Mechanisms of UVA-mutagenesis remain a matter of debate. Earlier described higher rates of mutation formation per pyrimidine dimer with UVA than with UVB and other evidence suggested that a non-pyrimidine dimer-type of DNA damage contributes more to UVA- than to UVB-mutagenesis. However, more recently published data on the spectra of UVA-induced mutations in primary human skin cells and in mice suggest that pyrimidine dimers are the most common type of DNA damage-inducing mutations not only with UVB, but also with UVA. As this rebuts a prominent role of non-dimer type of DNA damage in UVA-mutagenesis, we hypothesized that the higher mutation rate at UVA-induced pyrimidine dimers, as compared to UVB-induced ones, is caused by differences in the way UVA- and UVB-exposed cells process DNA damage. Therefore, we here compared cell cycle regulation, DNA repair, and apoptosis in primary human fibroblasts following UVB- and UVA-irradiation, using the same physiologic and roughly equimutagenic doses (100-300 J m(-2) UVB, 100-300 kJ m(-2) UVA) we have used previously for mutagenesis experiments with the same type of cells. ELISAs for the detection of pyrimidine dimers confirmed that much fewer dimers were formed with these doses of UVA, as compared to UVB. We found that cell cycle arrests (intra-S, G1/S, G2/M), mediated at least in part by activation of p53 and p95, are much more prominent and long-lasting with UVB than with UVA. In contrast, no prominent differences were found between UVA and UVB for other anti-mutagenic cellular responses (DNA repair, apoptosis). Our data suggest that less effective anti-mutagenic cellular responses, in particular different and shorter-lived cell cycle arrests, render pyrimidine dimers induced by UVA more mutagenic than pyrimidine dimers induced by UVB.

Recombination repair plays an important role in the processing of DNA double-strand breaks (DSB) and DNA cross-links, and has been suggested to be mediated by the activation of the Fanconi anemia (FA)/BRCA pathway. Unlike DNA damage generated by ionizing radiation or DNA crosslinking, UV light-induced DNA damage is not commonly thought to require recombination for processing, as UV light does not directly induce DSBs or DNA cross-links. To elucidate the role of recombination repair in the cellular response to UV, we studied the FA/BRCA pathway in primary skin cells exposed to solar-simulated light. UV-induced monoubiquitination of the FANCD2 protein and formation of FANCD2 nuclear foci confirmed the activation of the pathway by UV light. This was only observed when cells were irradiated during S phase and was not caused by directly UV-induced DSBs. UV-exposed cells did not exhibit FANCD2 nuclear foci once they entered mitosis or when growth-arrested. In addition, UV-induced nuclear foci of the recombination proteins, RAD51 and BRCA1, colocalized with FANCD2 foci. We suggest that in response to UV light, when nucleotide excision repair failed to repair, or when translesional DNA synthesis failed to bypass UV-induced DNA photoproducts, the FA/BRCA pathway mediates the recombination repair of replication forks stalled at DNA photoproducts as a third line of defense. (Cancer Res 2006; 66(23): 11140-7)

Longwave UVA is an independent class I carcinogen. A complete understanding of UVA-induced DNA damage and how this damage is processed in skin cells is therefore of utmost importance. A particular question that has remained contentious is whether UVA induces DNA double-strand breaks (DSBs), either directly or through processing of other types of DNA damage, such as recombination repair of replication forks stalled at DNA photoproducts. We therefore studied activation of the recombination repair pathway by solar available doses of UVA and assessed formation of DNA DSBs in primary skin fibroblasts. We found that, unlike ionizing radiation or UVB, UVA does not activate the Fanconi anemia/BRCA DNA damage response pathway or the "recombinase" RAD51 in primary skin fibroblasts. The fact that this pathway mediates recombination repair of DNA DSBs suggests that DNA DSBs are not formed by UVA. This is further supported by findings that UVA did not induce DNA DSBs, as assayed by neutral single-cell electrophoresis or by formation of γ-H2AX nuclear foci, considered the most sensitive assay for DNA DSBs. The lack of sufficient evidence for formation of DNA DSBs underlines the pivotal role of UVA-induced DNA photoproducts in UVA mutagenesis and carcinogenesis.