of infected cell nuclei and decreased protein, RNA and acid phosphatase levels in infected cells, suggesting a causal relationship. Hence, methods are available to investigate parasite products that we hypothesize to play a central role in regulating the infected cell phenotype. Specific aims to investigate the functions of NA in the host cell are as follows: 1. Clone and evaluate NA genes from T. spiralis by screening a parasite cDNA expression library with antibodies made against NA from infected cell nuclei. 2. Determine the effects of NA gene expression on differentiation and reversal of differentiation in mammalian skeletal muscle cells. 3. Use the yeast two-hybrid system to identify host proteins that interact with NA proteins. Results from the described research are expected to clarify important molecular interactions that regulate this host/parasite relationship. The parasite undermines basic, host cell-cycle and gene regulatory pathways by mechanisms which might provide targets for improved treatments of trichinellosis. Understanding these mechanisms may also contribute unique insight on mechanisms of muscle differentiation and cell cycle regulation in mammalian cells.

旋毛虫是一种寄生于哺乳动物细胞内的线虫 骨骼肌细胞会导致严重的肌炎，有时甚至死亡 在人类身上。 肌肉感染持续数月至数年， 野生哺乳动物是人类赖以生存的环境资源 感染 当被感染时，终末分化的骨骼肌 细胞被诱导重新进入细胞周期，复制DNA， 终末悬浮于G2/M期并停止肌肉基因表达。 的 这些细胞的核也变得异常大（14-17 μ m）， 表明高转录活性。 这些变化被认为是 促进细胞内寄生虫的持续存在。 我们已经确定 寄生虫抗原定位于宿主细胞核，并可能调节一些 这些变化。 开发了提取这些核的方法 抗原（NA）。 进一步表明甲苯咪唑治疗小鼠 消耗宿主细胞核中的NA，同时导致 感染细胞核的大小和蛋白质、RNA和酸的减少 感染细胞中的磷酸酶水平，表明因果关系。 因此，方法可用于研究寄生虫产品， 假设在调节受感染细胞中起着核心作用， 表型 具体目的是研究NA在宿主中的功能 细胞如下：1.克隆并鉴定了T.旋毛虫 通过用制备的抗体筛选寄生虫cDNA表达文库， 从感染的细胞核中分离NA。 2.确定NA的影响 分化和逆转分化的基因表达 哺乳动物骨骼肌细胞 3.利用酵母双杂交系统 鉴定与NA蛋白相互作用的宿主蛋白。 结果 所描述的研究预计将阐明重要的分子 调节这种宿主/寄生虫关系的相互作用。 的 寄生虫破坏基本的宿主细胞周期和基因调控途径 通过可能为改善治疗提供目标的机制， 旋毛虫病 了解这些机制也可能有助于 对肌肉分化和细胞周期机制的独特见解 在哺乳动物细胞中的调节。