Mechanism of VEE Attenuation

描述：（改编自研究者摘要）：委内瑞拉马 脑炎病毒（VEE）是一种重要的马和人类病原体。它 周期性地出现在由高毒性动物流行菌株引起的流行病中。 天然VEE菌株的毒力决定因素尚未完全了解， 分子水平。在疫苗株和实验室减毒株中， 小鼠的毒力决定簇已经定位于E1和E2糖蛋白， 5'非翻译区（UTR）。在核苷酸3（nt 3）的单一变化， 将VEE基因组的5' UTR（G至A置换）引入分子生物学系统中， 野生型毒性VEE的克隆导致完全 在小鼠中减毒。长期目标是确定 VEE基因组的5' UTR中的特异性突变使病毒减毒， 使用VEE系统来阐明IFNAB在甲病毒发病机制中的作用。 我们假设nt 3A突变导致病毒增加 对IFNaB的抗病毒作用的敏感性由于病毒水平的增加而降低。 dsRNA复制中间体，因此（a）增加dsRNA复制中间体的诱导， IFN-刺激的抗病毒基因，（B）增强IFN-诱导的抗病毒基因的活化， 需要dsRNA作为辅因子的抗病毒蛋白，或者可替代地，（C） nt 3A突变病毒可能在抑制宿主蛋白质合成方面较慢， 继续合成关键的抗病毒活性。我们将检验这些假设 在以下具体问题的背景下：1）什么是相对 INFaB诱导的抗病毒途径PKR和2 '-5' OAS/RNase L在 与nt 3A突变体VEE相比，野生型VEE体外复制抑制 在vivo？2)nt 3A突变是否影响诱导和/或 下游IFNaB效应子机制如PKR，2 ′-5 ′ OAS， RNase L和阿达尔？和3）nt 3A突变对 病毒诱导的宿主蛋白质合成抑制？

DESCRIPTION: (Adapted from the Investigator's abstract): Venezuelan equine encephalitis virus (VEE) is an important equine and human pathogen. It periodically emerges in epidemics caused by highly virulent epizootic strains. Virulence determinants of the natural VEE strains are not well understood at the molecular level. In vaccine strains and laboratory attenuated strains, virulence determinants for mice have been mapped to El and E2 glycoproteins and the 5' untranslated region (utr). A single change at nucleotide 3 (nt3) of the 5' utr of the VEE genome (G to A substitution) introduced into a molecular clone of wild-type virulent VEE results in a virus that is completely attenuated in mice. The long-term goal is to determine the mechanisms by which specific mutations in the 5' utr of the VEE genome attenuates the virus, and to use the VEE system to elucidate the role of lFNAB in alphaviruses pathogenesis. We hypothesize that the nt3A mutation results in a virus with increased sensitivity to the antiviral actions of lFNaB due to increased levels of viral dsRNA replication intermediates, consequently (a) increasing induction of IFN-stimulated antiviral genes, (b) enhancing activation of IFN-induced antiviral proteins that require dsRNA as cofactor, or alternatively, (C) the nt3A mutant virus may be slower in inhibiting host protein synthesis, allowing continued synthesis of key antiviral activities. We will test these hypotheses in the context of the following specific questions: 1) What is the relative contribution of INFaB-induced antiviral pathways PKR and 2'-5' OAS/RNase L in the inhibition of replication of wild type compared to nt3A mutant VEE in vitro and in vivo? 2) Does the nt3A mutation affect the induction and/or the activation of downstream IFNaB effector mechanisms such as PKR, 2'-5' OAS, RNase L, and ADAR? and 3) What is the effectof the nt3A mutation on virus-induced host protein synthesis inhibition?