Policastro P F, Fredholm M, Wilson M C
Department of Molecular Biology, Research Institute of Scripps Clinic, La Jolla, California 92037.
J Virol. 1989 Oct;63(10):4136-47. doi: 10.1128/JVI.63.10.4136-4147.1989.
The conversion of endogenous or exogenous murine retroviruses to a leukemogenic phenotype involves recombination with retroviral sequences present in host genomic DNA. In the 129 Gix+ inbred strain, these endogenous sequences are replication defective but still express retroviral proteins under the apparent transcriptional control of the Gv-1 regulatory locus. To study the protein-coding potential of Gv-1-regulated endogenous retroviral loci, we used oligonucleotide probes directed to env deletion breakpoints identified in previously characterized cDNA clones. Four endogenous retroviral loci were isolated from a library of 129 Gix+ genomic DNA with these probes. Three loci cloned with the env deletion probe del env-1 had virtually identical proviral inserts by restriction analysis. A unique locus was identified and cloned with the del env-2 probe, which must therefore represent a Gv-1-responsive element. Restriction enzyme and nucleotide sequence analyses indicated that the del env-1 and del env-2 loci represented members of the polytropic and modified polytropic classes of endogenous retrovirus, respectively. Despite this divergence, members of both classes contained identical deletions of 19 nucleotides within p30gag and of 1,474 nucleotides from p10gag into the reverse transcriptase-coding region of pol, suggesting that a recombination event had occurred between these proviral sequences prior to insertion within the genome. The del env-1 and del env-2 loci retained coding capacity for truncated gag polyproteins, confirmed by in vitro translation and immunoprecipitation of the protein products. Nucleotide sequence comparison of the untranslated leader (L) regions of the del env-1 and del env-2 loci to a replication-competent ecotropic virus indicated regions that might be important to dispersion of these endogenous retroviral elements throughout the host genome.
内源性或外源性鼠逆转录病毒转变为致白血病表型涉及与宿主基因组DNA中存在的逆转录病毒序列发生重组。在129 Gix +近交系中，这些内源性序列复制缺陷，但在Gv - 1调控位点的明显转录控制下仍表达逆转录病毒蛋白。为了研究Gv - 1调控的内源性逆转录病毒基因座的蛋白质编码潜力，我们使用了针对先前鉴定的cDNA克隆中确定的env缺失断点的寡核苷酸探针。用这些探针从129 Gix +基因组DNA文库中分离出四个内源性逆转录病毒基因座。用env缺失探针del env - 1克隆的三个基因座通过限制性分析具有几乎相同的前病毒插入片段。用del env - 2探针鉴定并克隆了一个独特的基因座，因此它必定代表一个Gv - 1反应元件。限制性内切酶和核苷酸序列分析表明，del env - 1和del env - 2基因座分别代表内源性逆转录病毒的多嗜性和修饰多嗜性类别成员。尽管存在这种差异，但这两类成员在p30gag内均含有19个核苷酸的相同缺失，从p10gag到pol的逆转录酶编码区域缺失1474个核苷酸，这表明在这些前病毒序列插入基因组之前发生了重组事件。del env - 1和del env - 2基因座保留了截短的gag多蛋白的编码能力，这通过体外翻译和蛋白质产物的免疫沉淀得到证实。将del env - 1和del env - 2基因座的非翻译前导（L）区域与具有复制能力的亲嗜性病毒进行核苷酸序列比较，发现了可能对这些内源性逆转录病毒元件在整个宿主基因组中的分散很重要的区域。
