MOLECULAR ANALYSIS OF RAT EMBRYO CELL TRANSFORMANTS INDUCED BY ALPHA-PARTICLES

Citation
B. Endlich et al., MOLECULAR ANALYSIS OF RAT EMBRYO CELL TRANSFORMANTS INDUCED BY ALPHA-PARTICLES, International journal of radiation biology, 64(6), 1993, pp. 715-726
Citations number
51
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Radiology,Nuclear Medicine & Medical Imaging","Nuclear Sciences & Tecnology
ISSN journal
0955-3002
Volume
64
Issue
6
Year of publication
1993
Pages
715 - 726
Database
ISI
SICI code
0955-3002(1993)64:6<715:MAOREC>2.0.ZU;2-7
Abstract
An immortal cell line was established by transfecting a myc oncogene i nto rat embryo cells (REC:myc)1. This cell line was diploid, contact i nhibited and grew well in culture. Exposure to a single 200 cGy dose o f 6 MeV alpha-particles transformed these cells with a frequency of fo cus formation of approximately 3.6 x 10(-4) compared with a transforma tion frequency of < 7.8 x 10(-6) for primary cultures of REC. Isolates of alpha-particle-induced REC:myc (REC:myc:alpha) foci displayed anch orage-independent growth in soft agar and were tumourigenic in nude mi ce. Molecular studies demonstrated no alteration of gene structure or expression of the transfected or of the endogenous c-myc genes. Simila rly, there was no alteration of the structure of Ha-ras, Ki-ras, or N- ras. The expression of Ha-ras, Ki-ras, N-ras and raf was not altered s ignificantly. Assay for dominant oncogenes via DNA-mediated gene trans fer into NIH3T3 cells was positive for nine of 13 REC:myc:alpha transf ormants. All NIH3T3 isolates contained bands hybridizing to rat repeti tive DNA. NIH3T3 transformants from a tertiary round of transfection w ere analysed by Southern blot analysis for the presence of Ki-ras, N-r as, raf, trk, abl, fms, src, mos, fos, sis, fps, erbA, erbB or neu onc ogenes of REC origin, and none were detected. Tertiary NIH3T3 transfor mants from three REC:myc:alpha transformants contained bands correspon ding to Ha-ras but no point mutations were identified at the known hot spots of exons 1 or 2 of the donor REC:myc:alpha transformants. The in activation of the tumour suppressor genes Rb, and p53, and the antimet astasis gene, nm23, was evaluated by Southern and Northern hybridizati on analysis. Southern blots demonstrated that at least one allele of R b, p53 and nm23 was present and no large scale structural changes were detected. No expression of Rb or p53 was detected in REC:myc or the a lpha-particle-induced REC:myc transformants. The expression of nm23 wa s not altered in the transformed cell lines. While the analysis of the role of tumour suppressor gene inactivation in radiation-induced cell transformation is only in the initial stages, the results of DNA-medi ated gene transfer into NIH3T3 cells suggest that unidentified dominan t oncogenes are associated with alpha-particle-induced transformation in vitro.