Delayed reactivation of p53 in the progeny of cells surviving ionizing radiation.
Ionizing radiation induces genomic instability, which is transmitted through many generations after irradiation in the progeny of surviving cells. To detect delayed activation of p53, we constructed a reporter plasmid containing the p53-responsible promoter and the bacterial beta-galactosidase (beta-gal) gene and introduced it into human fibrosarcoma (HT1080) cells, which retain wild-type p53 function. The resultant clones induce beta-gal protein after X-irradiation, and the induction kinetics were similar to those of p21(WAF1/CIP1) protein. More than 90% of the cells were stained blue when the cells were incubated with X-gal 4 h after 6 Gy of X-rays, whereas very few control cells were beta-gal positive. The primary colonies formed after 6 Gy of X-rays were collected, and they were subjected to secondary colony formation. We observed that a significant number of surviving colonies contained beta-gal-positive cells, suggesting that delayed activation of p53 occurred in the progeny of irradiated cells. We also found higher frequency of phosphorylation of p53, NBS1, and CHK2/Cds1 in the progeny of surviving cells. Furthermore, foci formation of phosphorylated histone H2AX was detected in the progeny of surviving cells. These findings provide the possibility that the observed instability results from these DNA breaks, i.e., the breaks lead to delayed chromosome rearrangements, delayed cell death, and so forth, many generations after irradiation and that activation of p53 function may eliminate cells that have potentially accumulated genomic alterations.
Suzuki
Keiji
K
Laboratory of Radiation and Life Science, School of Pharmaceutical Sciences, Nagasaki University, Nagasaki 852-8521, Japan. kzsuzuki@net.nagasaki-u.ac.jp
Yokoyama
Satoshi
S
Waseda
Satomi
S
Kodama
Seiji
S
Watanabe
Masami
M
D018797:Cell Cycle Proteins / Q000378:metabolism; D002470:Cell Survival / Q000528:radiation effects; D064447:Checkpoint Kinase 2; D002999:Clone Cells; D005354:Fibrosarcoma / Q000235:genetics; D005786:Gene Expression Regulation / Q000528:radiation effects; D016158:Genes, p53 / Q000528:radiation effects*; D006657:Histones / Q000378:metabolism / Q000528:radiation effects; D006801:Humans; D009687:Nuclear Proteins / Q000378:metabolism; D010766:Phosphorylation / Q000528:radiation effects; D010957:Plasmids / Q000235:genetics; D011494:Protein Kinases / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases; D011836:Radiation Tolerance / Q000235:genetics*; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D001616:beta-Galactosidase / Q000096:biosynthesis / Q000235:genetics
D018797:Cell Cycle Proteins; D006657:Histones; C112156:NBN protein, human; D009687:Nuclear Proteins; D016159:Tumor Suppressor Protein p53; D011494:Protein Kinases; D064447:Checkpoint Kinase 2; C578634:CHEK2 protein, human; D017346:Protein Serine-Threonine Kinases; D001616:beta-Galactosidase