P53-MEDIATED TRANSCRIPTIONAL ACTIVITY INCREASES IN DIFFERENTIATING EPIDERMAL-KERATINOCYTES IN ASSOCIATION WITH DECREASED P53 PROTEIN

The regulation of p53 protein synthesis and p53-mediated gene transact ivation mere evaluated in cultured mouse keratinocytes maintained as b asal cells or induced to differentiate by Ca2+>0.1 mM. p53 protein hal f-life, p53 protein synthesis and the level of p53 mRNA decreased duri ng terminal differentiation, as detected by immunoprecipitation ,vith a panel of p53-specific antibodies and Northern blotting. Thus differe ntiating keratinocytes have lower levels of p53 protein. This decline is not observed following growth arrest alone, or in papilloma cell li nes which do not terminally differentiate in response to Ca2+. In cont rast, the ability of endogenous p53 to transactivate transcription fro m the PGI, CAT plasmid increased during differentiation in vitro. This change in activity cannot be explained by changes in p53 conformation or nuclear localization. Consistent with these findings, mRNA for the p53-mediated genes WAF1 and mdm-2 increased with Ca2+-induced differe ntiation in a time dependent manner, suggesting activation of p53 cont ributes to the differentiated phenotype. However, p53-null mice exhibi t histologically normal skin and epidermal keratinocytes from these mi ce express the appropriate markers of differentiation and suppression of DNA synthesis in vitro when the [Ca2+] is >0.1 mM. The observation that proliferating cells have higher levels of p53 protein which is le ss active for its function than differentiated cell types could have a consequence for the selection of p53 gene mutations during carcinogen esis, depending upon the stage of differentiation of the tumor cell ty pe.