Velu T J, Vass W C, Lowy D R, Tambourin P E
Laboratory of Cellular Oncology, National Cancer Institute, Bethesda, Maryland 20892.
J Virol. 1989 Mar;63(3):1384-92. doi: 10.1128/JVI.63.3.1384-1392.1989.
The rat-derived Harvey murine sarcoma virus (Ha-MuSV) contains a transduced ras oncogene activated by two missense mutations and flanked by rat retroviruslike VL30 sequences. Ha-MuSV induces focal transformation of mouse NIH 3T3 cells in vitro and tumors (fibrosarcomas and splenic erythroleukemias) in newborn mice. We have used these two assays to study the contribution of coding and noncoding viral sequences to the biological activity of Ha-MuSV. A good correlation was found between the in vitro and in vivo assays. In several different isogenic Ha-MuSV variants, those with a rasH gene that had one or both of the Ha-MuSV missense mutations were much more active biologically than the corresponding proto-oncogene. A Ha-MuSV variant that encoded the proto-oncogene protein induced lymphoid leukemias (with thymomas), with a relatively long latent period, rather than the fibrosarcomas and erythroleukemias characteristic of Ha-MuSV with one or both missense mutations. A VL30-derived segment with enhancer activity was identified downstream from v-rasH. A mutant Ha-MuSV from which this 3' noncoding segment was deleted expressed lower levels of the wild-type viral protein, displayed impaired transforming activity in vitro, and induced lymphoid leukemias (with thymomas). 5' noncoding rat c-rasH sequences were found to increase the biological activity of the virus when substituted for the corresponding segment of v-rasH. We conclude that (i) the biological activity of Ha-MuSV can be influence significantly by noncoding sequences located outside the long terminal repeat as well as by coding sequences, (ii) VL30 sequences positively regulate the expression of v-rasH, (iii) relatively low biological levels of ras, whether resulting from low-level expression of wild type v-rasH or high-levels of ras proto-oncogene protein, induce a type of tumor that differs from tumors induced by high biological levels of ras, and (iv) the in vivo pathogenicity of the Ha-MuSV variants correlated with their transforming activity on NIH 3T3 cells.
源自大鼠的哈维鼠肉瘤病毒（Ha-MuSV）含有一个经转导的ras癌基因，该基因由两个错义突变激活，并两侧伴有大鼠逆转录病毒样VL30序列。Ha-MuSV在体外诱导小鼠NIH 3T3细胞发生灶性转化，并在新生小鼠中诱发肿瘤（纤维肉瘤和脾红细胞白血病）。我们利用这两种检测方法来研究编码和非编码病毒序列对Ha-MuSV生物学活性的贡献。体外和体内检测之间发现了良好的相关性。在几种不同的同基因Ha-MuSV变体中，那些具有一个或两个Ha-MuSV错义突变的rasH基因的变体在生物学上比相应的原癌基因更具活性。一个编码原癌基因蛋白的Ha-MuSV变体诱发了淋巴白血病（伴有胸腺瘤），潜伏期相对较长，而不是具有一个或两个错义突变的Ha-MuSV所特有的纤维肉瘤和红细胞白血病。在v-rasH下游鉴定出一个具有增强子活性的VL30衍生片段。一个缺失该3'非编码片段的突变Ha-MuSV表达的野生型病毒蛋白水平较低，在体外显示出受损的转化活性，并诱发淋巴白血病（伴有胸腺瘤）。当用5'非编码大鼠c-rasH序列替代v-rasH的相应片段时，发现其可增加病毒的生物学活性。我们得出结论：（i）Ha-MuSV的生物学活性可受到长末端重复序列以外的非编码序列以及编码序列的显著影响；（ii）VL30序列正向调节v-rasH的表达；（iii）无论是野生型v-rasH的低水平表达还是原癌基因蛋白的高水平表达导致的相对低生物学水平的ras，都会诱发一种与高生物学水平的ras所诱发的肿瘤不同的肿瘤类型；（iv）Ha-MuSV变体的体内致病性与其对NIH 3T3细胞的转化活性相关。
