ISOLATION OF GENES SPECIFICALLY EXPRESSED IN FLAT REVERTANT CELLS DERIVED FROM ACTIVATED RAS-TRANSFORMED NIH-3T3 CELLS BY TREATMENT WITH AZATYROSINE
ISOLATION OF GENES SPECIFICALLY EXPRESSED IN FLAT REVERTANT CELLS DERIVED FROM ACTIVATED RAS-TRANSFORMED NIH-3T3 CELLS BY TREATMENT WITH AZATYROSINE
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DOI：
10.1073/pnas.89.11.4879
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发表时间：
1992-06-01
影响因子：
11.1
通讯作者：
NISHIMURA, S
中科院分区：
文献类型：
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作者：
KRZYZOSIAK, WJ;SHINDOOKADA, N;NISHIMURA, S
我们之前报道过，通过转染活化的人c-Ha-ras 转化的小鼠NIH 3T3 细胞在用抗生素氮杂酪氨酸治疗后变得明显正常。尽管激活的 p21ras 仍然表达，但回复体细胞在没有氮杂酪氨酸的情况下长时间培养期间仍保持其正常表型。氮杂酪氨酸诱导的正常表型可能是由于细胞中某些细胞基因表达的激活导致 ras 功能的抑制。为了鉴定回复体细胞中表达增加的基因，我们对由回复体细胞的mRNA制成的噬菌体cDNA文库中的重组体进行差异筛选，并与由ras转化的NIH 3T3细胞和回复体细胞的mRNA制成的P-32标记的cDNA杂交。发现如此分离的两个克隆与 ras 切除基因 (rrg) 几乎相同，该基因被 Contente 等人鉴定为肿瘤抑制基因。 [Contente, S.、Kenyon, K.、Rimoldi, D. 和 Friedman, R. M. (1990) Science 249, 796-798]。其他确定的基因是 III 型胶原蛋白和 rhoB 基因。发现大约一半的克隆含有与鼠逆转录病毒样脑池内 A 颗粒相对应的序列。我们推测氮杂酪氨酸激活ras转化细胞中的几个细胞基因，其中一些基因，包括rrg，协同作用以抵消ras功能，导致ras转化细胞回复到正常表型。
We previously reported that mouse NIH 3T3 cells transformed by transfection of activated human c-Ha-ras become apparently normal upon treatment with the antibiotic azatyrosine. The revertant cells maintain their normal phenotype during prolonged culture in the absence of azatyrosine, although activated p21ras is still expressed. The normal phenotype induced by azatyrosine could be due to activation of expression of some cellular gene(s) in the cells that results in suppression of ras function. To identify the genes with increased expression in the revertant cells, we adopted differential screening of recombinants from a phage cDNA library made from mRNA of the revertant cells, hybridized with P-32-labeled cDNAs made from mRNAs of the ras-transformed NIH 3T3 cells and the revertant cells. Two clones thus isolated were found to be almost identical to the ras recision gene (rrg), which was identified as a tumor-suppressor gene by Contente et al. [Contente, S., Kenyon, K., Rimoldi, D. & Friedman, R. M. (1990) Science 249, 796-798]. Other genes identified were the collagen type III and rhoB genes. Approximately half the clones were found to contain a sequence corresponding to that of the murine retrovirus-like intracisternal A particle. We speculate that azatyrosine activates several cellular genes in the ras-transformed cells and that some of these genes, including rrg, act cooperatively to counteract ras function, resulting in reversion of the ras-transformed cells to the normal phenotype.