The damage of DNA by ultraviolet (UV) light from the sun is assumed to be responsible for the induction of mutations and the development of skin cancer in humans. To understand UV carcinogenesis, a more detailed knowledge of the molecular mechanisms of UV damage, repair and mutagenesis is necessary. In this competing continuation application, we will address the following issues:
In Specific Aim I, we will characterize the DNA damaging and mutagenic properties of UVA irradiation, a component of the solar spectrum that has been linked to melanoma.
In Specific Aim II, we will try to demonstrate a molecular link between sunlight exposure and melanoma. We will attempt to identify a UVB signature in the human p16 gene of melanoma patients and a UVA signature in the BRAF gene.
In Specific Aim III, we will test the hypothesis that specific lesion-tolerant DNA polymerases and additional factors are involved in UV mutagenesis. We will determine the in vivo roles of DNA polymerases eta, iota, kappa, beta, RAD3/RAD7 and REV1, as well as other proteins in UV mutagenesis. These studies will employ siRNA technology to knock down specific proteins involved in UV mutagenesis. The experiments will use supF plasmids containing CPDs or (6-4) photoproducts, which are replicated in cells treated with siRNA against the same polymerases. These studies will be complemented by ones using mouse embryonic fibroblasts carrying mutation reporter transgenes and derivatives of these cells expressing photoproduct-specific photolyases and specific siRNAs.
In Specific Aim I V we will test the hypothesis that 5-methylcytosine plays a crucial role in UV mutagenesis in mammalian cells. We will study the effects of 5-methylcytosine and 5-methylcytosinebinding proteins (mCpG-binding proteins, MeCP2, MBD2, MBD1) on UV damage induction and repair in vitro, on the deamination of 5-methylcytosine within cyclobutane pyrimidine dimers, and on mutagenesis in a CpG-methylated mutation reporter gene. Altogether, these studies should increase our understanding of the mechanisms of UV-induced DNA damage, its repair and the mechanisms of mutagenesis that lead to alterations in genes that are implicated in nonmelanoma and melanoma skin cancers.
|Pfeifer, Gerd P; Besaratinia, Ahmad (2012) UV wavelength-dependent DNA damage and human non-melanoma and melanoma skin cancer. Photochem Photobiol Sci 11:90-7|
|Lahtz, Christoph; Pfeifer, Gerd P (2011) Epigenetic changes of DNA repair genes in cancer. J Mol Cell Biol 3:51-8|
|Besaratinia, Ahmad; Yoon, Jae-In; Schroeder, Christi et al. (2011) Wavelength dependence of ultraviolet radiation-induced DNA damage as determined by laser irradiation suggests that cyclobutane pyrimidine dimers are the principal DNA lesions produced by terrestrial sunlight. FASEB J 25:3079-91|
|Hendriks, Giel; Calleja, Fabienne; Besaratinia, Ahmad et al. (2010) Transcription-dependent cytosine deamination is a novel mechanism in ultraviolet light-induced mutagenesis. Curr Biol 20:170-5|
|Besaratinia, Ahmad; Pfeifer, Gerd P (2009) DNA-lesion mapping in mammalian cells. Methods 48:35-9|
|Besaratinia, Ahmad; Kim, Sang-In; Pfeifer, Gerd P (2008) Rapid repair of UVA-induced oxidized purines and persistence of UVB-induced dipyrimidine lesions determine the mutagenicity of sunlight in mouse cells. FASEB J 22:2379-92|
|Jin, Seung-Gi; Tsark, Walter; Szabo, Piroska E et al. (2008) Haploid male germ cell- and oocyte-specific Mbd3l1 and Mbd3l2 genes are dispensable for early development, fertility, and zygotic DNA demethylation in the mouse. Dev Dyn 237:3435-43|
|Besaratinia, Ahmad; Pfeifer, Gerd P (2008) Sunlight ultraviolet irradiation and BRAF V600 mutagenesis in human melanoma. Hum Mutat 29:983-91|
|Lee, Dong-Hyun; Pfeifer, Gerd P (2008) Translesion synthesis of 7,8-dihydro-8-oxo-2'-deoxyguanosine by DNA polymerase eta in vivo. Mutat Res 641:19-26|
|Kim, Sang-in; Pfeifer, Gerd P; Besaratinia, Ahmad (2007) Mutagenicity of ultraviolet A radiation in the lacI transgene in Big Blue mouse embryonic fibroblasts. Mutat Res 617:71-8|
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