Strategies of lentivirus persistence

慢病毒持续进化和逃避免疫控制的能力是 开发一种有效的艾滋病毒疫苗。病毒在体内的终生持久性是其能力的函数 逃避免疫识别和消除以及它的复制能力，即它的复制能力。这个 拟议的研究将使用表征良好的马传染性贫血病毒(EIAV)模型来识别病毒 慢病毒进展过程中免疫逃逸变异体进化和选择的重要因素 疾病。将使用从感染EIAV的马身上分离出的、处于疾病连续阶段的回溯性样本 为了检验这样一种假设，即成功逃避广泛免疫反应的病毒变种含有 多个基因区域的突变，使复制表型发生可重复的变化。第一 AIM将确定在EIAV疾病连续阶段占主导地位的env/rev基因型是否在 复制表型。含有代表猪传染性支气管炎病毒主要env/rev基因的重组感染性克隆 将在生长动力学和生长竞争分析中测试疾病的每个阶段的复制能力。 将估计变种对的相对适合度，并使用标准统计检验来确定何时 适合性有很大不同。这些研究将确定复制表型是否会导致变异 疾病进展过程中病毒载量的选择和变化。随后的目标将确定是否 复制差异是免疫逃避的代价，是逃避免疫识别的病毒策略，或者是 两者都有。第二个目标的研究将确定REV和SU的变化对复制能力的影响 和免疫逃避。REV复制表型将被量化为核出口活性，并用于 根据CTL杀伤敏感性推断免疫逃避表型。SU复制表型将 被测量为传染性，免疫逃避表型由对中和的敏感性确定 抗体。这些值以及在特定目标1中获得的复制适合度分数将用于 确定哪些env/rev表型影响病毒载量变化的统计模型 疾病。第三个特定目标的实验将确定复制的分子决定因素 对广谱中和抗体敏感性不同的变异体的表型。基于EIAV的伪病毒 将用于绘制EIAV SU在感染性和免疫逃逸鉴定中重要的特定区域 导致临床疾病复发的SU基因改变。这一详细的、综合的分析 将确定关键的病毒决定因素，使平衡向病毒或宿主倾斜 慢病毒病的进展。成功的HIV和其他慢病毒疫苗的设计要求 我们了解、预测和阻止病毒逃避广泛反应性免疫反应的策略。这个 从拟议的研究中获得的信息将确定新的疫苗靶点，以限制病毒从 广泛的反应性免疫反应。项目叙事 拟议中的研究将确定使慢病毒能够修改其复制的遗传机制 并逃脱宿主免疫系统的清除。这一策略允许病毒在体内持续存在，并且 获得可能增加病毒载量并导致临床疾病进展的新突变。结果是 这项研究将确定新的HIV-1疫苗策略，以抑制病毒逃脱 宿主免疫反应。

The ability of lentiviruses to continually evolve and escape immune control is the central impediment in developing an effective vaccine for HIV. The lifelong persistence of virus in vivo is a function of its ability to evade immune recognition and elimination as well as its ability to replicate, i.e. it's replicative capacity. The proposed studies will use the well-characterized equine infectious anemia virus (EIAV) model to identify virus factors important in the evolution and selection of immune escape variants during progression of lentiviral disease. Retrospective samples isolated from EIAV-infected horses at sequential stages of disease will be used to test the hypothesis that virus variants which successfully evade a broadly reactive immune response contain mutations in multiple genetic regions, which confer reproducible changes in replication phenotype. The first aim will determine if env/rev genotypes that predominate at sequential stages of EIAV disease differ in replication phenotype. Recombinant infectious clones containing dominant env/rev genotypes representative of each stage of disease will be tested for replicative capacity in growth kinetic and growth competition assays. The relative fitness of pairs of variants will be estimated, and standard statistical tests used to determine when fitnesses differ significantly. These studies will establish if replication phenotype contributes to variant selection and changes in virus load during progression of disease. Subsequent aims will ascertain whether the replication differences are a cost of immune evasion, a virus strategy to evade immune recognition, or a mix of both. Studies in the second aim will determine the impact of variation in Rev and SU on replicative capacity and immune evasion. The replication phenotype of Rev will be quantified as nuclear export activity, and used to infer an immune evasion phenotype based on sensitivity to CTL killing. The SU replication phenotype will be measured as infectivity, and immune evasion phenotype determined by sensitivity to neutralizing antibody. These values, together with the replication fitness score obtained in Specific Aim 1, will be used in statistical models to determine which env/rev phenotypes affect changes in virus load during progression of disease. Experiments in the third specific aim will identify the molecular determinants of replication phenotype in variants that differ in susceptibility to broadly neutralizing antibody. EIAV-based pseudovirus will be used to map the specific regions of EIAV SU important in infectivity and immune escape identify genetic changes in SU that contribute to recrudescence of clinical disease. This detailed, integrative analyses will identify critical virus determinants that shift the balance in favor of either the virus, or the host, during progression of lentivirus disease. The design of successful vaccines for HIV and other lentiviruses requires that we understand, anticipate, and block viral strategies of evasion from a broadly reactive immune response. The information gained from the proposed studies will identify new vaccine targets that limit virus escape from broadly reactive immune responses. PROJECT NARRATIVE The proposed studies will identify the genetic mechanisms that enable lentiviruses to modify their replication and escape elimination by the host immune system. This strategy allows the virus to persist in vivo, and acquire new mutations that may increase virus load and lead to progression of clinical disease. The results of this study will identify new vaccine strategiesw for HIV-1 that inhibit the viruses ability to escape from the host immune response.