VACCINIA VIRUS BIOCHEMICAL GENETICS

近年来，人们越来越清楚地认识到， 延伸是调控真核生物的重要控制点， 基因表达。 我们最近的工作表明， 在牛痘病毒感染期间调节转录延伸。 研究这种调节对于理解 特别是疫苗病毒基因表达，该系统可以证明 是研究转录调控的重要模型 在真核生物中的延伸。 这个项目的目标是了解牛痘的调控 病毒复制后（中间和晚期）基因转录 伸长率 该项目以两个牛痘基因G2R和A18R为中心， 似乎调节复制后基因转录延长 补充活动。 遗传实验表明， A18R基因产物，一种DNA解旋酶，限制了细胞的持续合成能力。 病毒RNA聚合酶，因此作为负转录 延伸因子，而G2R基因产物增强持续合成能力 病毒RNA聚合酶，因此作为阳性 转录延伸因子 生化和遗传实验 涉及其他几种病毒基因产物的调节， 复制后转录延长，包括病毒晚期 转录起始因子（H5R），一种病毒多聚腺苷酸聚合酶亚单位 (J3R)和两个病毒RNA聚合酶亚基（J4R和A24R）。 我们 一个可行的假设是，这些基因产物可能一起起作用， 作为转录延伸复合物，调节3'末端的形成 中期和晚期的牛痘病毒mRNA。 我们建议测试， 通过1）体外转录， 延伸测定，2）现有病毒突变体的表型分析 转录延伸缺陷，和3）分离， 额外的延伸缺陷突变体的表征。

In recent years it has become increasingly clear that transcription elongation is an important control point for regulation of eucaryotic gene expression. Our recent work demonstrates that postreplicative gene transcription elongation is regulated during vaccinia virus infection. A study of this regulation is important for understanding regulation of vaccina virus gene expression in particular, and the system may prove to be an important model for study of regulation of transcription elongation in eucaryotes in general. The goal of this project is to understand the regulation of vaccinia virus postreplicative (intermediate and late) gene transcription elongation. The project centers on two vaccinia genes, G2R and A18R, which seem to regulate postreplicative gene transcription elongation with complementing activities. Genetic experiments indicate that the A18R gene product, a DNA helicase, restricts the processivity of the viral RNA polymerase and therefore acts as negative transcription elongation factor, while the G2R gene product enhances the processivity of the viral RNA polymerase and therefore acts as a positive transcription elongation factor. Biochemical and genetic experiments implicate several other viral gene products in the regulation of postreplicative transcription elongation, including a viral late transcription initiation factor (H5R), a viral poly A polymerase subunit (J3R), and two viral RNA polymerase subunits (J4R and A24R). Our working hypothesis is that these gene products work together, perhaps as a transcription elongation complex, to regulate formation of 3' ends of intermediate and late vaccinia viral mRNAs. We propose to test, refine, and extend this hypothesis through 1) an in vitro transcription elongation assay, 2) phenotypic analysis of existing virus mutants defective in transcription elongation, and 3) isolation and characterization of additional elongation defective mutants.