Molecular analysis of DNA isolated from the different stages of x-ray-induced transformation in vitro

A major challenge in radiation carcinogenesis is to identify the cellular gene or genes involved in initiating the process. We examined the transforming activities of DNAs obtained from C3H10T1/2 cells during x-ray-induced morphological transformation. DNAs extracted from mass cultures of 10T1/2 cells at different times after irradiation with 600 rad and from type III-transformed foci were transfected into NIH 3T3 cells. The results indicate that certain oncogenes are activated beginning 3 wk after irradiation, well before the appearance of macroscopically visible transformed foci. For DNA isolated from x-ray-transformed 10T1/2 cells (type III foci), the frequencies of transfection were 0.003-0.11 foci/microgram of genomic DNA with NIH 3T3 cells and 0.004-0.04 foci/microgram genomic DNA using 10T1/2 cells as recipients. Southern blot analysis of DNAs obtained from 23 primary transfectants and from 23 x-ray-transformed cell lines indicated no gross rearrangements or amplification of any of the 14 oncogenes screened (v-Ha-ras, v-Ki-ras, N-ras, v-myc, v-raf, v-src, v-fes, v-abl, v-mos, v-erbA, v-erbB, v-myb, v-fos, v-sis). This suggests that x-irradiation may activate as yet unidentified oncogenes. The occurrence of positive transfection 3 wk after irradiation is discussed in terms of the hypothesis that transformation may not occur as a direct consequence of the exposure to x-rays but develops as a rare event in the progeny of the irradiated cells at some later time, as a consequence of the delayed activation of certain genes.