A molecular approach to leukemogenesis: mouse lymphomas contain an activated c-ras oncogene.

Abstract

By inducing mouse thymomas with carcinogens and y-radiation, we have studied the potential of tumor DNA to induce foci in rodent fibroblasts. A high percentage of the tumors used transformed the cultured cells, and the oncogenic phenotype segregated with extra copies of the c-ras gene family. There appears to be selectivity in the activated gene because so far all analyzed tumors induced by carcinogen have activated the N-ras gene, and those induced by radiation have activated the K-ras gene. The K-ras gene is the cellular counterpart of the viral ras oncogene in Kirsten murine sarcoma virus, but the N-ras has not yet been found in a retrovirus. The transformed cells have a marked increase in expression of the oncogene at the RNA and protein level. This model system might be a powerful tool in the study of leukemogenesis.The study of thymic lymphoma development in inbred strains of mice is an excellent model system for tumor development. A defined set of protocols involving either radiation or various chemical carcinogens has been elaborated to induce reproducibly a high incidence of thymic lymphoma in treated mice (1-5), and the use of inbred laboratory strains provides a well-characterized genetic background.In the past few years, a powerful technique has been worked out to study the neoplastic process: high molecular weight DNA from several transformed cell lines and primary tumors "transforms" NIH 3T3 cells that are exposed to a calcium phosphate precipitate of DNA (6-12; for a review see ref. 13).Several genes from human tumor cell lines have been isolated (14-17), and the sequences responsible for transformation have been identified as cellular counterparts (c-ras) of the transforming genes of the viral ras family (18, 19) that were previously found in Kirsten and Harvey murine sarcoma viruses, designated K-ras and H-ras, respectively.Recently a report has appeared showing activation of the H-ras gene in carcinogen-induced (and subsequently transplanted) mouse skin carcinomas (20). We call activation of an oncogene the property of segregating with the transformed phenotype upon DNA-mediated gene transfer, although formal proof will come only after a change is demonstrated in the DNA sequence with respect to the normal gene. We report here induced activation of the N-and K-ras genes in murine tumors of the lymphohemopoietic system by means of carcinogen or -radiation. These findings should be instrumental in studying the early stages of carcinogenesis in this model in vivo system. MATERIALS AND METHODSInduction of the Tumors. Thymic lymphomas were induced by (i) intraperitoneal injection of mice with 30 mg of the chemical carcinogen N-nitrosomethyl urea per kg of body weight once a week for 5 consecutive weeks, starting with 35-to 60-day-old animals, and (ii) exposure at similar ages to whole-body y-ray doses of 150 rads once a week for 4 consecutive weeks. The mice used, derived from crosses between the inbred AKR and RF strains, had low incidence of spontaneous thymic lymphoma.Transformation of C...

The CD2-scl transgene alters the phenotype and frequency of T-lymphomas in N-ras transgenic or p53 deficient mice

“…In the light of these unexpected results (reduced thymic lymphomas and scl transgene expression in many of the histiocytic sarcomas) we decided to examine the co-operative e ects of the scl transgene and N-ras by chemical carcinogenesis of the CD2-scl transgenic mice. Intraperitoneal injection of N-methyl-N-nitrosourea (MNU) results in thymic T-cell lymphomas harbouring mutations of N-ras (Guerrero et al, 1984). We injected 6 ± 10 week old scl transgenic mice (n=23) or non-transgenic littermate controls (n=43) with intraperitoneal MNU.…”

“…Mutations of N-ras have been implicated in T-ALL (Lubbert et al, 1990). Furthermore, overexpression of an activated form of N-ras results in T-cell lymphoma (Guerrero et al, 1984;Harris et al, 1988). Examination of the co-operative e ects of N-ras with scl were chosen for a number of theoretical reasons.…”

rsc: a novel oncogene with structural and functional homology with the gene family of exchange factors for Ral

“…NIH3T3 cells were transfected with 1 mg of vector DNA and 10 mg of 3T3 carrier DNA per plate for colony selection or focus formation as previously described (Guerrero et al, 1984). Anchorage independence assays were performed following previously published protocols (Bouck and di Mayorca, 1979).…”

“…DNAs and RNAs were extracted as previously described (Guerrero et al, 1984). DNAs were digested with rsc oncogene homologous to Ral-GDS D D'Adamo et al restriction enzymes following the manufacturer's recommendations, subjected to agarose gel electrophoresis, and transferred for Southern blot as previously described (Guerrero et al, 1984).…”

“…DNAs were digested with rsc oncogene homologous to Ral-GDS D D'Adamo et al restriction enzymes following the manufacturer's recommendations, subjected to agarose gel electrophoresis, and transferred for Southern blot as previously described (Guerrero et al, 1984). RNAs were, when indicated, subjected to further puri®cation to isolate poly (A) + transcripts.…”