REGULATION OF VIRAL NUCLEIC ACIDS SYNTHESIS IN ANIMAL CELLS

动物细胞中病毒核酸合成的调控

• 批准号: 4696804
• 负责人: M SCHUBERT
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF NEUROLOGICAL DISORDERS AND STROKE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/4696804
• 关键词: DNA directed RNA polymerase; Orthomyxoviridae; Paramyxoviridae; Rhabdoviridae; Vesiculovirus; complementary DNA; defective virus; gene expression; genetic manipulation; genetic mapping; genetic transcription; genome; hybridomas; interfering virus; messenger RNA; molecular cloning; molecular pathology; neurotropic virus; nucleic acid sequence; tissue /cell culture; virus RNA; virus genetics; virus replication; viruslike particle

The RNA dependent RNA polymerase of negative strand RNA viruses transcribes and replicates the viral genome. Despite the fact that many members of these viruses cause disease of the central nervous system, little is known about the RNA polymerase and the multiple enzymatic functions it performs. Since many of these funcions are essential to the virus and differ from those of the host cell, they represent potential targets that could specifically be of the host cell, they represent cellular mechanisms. Therefore, the study of these viral specific mechanisms could be exploited in the treatment or prevntion of viral infections in the central nervous system. In order to study the multiple functions of the polymerase complex, we have cDNA cloned, sequenced, assembled and expressd a functional 241 kilodalton polymease protein of Vesicular stomatitis virus (VSV). We have currently established a eukaryotic cell line which constitutively expresses the polymerase protein. Functionality of the polymerase was demonstrated by complementation and rescue of conditional polymerase mutants of VSV. Hence, this unique system will allow for the first time the identification and dissection of the functional domains of the polymerase of negative strand viruses through site specific mutagenesis. The effect of these mutations on the mutation rate of the polymerase, itself, will also be studied with respect to the establishment and maintenance of persistent infections. Towards these ends, we have generated over 20 mutant recombinant L genes and are currently studying their effects on the performance of the multiple functions of the polymerase complex.

负链RNA病毒的RNA依赖性RNA聚合酶进行转录 并复制病毒基因组。 尽管事实上许多成员 这些病毒会导致中枢神经系统疾病，但人们知之甚少 关于 RNA 聚合酶及其执行的多种酶功能。 由于其中许多功能对于病毒来说是必需的并且不同于病毒 那些宿主细胞的细胞，它们代表了可能的潜在目标 特别是宿主细胞，它们代表细胞机制。 因此，可以利用对这些病毒特异性机制的研究 治疗或预防中枢神经病毒感染 系统。 为了研究聚合酶复合物的多种功能，我们有 cDNA 克隆、测序、组装并表达功能性 241 千道尔顿 水泡性口炎病毒（VSV）的聚合酶蛋白。 我们目前有 建立了组成型表达的真核细胞系 聚合酶蛋白。 聚合酶的功能通过以下方式证明 VSV条件聚合酶突变体的互补和拯救。 因此，这个独特的系统将首次允许识别 以及阴性聚合酶功能域的解剖 通过位点特异性诱变形成链病毒。 这些效果 聚合酶本身的突变率也将受到突变的影响 研究了持久性的建立和维护 感染。 为了实现这些目标，我们已经生成了 20 多个突变体 重组L基因，目前正在研究它们对 聚合酶复合物的多种功能的性能。