Souyri M, Vigon I, Charon M, Tambourin P
Laboratoire d'Immunologie et Oncologie des Maladies Rétrovirales, Hôpital Cochin, INSERM U152, CNRS UA628, Paris, France.
J Virol. 1989 Sep;63(9):3944-53. doi: 10.1128/JVI.63.9.3944-3953.1989.
The N-ras gene is the only member of the ras family which has never been naturally transduced into a retrovirus. In order to study the in vitro and in vivo oncogenicity of N-ras and to compare its pathogenicity to that of H-ras, we have inserted an activated or a normal form of human N-ras cDNA into a slightly modified Harvey murine sarcoma virus-derived vector in which the H-ras p21 coding region had been deleted. The resulting constructions were transfected into NIH 3T3 cells. The activated N-ras-containing construct (HSN) induced 10(4) foci per microgram of DNA and was found to be as transforming as H-ras was. After infection of the transfected cells by either the ecotropic Moloney murine leukemia virus or the amphotropic 4070A helper viruses, rescued transforming viruses were injected into newborn mice. Both pseudotypes of HSN virus containing activated N-ras induced the typical Harvey disease with similar latency. However, we found that the virus which contained normal N-ras p21 (HSn) was also pathogenic and induced splenomegaly, lymphadenopathies, and sarcoma in mice after a latency of 3 to 7 weeks. In addition, Moloney murine leukemia virus pseudotypes of N-ras caused neurological disorders in 30% of the infected animals. These results differed markedly from those of previous experiments in which we had inserted the activated form of N-ras in the pSV(X) vector: the resulting SVN-ras virus was transforming on NIH 3T3 cells but was poorly oncogenic in vivo (M. Souyri, C. F. Koehne, P. V. O'Donnel, T. H. Aldrich, M. E. Furth, and E. Fleissner, Virology 158:69-78). However, similarly poor oncogenicity was also observed when the v-H-ras coding sequence was inserted in pSV(X) vector, which indicated that the vector sequences play a crucial role in the pathogenicity of a given oncogene. Altogether, these data demonstrated unequivocally that N-ras is potentially as oncogenic as H-ras and that such oncogenic effect could depend on the vector environment.
N-ras基因是ras家族中唯一从未自然转导进逆转录病毒的成员。为了研究N-ras在体外和体内的致癌性，并将其致病性与H-ras的致病性进行比较，我们已将人N-ras cDNA的活化形式或正常形式插入到一种经轻微修饰的源自哈维鼠肉瘤病毒的载体中，该载体中的H-ras p21编码区已被删除。将所得构建体转染到NIH 3T3细胞中。含活化N-ras的构建体（HSN）每微克DNA诱导产生10⁴个集落，并且发现其转化能力与H-ras相同。在用亲嗜性莫洛尼鼠白血病病毒或双嗜性4070A辅助病毒感染转染细胞后，将拯救出的转化病毒注射到新生小鼠体内。含有活化N-ras的HSN病毒的两种假型均诱导出具有相似潜伏期的典型哈维病。然而，我们发现含有正常N-ras p21的病毒（HSn）也具有致病性，并且在3至7周的潜伏期后在小鼠中诱导脾肿大、淋巴结病和肉瘤。此外，N-ras的莫洛尼鼠白血病病毒假型在30%的感染动物中引起神经紊乱。这些结果与我们之前将活化形式的N-ras插入pSV(X)载体的实验结果明显不同：所得的SVN-ras病毒在NIH 3T3细胞上具有转化能力，但在体内致癌性较差（M. 苏伊里、C. F. 克内、P. V. 奥唐纳、T. H. 奥尔德里奇、M. E. 弗思和E. 弗莱斯纳，《病毒学》158:69 - 78）。然而，当v-H-ras编码序列插入pSV(X)载体时也观察到类似的低致癌性，这表明载体序列在给定癌基因的致病性中起关键作用。总之，这些数据明确证明N-ras潜在致癌性与H-ras相同，并且这种致癌作用可能取决于载体环境。
