DIFFERENTIATION-DEPENDENT P53 REGULATION OF NUCLEOTIDE EXCISION-REPAIR IN KERATINOCYTES

The role of the tumor suppressor p53 in repair of ultraviolet light (U V)-induced DNA damage was evaluated using a host-cell reactivation (HC R) assay. HCR determines a cell's ability to repair W-damaged DNA thro ugh reactivation of a transfected CAT reporter plasmid. Most UV damage is removed through nucleotide excision repair (NER). Primary murine k eratinocytes isolated from p53-deficient and wild-type p53 mice were u sed in the HCR assay The NER was reduced in p53(-/-) keratinocytes as compared with p53(-/-) keratinocytes. The reduced DNA repair in p53(-/ -) mice was confirmed with a radioimmunoassay comparing cyclobutane di mers (CPDs) and (6-4) photoproducts in P53(+/+) and p53(-/-) keratinoc ytes after the cells were exposed to UV irradiation. Our results demon strate that wildtype p53 play's a significant role in regulating NER. Furthermore, as there is evidence that p53 protein levels decrease aft er keratinocytes become differentiated, we sought to determine whether p53 plays a role in NER in differentiated keratinocytes. Differentiat ion of the keratinocytes by increasing the Ca2+ concentration in the c ulture media resulted in a marked reduction in NER equally in both p53 (+/+) and p53(-/-) groups. This finding suggests that reduced DNA repa ir after differentiation is p53 independent. A similar reduction in HC R was confirmed in differentiated human keratinocytes. These data, tak en together, indicate that P53 or p53-regulated proteins enhance NER i n basal undifferentiated keratinocytes but not in differentiated cells . As non-melanoma skin cancers originate from the basal keratinocytes, our findings suggest that loss of p53 may contribute to the pathogene sis of this common skin cancer.