E3L AND POXVIRUS REPLICATION/INTERFERON RESISTANCE

干扰素系统是脊椎动物的主要防御系统之一。 防止病毒感染。最近，一些具有干扰素耐药性的病毒 被刻画出来了。拟议研究的长期目标是 了解病毒对干扰素耐药的机制。 具体目的是了解其中一种痘病毒的作用。 干扰素系统的抑制剂在这些病毒的复制和它们的 对干扰素的耐药性。由于痘病毒是作为媒介发展起来的 对于人类疫苗，可以从干扰素的影响中拯救其他病毒，以及 由于导致人类严重腹泻的C组轮状病毒编码 作为一种类似的PKR抑制剂，这项研究可能会影响健康问题。 痘苗病毒在病毒复制和对干扰素耐药性中的作用 干扰素系统的抑制剂将通过使用突变菌株进行评估 病毒vPl080的一种，该病毒与抗性有关的基因已被删除 干扰素，E3L基因。该基因也被认为与抑制 痘苗病毒感染细胞的凋亡，并可将细胞转化为 文化。激活vp1080中干扰素系统的各种组件- 被感染的细胞将被分析。蜕皮病病毒(另一种痘病毒) 对于删除的E3L同源基因将被制备并致病 改变后的病毒将在小鼠身上进行测试。结构函数关系 对于E3L编码的蛋白质，将确定p25。具体来说，P25 结合双链(DS)RNA，并与其他几个 DsRNA结合蛋白，包括干扰素诱导的蛋白激酶， 结合到HIV反式激活(TAR)位点的人类蛋白，TRBP，和 轮状病毒NSP3蛋白。此中的结构/功能关系 保守的dsRNA结合域将由定点确定 诱变。含有这个结构域的蛋白质的晶体结构将是 下定决心。这项工作可能会为了解蛋白质如何识别提供线索。 DsRNA。其他痘苗病毒--可能影响病毒对 通过选择能够允许植物生长的基因，将分离出干扰素 痘苗病毒干扰素的存在使E3L基因缺失。这项工作将 洞察痘病毒是如何进化出对干扰素的抗药性的。

The interferon (IFN) system is one of the primary vertebrate defenses against virus-infection. A number of IFN-resistant virus have recently been characterized. The long-term objectives of the proposed research are to understand the mechanisms involved in resistance of viruses to IFN. The specific aims are to understand the role of one of the poxvirus inhibitors of the IFN system in replication of these viruses and in their resistance to IFN. Since poxviruses, which are being developed as vectors for human vaccines, can rescue other viruses from the effects of IFN, and since group C rotaviruses, which cause severe diarrhea in humans, encode a similar PKR inhibitor, this research may impact health issues. The role in virus replication and resistance to IFN of the vaccinia virus inhibitor of the IFN system will be evaluated by use of a mutant strain of virus, vPl080, that has been deleted for a gene involved in resistance IFN, the E3L gene. This gene has also been implicated in suppression of apoptosis in vaccinia virus infected cells, and can transform cells in culture. Activation of various components of the IFN system in vp1080- infected cells will be analyzed. Ectromelia virus (another poxvirus) deleted for the E3L homologue will be prepared and pathogenesis of this altered virus will be tested in mice. Structure function relationships for the protein encoded by E3L, p25, will be determined. p25 specifically binds double-stranded (ds)RNA, and shares homology with several other dsRNA-binding proteins, including the IFN-induced protein kinase, the human protein that binds to the HIV transactivation (tar) site, TRBP, and the rotavirus NSP3 protein. Structure/function relationships in this conserved dsRNA-binding domain will be determined by site-directed mutagenesis. Crystal structures for proteins bearing this domain will be determined. This work may provide light into how proteins recognize dsRNA. Other vaccinia virus-genes that may affect viral sensitivity to IFN will be isolated, by selecting for genes that can allow growth in the presence of IFN of vaccinia virus deleted the E3L gene. This work will provide insight into how the poxviruses have evolved resistance to IFN.