Overexpression of p21 protein in radiation-transformed mouse 10T(1)/(2) cell clones

In order to investigate the hypothesis that aberrant expression of cell-cyc le regulatory proteins may represent early events in the process of carcino genesis, levels of expression of the negative regulators p21(waf1/cip1) (p2 1), p27(kip1) (p27), and p16(ink4a) (P16) and/or the positive regulators cy clin D-1 and cyclin E were examined by western blot analysis in cells trans formed in vitro by ionizing radiation. The levels of these proteins in 12 i ndependently derived mouse 10T1/2 cell clones transformed by 1.5 Gy of alph a radiation were compared with those in nine similarly derived nontransform ed control clones. Constitutive levels of p21 were very low in all control clones, whereas p21 expression was significantly elevated in nine of 12 tra nsformed clones. Two of the three transformed clones displaying low levels of p21 expressed increased levels of p53. p21 regulation was also altered i n response to radiation in transformed clones as compared with controls, on ly minimal induction was observed 4h following gamma irradiation. Western b lot analysis indicated a constant expression of p27 protein but slightly de creased levels of p16 in these transformed clones. Cyclin D-1 was overexpre ssed in 11 of 12 transformed clones; in only two of these were the levels o f cyclin E elevated. Overall, the results suggest that alterations in the e xpression of cell cycle regulatory proteins may represent important events in radiation-induced oncogenic transformation in vitro. Although the specif ic alterations vary among different transformed clones, overexpression and aberrant regulation of p21 appear to be the most frequent ones. (C) 2000 Wi ley-Liss, Inc.