Molecular mechanisms of DNA replication across UV-induced DNA lesions and novel approaches to suppress UV mutagenesis
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Skin cancer is the most prevalent cancer in humans, with an annual incidence of 5.4 million in the U.S. Ultra Violet (UV) irradiation generates DNA lesions that are potentially mutagenic, and accumulation of mutations eventually leads to skin cancers. UV irradiation generates two major types of DNA lesions: cyclobutane pyrimidine dimers (CPDs) and 6-4 photoproducts (6-4PPs) that can cause replication stalling. To cope with the replication-blocking lesions, cells employ a specialized damage tolerance process known as translesion synthesis (TLS), that is potentially mutagenic. The two types of UV lesions are structurally distinct and are bypassed by DNA polymerases differently. Importantly, 6-4PP is more mutagenic than CPD. A major cell cycle checkpoint kinase known as Ataxia-Telangiectasia Mutated and RAD3-related (ATR) is activated after UV. ATR is critical for the cells to survive UV damage, but could also increase the risk of mutations by increasing the survival of DNA-damaged cells. ATR inhibition augments the apoptosis of UV-damaged cells, thereby suppressing UV carcinogenesis. However, whether ATR promotes mutations in UV-damaged cells remains unclear. It is of interest to investigate whether ATR inhibition reduces mutations in cells surviving UV damage. We hypothesized that ATR is required for promoting error-prone TLS. TLS bypass of 6-4PP lesions increases mutations leading to UV-induced carcinogenesis. In this study, we analyzed the effect of ATR inhibition in TLS activity at 6-4PP lesions. We measured the percentage of unbypassed 6-4PP lesions surrounded by single-stranded DNA by using a lesion-specific antibody using flow cytometry. If ATR is facilitating 6-4PP bypass, ATR inhibition is expected to increase the number of unbypassed 6-4PP lesions. Our results showed that the percentage of unbypassed 6-4PP lesions were increased when ATR is inhibited by pharmacologic inhibition (VE-821/caffeine) compared to cells without ATR inhibition. This implies that ATR is likely involved in 6-4PP lesion bypass. The unbypassed 6-4PP lesions might eventually get repaired or progress into double-strand DNA breaks leading to apoptosis. Therefore, blocking error-prone TLS activity by ATR inhibition using VE-821 or caffeine might reduce mutation incorporation and thereby suppression of UV carcinogenesis.
- Dentistry