Genetic Aspects Of Viral Oncogenesis In Wild Mouse Species

小鼠白血病病毒（MLV）是与诱发肿瘤、神经和免疫缺陷疾病相关的γ逆转录病毒。 实验室小鼠和野生小鼠物种的近交系在它们对小鼠γ逆转录病毒感染和病毒诱导的疾病的易感性方面不同，并且它们在它们携带的MLV类型方面也不同。 易感性差异是由于特定宿主基因的变化，我们一直在进行持续的努力，以确定和表征几个宿主基因，无论是参与病毒抗性或有助于疾病的过程。 有两种类型的宿主基因参与病毒诱导的疾病。 首先，小鼠基因组包含小鼠γ逆转录病毒基因组的拷贝，其中许多可以产生感染性和致病性病毒。 其次，也有直接干扰病毒感染和复制的宿主因素，我们特别感兴趣的是那些抑制病毒进入和病毒复制周期的早期进入后阶段的因素。 在进入水平上，抗性可由细胞表面受体的多态性引起。 在γ-逆转录病毒进入受体细胞后，逆转录和向细胞核的易位可被病毒抗性因子Fv 1、mApobec 3/Rfv 3和TRIM 5 α抑制或改变。 我们目前的目标是表征这些活性内源性逆转录病毒和宿主编码的抗性因子及其病毒靶点。 最终目的是确定小鼠进化中抗病毒活性的起源和程度，并阐明负责的机制。 这项工作在很大程度上依赖于野生小鼠，因为实验室菌株只提供了一个有限的采样Mus的遗传多样性。此外，野生小鼠物种使我们能够研究在自然种群中的生存策略，窝藏病毒，并遵循抗性基因的进化。 这些小鼠还提供了新的抗性基因和病毒变体的来源。 一组项目旨在确定负责病毒结合和进入的病毒和细胞受体决定因素。 我们目前正在研究异嗜性/多变性MLV（XP-MLV）的XPR 1受体。我们已经确定，在暴露于感染性病毒的小鼠群体中，病毒抗性是由细胞表面受体的多态性介导的。我们已经确定了野生小鼠共6个XPR 1易感性变体，并描述了这些小鼠Xpr 1变体的地理和物种分布。其中五种受体限制两种或更多种依赖于XPR 1的病毒宿主范围变体的进入，并且所有这些受体都在暴露于X-MLV的人群中进化。 几乎所有的哺乳动物物种都可以被X-MLV感染，尽管它们用于进入的细胞表面受体XPR 1存在实质性的序列变异。 为了确定为什么逃逸变体在哺乳动物中很罕见，我们研究了XPR 1中进入决定簇的进化，这些决定簇位于其第三和第四个假定的细胞外环（ECL）中。 关键的ECL 3受体决定簇覆盖剪接供体位点，并且在脊椎动物XPR 1基因中是进化保守的。 13个残基的ECL 4是高变的，但即使整个ECL被水母基因的相应片段取代，这种变异性也不会消除受体功能。 沿着该ECL长度的缺失沿着可以影响但不能消除受体功能，并且不同的缺失影响不同的XP-MLV。 因此，受体使用受约束的剪接位点和耐受突变的环限制了宿主逃逸突变的可能性。 在另一系列实验中，我们使用系统发育和分子生物学方法来鉴定在测序的C57 BL小鼠基因组中发现的内源性XP-MLV的野生小鼠来源。 我们追踪了12种X-MLV到亚洲小鼠物种，但未能在任何野生小鼠中鉴定出任何P-MLV内源性逆转录病毒（ERV）。 我们现在正在研究这些ERV产生传染性病毒的能力。 我们正在对25个特征良好的病毒分离株进行测序，其中7个是从地理上分离的小家鼠亚种中分离的天然MLV，18个是从病毒诱导的肿瘤中分离的分离株。所有18个代表亲嗜性MLV（E-MLV）和非亲嗜性ERV的从头重组体。 这些病毒分为两类：致白血病病毒和非致白血病病毒。 在接种的新生儿中不引起白血病发生的分离株是由P-MLV和E-MLV的重组引起的。 然而，致白血病分离株是E-MLV、P-MLV和X-MLV的重组体。 这两种表型不同的病毒在病毒包膜基因的跨膜亚基的区段中彼此不同。 我们还鉴定了7种来自野生小鼠物种的病毒，以记录这些病毒在自然种群中的进化历史，并确定它们的起源。 这些病毒的分析表明，与特定的主机限制因素的共同进化的证据，表现为在复制过程中与主机因素相互作用的病毒网站的多态性的时间和地理分布。

Mouse leukemia viruses (MLVs) are gammaretroviruses linked to induction of neoplasms, and neurological and immunodeficiency diseases. Inbred strains of laboratory mice and wild mouse species differ in their susceptibility to mouse gammaretrovirus infection and to virus-induced diseases, and they also differ in the types of MLVs that they carry. Susceptibility differences are due to variations in specific host genes, and we have been engaged in an ongoing effort to identify and characterize several host genes that are either involved in virus resistance or that contribute to the disease process. There are two types of host genes involved in virus-induced disease. First, the mouse genome contains copies of mouse gammaretrovirus genomes, many of which can produce infectious and pathogenic viruses. Second, there are also host factors that interfere directly with virus infection and replication, and we are particularly interested in those factors that inhibit virus entry and the early post-entry stages of the virus replicative cycle. At the level of entry, resistance can be caused by polymorphisms in the cell surface receptors. After the gammaretrovirus enters the receptive cell, reverse transcription and translocation to the nucleus can be inhibited or altered by virus resistance factors Fv1, mApobec3/Rfv3, and TRIM5alpha. Our current aim is to characterize these active endogenous retroviruses and the host encoded resistance factors and their viral targets. The ultimate goal is to define the origin and extent of antiviral activity in Mus evolution, and elucidate the responsible mechanisms. This work relies heavily on wild mice because laboratory strains provide only a limited sampling of the genetic diversity in Mus. Also, wild mouse species allow us to examine survival strategies in natural populations that harbor virus and to follow the evolution of the resistance genes. These mice additionally provide a source of novel resistance genes and virus variants. One set of projects aims to identify viral and cell receptor determinants responsible for virus binding and entry. We are currently working on the XPR1 receptor for the xenotropic/polytropic MLVs (XP-MLVs). We have determined that, in mouse populations exposed to infectious virus, virus resistance is mediated by polymorphisms of the cell surface receptor. We have identified a total of six XPR1 susceptibility variants in wild mice and described the geographic and species distribution of these Mus Xpr1 variants. Five of these receptors restrict entry by two or more of the virus host range variants that rely on XPR1, and all of these receptors evolved in populations exposed to X-MLVs. Virtually all mammalian species can be infected by X-MLVs, despite substantial sequence variation in the cell surface receptor they use for entry, XPR1. To determine why escape variants are rare in mammals, we examined the evolution of the entry determinants in XPR1, which lie in its third and fourth putative extracellular loops (ECLs). The critical ECL3 receptor determinant overlies a splice donor site and is evolutionarily conserved in vertegrate XPR1 genes. The 13 residue ECL4 is hypervariable, but this variability does not abolish receptor function, even when the entire ECL is replaced by the corresponding segment of the jellyfish gene. Deletions along the length of this ECL can influence but not abolish receptor function, and different deletions affect different XP-MLVs. Thus, receptor usage of a constrained splice site and a loop that tolerates mutations limits the likelihood of host escape mutations. In another series of experiments, we used phylogenetic and molecular biological methods to identify the wild mouse origins of the endogenous XP-MLVs found in the sequenced C57BL mouse genome. We traced 12 X-MLVs to Asian mouse species, but failed to identify any of the P-MLV endogenous retroviruses (ERVs) in any wild mice. We are now looking at the ability of these ERVs to generate infectious virus. We are in the process of sequencing 25 well characterized virus isolates, 7 of which are naturally occurring MLVs isolated from geographically separated Mus musculus subspecies, and 18 are isolates from virus-induced tumors. All of the 18 represent de novo recombinants of ecotropic MLVs (E-MLVs) and nonecotropic ERVs. These viruses fall into two groups: leukomogenic and nonleukemogenic. The isolates that do not cause leukemogenesis in inoculated neonates result from recombination of P-MLVs and E-MLVs. The leukemogenic isolates, however, are recombinants of E-MLVs, P-MLVs and X-MLVs. These two phenotypically distinct viruses differ from one another in a segment of the transmembrane subunit of the virus envelope gene. We have also characterized 7 viruses from wild mouse species to document the evolutionary history of these viruses in natural populations and to determine their origins. Analysis of these viruses show evidence of coevolution with specific host restriction factors as demonstrated by temporal and geographic distribution of polymorphisms at the virus sites that interact with host factors during replication.