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  • Title: Isolation of azatyrosine-induced revertants from ras-transformed human mammary epithelial cells.
    Author: Kyprianou N, Taylor-Papadimitriou J.
    Journal: Oncogene; 1992 Jan; 7(1):57-63. PubMed ID: 1741165.
    Abstract:
    Non-transformed revertant clones were isolated from the ras-transformed MTSV1-7 (ras) cell line, after treatment with the antibiotic azatyrosine. Azatyrosine significantly inhibited the growth of the ras-transformed cells but not of the normal MTSV1-7. After 7 days of azatyrosine treatment, approximately 30% of MTSV1-7 (ras) cells survived, and revertant cell lines were selected by random cloning. The azatyrosine-induced revertants (six clones) were considered non-transformed on the basis of (a) their substantially reduced ability to form colonies in soft agar, and (b) their inability to produce tumours in nude mice. Molecular analysis of the revertants revealed that each contains multiple copies of the v-H-ras gene and expresses high levels of v-H-ras mRNA, and all revertants sustain elevated levels of p21ras protein. Thus, the revertant phenotype induced by azatyrosine does not result from inactivation of v-H-ras oncogene or inhibition of its expression. In vivo guanine nucleotide binding to p21ras in the revertant cell lines demonstrated binding of both GTP and GDP, indicating that reversion to the non-transformed phenotype was not due to inability of p21ras to bind GTP. The expression of the human K-rev-1 gene, a known tumour-suppressor gene in ras-transformed NIH3T3 cells, was studied in the isolated azatyrosine revertants. All six revertants showed a significant increase in the K-rev-1 transcript levels compared with the ras-transformed MTSV1-7 cells. These results suggest that tumorigenic transformation of human mammary epithelial cells by v-H-ras may be influenced by the level of expression of the tumour-suppressor gene, K-rev-1.
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