EXPRESSION AND REGULATION OF VACCINIA VIRUS LATE GENES

负责基因表达和基因表达的分子机制 痘苗病毒(VV)晚期基因的调控仍然是一个谜。这节课 代表大约一半的VV遗传潜力的VV基因是 仅在病毒DNA合成开始后表达。VV晚期转录本 显然，来自任何单个基因座的基因大小高度不同 从多个不同的5‘端开始并随机终止。 到目前为止，少数几个VV晚期基因已经被定位和测序， 披露了之前明显缺乏监管要素的情况 在其他真核或原核系统中被认为是必需的，甚至 VV早期基因。因此，VV晚期基因控制区和 据推测，识别它们的病毒蛋白是未知的。由于 情况的复杂性，解决这些问题的一种方法是 选择几个感兴趣的VV晚期基因，并对它们进行密集的 分子生物学检查。为此，一些VV基因 它们参与VV复制的后期[Alpha-Amanitin 电阻、TS17和六个非配位表达的Vv的串联阵列 晚期基因]最近已经被识别、绘制和测序。这个 实验试图使用到目前为止获得的信息作为基础： 1)对这些基因如何以及何时进行比较动力学分析 表达。2)制备足以使 编码基因产物的功能鉴定以及它们如何 参与病毒生命周期。3)使用定向遗传学，基因 融合，和标记救援技术，揭示显著的监管 每个基因的特征。4)使用足迹和凝胶延迟 识别相互作用的病毒和/或细胞蛋白的方法学 带有VV启动子和终止子区域。预计这样的情况 实验将提供相当大的洞察力来了解VV 用来实现其复杂发展的有序表达 在受感染的细胞内编程。这一信息可能有助于 未来VV重组疫苗株的设计和构建 用于预防传染病。

The molecular mechanisms that are responsible for the expression and regulation of vaccinia virus (VV) late genes remain an enigma. This class of VV genes, which represents about half of the VV genetic potential, is expressed only after viral DNA synthesis has begun. VV late transcripts from any single locus are highly heterogeneous in size, apparently initiating from a number of distinct 5' sites and terminating randomly. The few VV late genes that have been mapped and sequenced to date, have revealed the conspicious absence of regulatory elements previously recognized as essential in other eukaryotic or prokaryotic systems, or even VV early genes. Thus, the identity of the VV late gene control regions and the viral proteins which presumably recognize them are unknown. Due to the complexity of the situation, one approach to unraveling these questions is to select a few VV late genes of interest, and to subject them to intense molecular biological scrutiny. Towards this end, a number of VV genes which participate in the late phase of VV replication [Alpha-amanitin resistance, ts17, and a tandem array of six non-coordinately expressed VV late genes] have recently been identified, mapped, and sequenced. The experiments seek to use the information obtained thus far as a basis for: 1) A comparative kinetic analysis of how and when these genes are expressed. 2) Preparation of immunological reagents sufficient to enable functional identification of the encoded gene products and how they participate in the viral life cycle. 3) Using directed-genetics, gene fusion, and marker rescue techniques to reveal the salient regulatory features of each gene. 4) Using footprinting and gel retardation methodologies to identify the viral and/or cellular proteins which interact with VV promoter and terminator regions. It is anticipated that such experiments will provide considerable insight into the mechanisms which VV employes to achieve the ordered expression of its complex developmental program within the infected cell. This information may facilitate the design and construction of future VV recombinant vaccine strains to be used for the prophylaxsis of infectious diseases.