Loss of p21WAF1/Cip1 in Gadd45-deficient keratinocytes restores DNA repair capacity

Abstract

Ultraviolet light (UV)-induced DNA damage is repaired primarily by the nucleotide excision repair (NER) pathway. Gadd45 is a multifunctional protein that regulates NER. Gadd45-deficient keratinocytes fail to repair UV-induced DNA damage, but the mechanism by which Gadd45 stimulates repair of UV-induced DNA damage is unknown. p21WAF1/Cip1 (p21) is a well-characterized downstream target of p53 that binds to Gadd45 and proliferating cell nuclear antigen (PCNA). The role of p21 in NER is somewhat controversial, however, recent studies appear to suggest that it inhibits DNA repair by inhibiting PCNA activity. Since a physical interplay exists between p21, Gadd45 and PCNA, we hypothesized that Gadd45 promoted DNA repair via p21. Initially, we examined p21 protein expression in Gadd45-deficient and proficient mice and found a higher base level of p21 protein in Gadd45-deficient keratinocytes and in most other tissues. With these results, we next speculated on the role played by p21 in Gadd45 regulated NER, by exposing keratinocytes from wild-type, single and double knockout (Gadd45 and p21) mice to UV, and measuring the responses. We confirmed that Gadd45-deficient keratinocytes were defective in UV-induced NER, but interestingly Gadd45/p21-null keratinocytes had normal NER in response to UV. Furthermore, Gadd45/p21-null keratinocytes were more resistant to UV-induced cell death than Gadd45-deficient keratinocytes. These results support the hypothesis that Gadd45 enhances NER by negatively regulating basal p21 expression in keratinocytes.