The overall aim of this project is to apply new knowledge of fundamental mechanisms of mutagenesis to directly test the somatic mutation hypothesis of cancer, and to translate this knowledge into a clinically useful regimen to reduce the incidence of sunlight-associated skin cancer. Mutations in DNA are generally considered to have an etiologic role in the development of cancer, but until recently it has not been possible to directly examine the relative contribution of mutagenesis and epigenetic alterations in cellular physiology induced by carcinogen exposure. We hypothesize that mutations induced by ultraviolet light (UV) in the DNA of actively dividing skin cells are causally involved in skin carcinogenesis, and reducing the frequency of such mutations will reduce the risk of skin cancer induced by UV. There is now abundant evidence that virtually all mutations induced by UV in eukaryotic cells are dependent on the gene products encoded by the REV1 and REV3 genes. We showed further that targeting the mRNA of one of these genes (REV1) with gene-specific ribozymes virtually abolishes UV mutagenesis. To test our hypothesis, we propose to expose newly-developed REV3-depleted transgenic mice to UV. We anticipate that these mice will have a greatly reduced risk of developing skin cancer compared with control animals. Further, we propose to translate our data into a practical method to prevent UV-induced skin cancer. We will develop and optimize gene-specific ribozymes expressed in cultured cells using several different systems to target them to the cytoplasm or to the nucleus. Then, we propose to deliver such ribozyme expression vectors into mouse skin cells. These ribozymes will specifically target REV1 mRNA, a protein that may coordinate the activity of several proteins that are required for the mutagenic replication of UVdamaged DNA. The mice and the appropriate controls will be exposed to UV protocols that are known to induce skin cancer. Our hypothesis predicts that reducing the level of this protein will reduce the mutagenic response of skin cells to UV, and will result in a greatly reduced incidence of skin cancer. If so, these data will provide the experimental rationale to develop similar strategies to reduce the incidence of skin cancer in humans. Most importantly, such a strategy should be successful as a preventive measure after UV exposure and prior to appearance of disease symptoms.
