PACKAGING OF THE SEGMENTED GENOME OF BACTERIOPHAGE PHI 6

噬菌体 PHI 6 分段基因组的包装

基本信息

批准号：
2391966
负责人：
LEONARD E MINDICH
金额：
$ 28.43万
依托单位：
PUBLIC HEALTH RESEARCH INSTITUTE
依托单位国家：
美国
项目类别：
财政年份：
1985
资助国家：
美国
起止时间：
1985-01-01 至 1999-03-31
项目状态：
已结题

项目摘要

Bacteriophage theta6 contains three pieces of double-stranded RNA (S, M and L) in its nucleocapsid. The pieces possess different genetic information but have similar termini. The packaging of the genomic segments is precise in that virions contain one each of the three normal segments. The aim of this proposal is to determine how the packing system is able to select one of each of the chromosomes and how the system acts to replicate the genome. The entire genome of theta6 has been cloned as cDNA and completely sequenced. Procapsids are produced from cDNA copies of segment L in both Escherichia coli and Pseudomonas phaseolicola, the natural host of theta6. These procapsids care capable of packaging single stranded viral RNA, synthesizing the complementary minus strand to form double stranded RNA and then transcribing this dsRNA to form plus ssRNA. Procapsids will be isolated and used to study both the replicase and packaging reactions. RNA transcripts made from cDNA constructions of the genomic segments can be packaged by the procapsids. The regions of the RNA involved with packaging will be manipulated so as to clarify the interaction between the RNA and the packaging and replication machinery. We will test a new model of packaging that involves retention of the correct complement of genomic segments by the formation of a ternary complex inside the procapsid after non selective entry. A system has been developed that facilitates the isolation and analysis of mutants in three of the replicase proteins. We will use this system to define the domains in proteins P2, P4 and P7 that are involved in packaging, minus and plus strand synthesis. The nature of the mechanisms of the mechanisms of selective packaging in viruses of segmented genomes is currently unknown. We have developed a novel model that explains the specific packaging. The elucidation of the mechanism for theta6 has relevance to the packaging of other segmented- genome viruses. Several members of the reoviridae, e.g. rotavirus and blue tongue virus, are the etiologic agents of important human and animal diseases.

噬菌体 theta6 含有三段双链 RNA（S、M 和L)在其核衣壳中。这些碎片拥有不同的基因信息但具有相似的术语。基因组的包装片段是精确的，因为病毒粒子包含三个正常片段中的每一个段。该提案的目的是确定包装系统如何能够选择每条染色体之一以及系统如何运作来复制基因组。 theta6的整个基因组已被克隆为cDNA并完全已测序。原衣壳是由片段 L 的 cDNA 拷贝产生的大肠杆菌和绿豆假单胞菌，theta6 的天然宿主。这些衣壳能够包装单链病毒RNA，合成互补负链以形成双链RNA 然后转录该 dsRNA 形成 plus ssRNA。原衣壳将是分离并用于研究复制酶和包装反应。由基因组片段的 cDNA 构建而成的 RNA 转录物可以由原衣壳包装。 RNA 涉及的区域包装将被操纵，以澄清之间的相互作用 RNA 以及包装和复制机器。我们将测试新模型涉及保留基因组正确补体的包装通过在原衣壳内形成三元复合物来分割非选择性进入。已经开发出一个系统，有助于隔离和分析三种复制酶蛋白的突变体。我们将使用这个系统来定义蛋白质 P2、P4 和 P7 中涉及的结构域包装、负链和正链合成。选择性包装机制的本质分段基因组的病毒目前未知。我们开发了一个新颖的模型，解释了具体的包装。的阐明 Theta6 的机制与其他分段的包装相关基因组病毒。呼肠孤病毒科的几个成员，例如呼肠孤病毒科。轮状病毒和蓝舌病毒是重要的人类和动物的病原体疾病。