Mutational Analysis of the C8-Guanine Adduct of the Environmental Carcinogen 3-Nitrobenzanthrone in Human Cells: Critical Roles of DNA Polymerases 畏 and 魏 and Rev1 in Error-Prone Translesion Synthesis

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• 作者：Paritosh Pande ; Chanchal K. Malik ; Arindam Bose ; Vijay P. Jasti ; Ashis K. Basu
• 刊名：Biochemistry
• 出版年：2014
• 出版时间：August 19, 2014
• 年：2014
• 卷：53
• 期：32
• 页码：5323-5331
• 全文大小：427K
• ISSN：1520-4995

文摘

3-Nitrobenzanthrone (3-NBA), a potent mutagen and suspected human carcinogen, is a common environmental pollutant. The genotoxicity of 3-NBA has been associated with its ability to form DNA adducts, including N-(2鈥?deoxyguanosin-8-yl)-3-aminobenzanthrone (C8-dG-ABA). To investigate the molecular mechanism of C8-dG-ABA mutagenesis in human cells, we have replicated a plasmid containing a single C8-dG-ABA in human embryonic kidney 293T (HEK293T) cells, which yielded 14% mutant progeny. The major types of mutations induced by C8-dG-ABA were G 鈫?T > G 鈫?A > G 鈫?C. siRNA knockdown of the translesion synthesis (TLS) DNA polymerases (pols) in HEK293T cells indicated that pol 畏, pol 魏, pol 喂, pol 味, and Rev1 each have a role in replication across this adduct. The extent of TLS was reduced with each pol knockdown, but the largest decrease (of 55% reduction) in the level of TLS occurred in cells with knockdown of pol 味. Pol 畏 and pol 魏 were considered the major contributors of the mutagenic TLS, because the mutation frequency (MF) decreased by 70%, when these pols were simultaneously knocked down. Rev1 also is important for mutagenesis, as reflected by the 60% reduction in MF upon Rev1 knockdown, but it probably plays a noncatalytic role by physically interacting with the other two Y-family pols. In contrast, pol 味 appeared to be involved in the error-free bypass of the lesion, because MF increased by 60% in pol 味 knockdown cells. These results provide important mechanistic insight into the bypass of the C8-dG-ABA adduct.