GENETIC STRUCTURE OF MURINE RETROVIRUSES

许多逆转录病毒在体内经历基因改变，这可能会影响 病毒致癌、细胞趋向性或免疫消除。例如 转化的禽类和鼠类逆转录病毒已经获得了基因 直接致癌的宿主序列。小鼠白血病 病毒(MuLV)很容易与小鼠基因组序列重组 以产生表现出改变宿主感染性范围的变异体 可以激活细胞致癌基因。慢病毒的点突变 马传染性贫血病毒(EIAV)或人类免疫缺陷 病毒(HIV)可能会导致变异株的产生，这些变异株可以逃脱 宿主免疫反应。这个项目的主要重点是发生 和逆转录病毒基因改变的机制及其后果 这种变化的可能性。在之前的研究中，我们已经检查了组织- 在此之后生成的寄主范围变体的特定表达 接种一种小病毒。因为这种变异的产生会导致 一种混合感染不同宿主范围的病毒，有 一种病毒的基因组有可能出现伪分型 被包裹在另一个病毒粒子的外套里。我们发现了一种独特的 与小鼠白血病发病相关的伪分型模式 感染了淋巴细胞白血病病毒M-MuLV。在两个脾中 而胸腺、寄主范围变体完全被M-MuLV伪分型 在疾病的大部分白血病前期。因此，所有的病毒粒子都表现出 M-MuLV的宿主范围，无论它们包含哪种病毒基因组。 完整的伪分型在整个过程中在脾中持续存在 感染。然而，在胸腺中有很高比例的病毒粒子 展品变种寄主范围最早出现的立即后退 白血病的发病。进一步的研究将集中在这一关系上。 变异的病毒粒子突然向恶性转化。 对逆转录病毒点突变率的研究已经确定了这一比率。 可存活的子代病毒的数量将比以前慢约20倍 报告。这项研究目前正在扩展以确定体内的 MULV的聚合酶错误率。几个前苏联经历了 单个复制周期已经被分子克隆到一个载体中，该载体 允许直接DNA测序以及体外转录和RNA 指纹识别。对这些克隆的分析将在体内产生第一个 真核病毒聚合酶错误率的测定。 在以前的研究中，许多针对MuLV的单抗(MAb) 抗原已经开发出来了。我们已经启动了合作实验 与LMSF的Pincus博士一起构建和测试蓖麻毒素A链 免疫毒素(IT)具有选择性地杀死受 MuLV。几种免疫毒素对受感染的细胞具有细胞毒活性。 其中包括一种免疫毒素，它显示出显著的细胞毒性 感染了所有类别的MuLV和另一种免疫毒素的细胞 几乎完全杀死感染了高度病毒的细胞 神经毒性的MULV。

Many retroviruses undergo genetic alterations in vivo which may affect the viral oncogenicity, cellular tropism, or immune elimination. For example the transforming avian and murine retroviruses have acquired genetic sequences of the host which are directly oncogenic. Murine leukemia viruses (MuLVs) readily undergo recombination with mouse genomic sequences to generate variants which exhibit ah altered host range of infectivity and can activate cellular oncogenic genes. Point mutation of lentiviruses such as the equine infectious anemia virus (EIAV) or the human immunodeficiency virus (HIV) may contribute to the generation of variants which escape the host immune response. The major emphasis of this project is the occurrence and mechanisms of genetic alteration in retroviruses and the consequences of such alterations. In previous studies we have examined the tissue- specific expression of host range variants which are generated after inoculation of an MuLV. Since the generation of such variants results in a mixed infections of viruses with different host ranges, there is a potential for viral pseudotyping to occur in which the genome of one virus is encapsulated in the virion coat of another. We have found a distinct pattern of pseudotyping which correlates with the onset of leukemia in mice infected with the lymphocytic leukemia virus M-MuLV. In both the spleen and thymus, host range variants are completely pseudotyped by the M-MuLV during most of the preleukemic phase of disease. Thus, all virions exhibit the host range of M-MuLV, regardless of which viral genome they contain. Complete pseudotyping persists in the spleen throughout the course of infection. However, in the thymus a high proportion of virions which exhibit the variant host range arise immediately receding the earliest onset of leukemia. Further studies will focus on the relationship of this burst of variant virions to malignant transformation. Studies on the point mutation rate of retroviruses have determined the rate of viable progeny viruses to be approximately 20-fold slower than previous reports. This study is currently being extended to determine the in vivo polymerase error rate for an MuLV. Several proviruses which have undergone a single replication cycle have been molecularly cloned into a vector which allows both direct DNA sequencing as well as in vitro transcription and RNA fingerprinting. Analysis of these clones will yield the first in vivo polymerase error rate determination for a eukaryotic virus. In previous studies numerous monoclonal antibodies (mAb) directed at MuLV antigens have been developed. We have initiated collaborative experiments with Dr. Pincus of the LMSF to construct and test ricin A-chain immunotoxins (IT) for their ability to selectively kill cells infected by MuLVs. Several immunotoxins exhibit cytotoxic activity for infected cells. These include an immunotoxin which exhibits significant cytotoxicity of cells infected with all classes of MuLVs and another immunotoxin which exhibits virtually complete killing of cells infected with a highly neurovirulent MuLV.