Differential regulation of the p21/WAF-1 and mdm2 genes after high-dose UV irradiation: p53-dependent and p53-independent regulation of the mdm2 gene.
BACKGROUND
DNA damage in mammalian cells stabilizes the p53 protein which then functions as a cell cycle checkpoint by leading to growth arrest or apoptosis. p53 is a transcription factor and positively regulates the expression of the p21/WAF-1 gene and the mdm2 gene. After high-dose UV irradiation, p53 increases the expression of the p21/WAF-1 gene immediately (2 to 5 hours after irradiation) while the induction of the mdm2 gene is delayed (8 to 12 hours after irradiation). Experiments presented here explore this differential expression of two different p53-regulated genes.


METHODS
IP-Western (protein) and Northern (mRNA) blot experiments are used to follow mdm2 and p21/WAF-1 expression in primary rat or mouse cells after a low-dose (4 J/m2) or a high-dose (20 J/M2) of UV irradiation. Northern blot and nuclear run-on experiments are employed to study mRNA stability as well as transcription rates of selected genes.


RESULTS
After high-dose UV irradiation, p53 is rapidly stabilized and the expression of p21/WAF1 is immediately increased. By contrast, both protein and mRNA levels of mdm2 first decrease in a p53-independent manner, and later increase in a p53-dependent manner. The initial decline of mdm2 expression following high-dose UV irradiation is UV-dosage dependent and regulated at the level of transcription.


CONCLUSIONS
p53 regulates two genes, p21/WAF1 (blocks cell cycle progression) and mdm2 (reverses p53 activity), that mediate opposite actions. This process is regulated in a temporal fashion after high-dose UV irradiation, so that cell cycle progression can be halted while DNA repair continues prior to reversal of p53-mediated arrest by mdm2.
Wu
L
L
Department of Molecular Biology, Princeton University, NJ 08544-1014, USA.
Levine
A J
AJ
D000818:Animals; D002460:Cell Line; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins / Q000235:genetics* / Q000378:metabolism / Q000528:radiation effects*; D004307:Dose-Response Relationship, Radiation; D015536:Down-Regulation / Q000235:genetics / Q000528:radiation effects; D005347:Fibroblasts / Q000378:metabolism / Q000528:radiation effects; D005786:Gene Expression Regulation / Q000528:radiation effects*; D051379:Mice; D009687:Nuclear Proteins; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism / Q000528:radiation effects; D051736:Proto-Oncogene Proteins c-mdm2; D011519:Proto-Oncogenes / Q000528:radiation effects*; D012333:RNA, Messenger / Q000528:radiation effects; D051381:Rats; D014158:Transcription, Genetic / Q000528:radiation effects; D016159:Tumor Suppressor Protein p53 / Q000502:physiology* / Q000528:radiation effects; D014466:Ultraviolet Rays
C493957:Cdkn1a protein, mouse; C493958:Cdkn1a protein, rat; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D009687:Nuclear Proteins; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger; D016159:Tumor Suppressor Protein p53; C497207:Mdm2 protein, mouse; C497208:Mdm2 protein, rat; D051736:Proto-Oncogene Proteins c-mdm2