Adduct formation and repair, and translesion DNA synthesis across the adducts in human cells exposed to 3-nitrobenzanthrone

作者：Kawanishi Masanobu; Fujikawa Yoshihiro; Ishii Hiroshi; Nishida Hiroshi; Higashigaki Yuka; Kanno Takaharu; Matsuda Tomonari; Takamura Enya Takeji; Yagi Takashi^*

来源：Mutation Research: Genetic Toxicology and Environmental Mutagenesis , 2013, 753(2): 93-100.

DOI：10.1016/j.mrgentox.2013.03.005

• 3-Nitrobenzanthrone
• DNA adduct
• DNA repair
• Translesion DNA synthesis
• Mutation

3-Nitrobenzanthrone (3-nitro-7H-benz[d,e]anthracen-7-one, 3-NBA) is a potent environmental mutagen that is found in diesel exhaust fumes and airborne particulates. It is known to produce several DNA adducts, including three major adducts N-(2%26apos;-deoxyguanosin-8-yl)-3-aminobenzanthrone (dG-C8-N-ABA), 2-(2%26apos;-deoxyadenosin-N-6-yl)-3-aminobenzanthrone (dA-N-6-C2-ABA), and 2-(2%26apos;-deoxyguanosin-N-2-yl)-3-aminobenzanthrone (dG-N-2-C2-ABA) in mammalian cells. In the present study, we measured the quantity of the formation and subsequent reduction of these adducts in human hepatoma HepG2 cells that had been treated with 3-NBA using LC-MS/MS analysis. As a result, dG-C8-N-ABA and dG-N-2-C2-ABA were identified as major adducts in the HepG2 cells, and dA-N-6-C2-ABA was found to be a minor adduct. Treatment with 1 mu g/mL 3-NBA for 24h induced the formation of 2835 +/- 1509 dG-C8-N-ABA and 3373 +/- 1173 dG-N-2-C2-ABA per 10(7) dG and 877 +/- 330 dA-N-6-C2-ABA per 10(7) dA in the cells. The cellular DNA repair system removed the dG-C8-N-ABA and dA-N-6-C2-ABA adducts more efficiently than the dG-N-2-C2-ABA adducts. After a 24-h repair period, 86.4 +/- 11.1% of the dG-N-2-C2-ABA adducts remained, whereas only 51.7 +/- 2.7% of the dG-C8-N-ABA adducts and 37.8 +/- 1.7% of the dA-N-6-C2-ABA adducts were present in the cells. We also evaluated the efficiency of bypasses across these three adducts and their mutagenic potency by introducing site-specific mono-modified plasmids into human cells. This translesion DNA synthesis (TLS) assay showed that dG-C8-N-ABA blocked DNA replication markedly (its replication frequency was 16.9 +/- 2.7%), while the replication arrests induced by dG-N-2-C2-ABA and dA-N-6-C2-ABA were more moderate (their replication frequencies were 33.3 +/- 6.2% and 43.1 +/- 7.5%, respectively). Mutagenic TLS was observed more frequently in replication across dG-C8-N-ABA (30.6%) than in replication across dG-N-2-C2-ABA (12.1%) or dA-N-6-C2-ABA (12.1%). These findings provide important insights into the molecular mechanism of 3-NBA-mutagenesis.

• 出版日期2013-5-15