IDENTIFICATION OF HUMAN N-ras AS THE COMMON ONCOGENE IN NIH 3T3 CELLS TRANSFORMED WITH DNAs FROM HUMAN PRIMARY HEPATIC CANCER AND HEPATOMA 7402 LINE

DNA was extracted from NIH 3T3 cells transformed with DNAs from human primary hepatic cancer (PHC) and Hepatoma 7402 cell line. The transformant DNA was analyzed by Southern transfer and hybridization with ~(32)P-labeled probes of various oncogenes.The EcoRI 7.2 and 9.0 kb bands characteristic of human N-ras gene were identified in transformed NIH 3T3 cells derived both from PHC and 7402 DNA. The BamHI 6.6kb band characteristic of human c-Ha-ras I was present only in 7402 transformants, but not in PHC transformants.Using ~(35)S-methionine incorporation, immunoprecipitation with anti-p21 monoclonal antibodies, SDS-PAGE and autoradiography, it was demonstrated that p21 synthesis was remarkably enhanced in 1402 cells as well as in transformed cells derived from both 7402 and PHC DNA.Taking the data together, it. strongly implies that N-ras is one of the transforming genes for human liver cancer.     

人原发性肝癌中N-ras基因的表达

对9例人原发性肝癌,1例癌旁组织,1例正常肝的poly(A)~ RNA进行了分析。用各种癌基因探针作分子杂交,发现在6例原发性肝癌中,有二条增强表达的区带:2.2kb和5.6kb。肝癌组织较正常肝的mRNA在2.2kb处有明显的增强。提示人N-ras基因的转录产物明显增高。癌旁,正常肝中N-ras基因的专一的mRNA很弱或低于检测水平。由于癌的发生是通过癌基因产物发生作用,因此N-ras基因在多数的人原发性肝癌中的表达明显增强,提示了N-ras基因是人原发性肝癌的重要转化基因之一。     

EXPRESSION OF N-ras GENE IN HUMAN PRIMARY HEPATOCARCINOMA

Poly(A)~+ RNA was isolated from 9 specimens of human primary hepatic carcinoma, 1 non-tumorous liver tissue adjacent to cancer and 1 normal liver tissue samples. The Oligo-dT cellulose-purified poly(A)~+ RNAs were subjected to formaldehyde agarose gel electrophoresis, Northern transfer and hybridization with various ancogene probes. Two RNA species, 5.6 kb and 2.2 kb wore identified by N-ras gone hybridization in 6 out of 9 mRNA samples from primary hepatic carcinoma specimen. N-ras specific mRNA was not detectable in mRNA samples from normal human liver and tumor surrounding cirrhotic tissue. No detectable hybridization of mRNA from hepatoma and normal liver with Ki-ras or Ha-ras was observed.As human N-ras gene has been identified in DNA of mouse transfeetants transformed with PHC DNA, it strongly suggests that N-ras gene might be responsible for the transforming activity of part of cases of human liver cancer.     

人原发性肝癌及7402细胞株和髓细胞白血病细胞株(K562)的癌基因(转化基因)的共同属性——N-ras基因

用人的原发性肝癌(PHC)、肝癌7402细胞株(郭婵等,1984)及髓细胞白血病K562细胞株的DNA转染的NIH/3T3转化株DNA,以Southern吸印和~(32)P-标记的癌基因探针做分子杂交来分析,结果如下:在PHC DNA转化的NIH/3T3细胞DNA中,鉴定有N-ras基因的7.2及9.0kb(EcoRI酶切)的带,与人白细胞和肝癌DNA中出现的专一性条带相同,但是没有人Ha-ras(BamHI 6.6kb)或Ki-ras(EcoRI 3.0kb)的专一性条带。在以7402及K562细胞株DNA转化的细胞DNA中,同时发现有人的N-ras和Ha-ras基因(6.6kb,BamHI酶切),人的Ki-ras特异片段(3.0kb,EcoRI酶切)没有被检测到。PHC的DNA中N-ras基因在带型及杂交强度上没有大的改变,由于N-ras基因(非Ki-ras,Ha-ras)的表达,在多数PHC中明显地增加,提示了N-ras至少是人肝癌的转化基因之一。