STRUCTURE AND REPLICATION OF HEPATITIS DELTA VIRUS

HDV是一种人类病原体，通常与更具破坏性的HBV相关 肝脏感染。长期目标是理解小说 HDV复制的机制，重点是迄今为止还不知道的功能， 发生在其他动物病毒的复制过程中。四个目标是 建议：I.体外RNA合成：令人惊讶的核提取物 正常人肝细胞转录基因组和反基因组HDV RNA。为了表征这种不寻常的RNA指导的宿主酶合成， 将制备核提取物，加入HDV RNA，并对HDV RNA进行研究。 转录复合物的组装，转录的起始，以及 合成全长或更长的转录物。RNA序列或 启动RNA定向合成所需的结构将是 评估，以及纯化的小δ蛋白的作用（如果有的话）。 二.体内RNA合成：基因组复制可以通过以下方式启动： 用体外组装的由HDV RNA制成的RNP复合物转染细胞 和重组δ蛋白。这种策略模仿自然感染 但不需要原代肝细胞（已知表达 病毒受体）。改变RNP的组成部分将决定 RNA和蛋白质的要求，包括检查 δ蛋白在感染起始中的作用。5 '-RACE程序 将绘制假定的转录起始位点， 改变邻近区域的RNA序列和结构的后果 该等网站三.细胞中mRNA转录物的5 '-加帽和核输出 体内：研究将集中在天然HDV mRNA是否具有帽结构 和顺式作用的核输出信号，并将有助于了解核 大HDV RNA的保留。四.δ的翻译后修饰 蛋白质：大δ蛋白可以磷酸化，是唯一的 蛋白质，来自任何病毒，已知被法尼基化。实验将 解决这两个修改的明显联系以及它们的 复制中的角色。总之，研究提出了四个具体目标， 将提供HDV生命周期独特方面的信息， 因为HDV可能比其他任何设备更依赖于主机功能， 动物病毒，关于宿主机械的信息，如宿主如何轮询 II被重定向到复制RNA模板以及如何翻译后 修饰可以被病毒蛋白质破坏。

HDV is a human pathogen typically associated with more damaging HBV infections of the liver. The long-range goal is to understand the novel mechanism of HDV replication, focusing on features not so far known to occur during the replication of other animal viruses. Four aims are proposed: I. RNA synthesis in vitro: Surprisingly nuclear extracts from normal human liver cells transcribe both genomic and anti-genomic HDV RNAs. To characterize this unusual RNA-directed synthesis by host enzymes, nuclear extracts will be prepared, HDV RNAs added, and studies made of the assembly of transcription complexes, initiation of transcription, and synthesis of full-length or longer transcripts. RNA sequences or structures needed for initiation of RNA-directed synthesis will be assessed, and also the effects, if any, of purified small delta protein. II. RNA synthesis in vivo: Genome replication can be initiated by transfecting cells with in vitro assembled RNP complexes, made of HDV RNAs and recombinant delta proteins. This strategy mimics natural infections but does not require primary hepatocytes (the only cells known to express the virus receptor). Altering components of the RNP will determine the requirements of both RNA and protein, including an examination of the roles of delta proteins in the initiation of infection. 5'-RACE procedures will map putative transcription initiation sites help determine the consequences of altering RNA sequence and structure in the vicinity of such sites. III. 5'-Capping and nuclear export of mRNA transcripts in vivo: Studies will focus on whether natural HDV mRNA has a cap structure and a cis-acting nuclear export signal, and will help understand nuclear retention of large HDV RNAs. IV. Post-translational modifications of delta proteins: Large delta protein can be phosphorylated and is the only protein, from any virus, known to be farnesylated. Experiments will address an apparent linkages of these two modifications as well as their roles in replication. In summary, studies proposed in four specific aims will provide information on unique aspects of the HDV life cycle and, because HDV is possibly more dependent on host functions than any other animal virus, information on the host machinery, such as how the host pol II is redirected to copy RNA templates and how post-translational modifications can be subverted by viral proteins.