科研之友
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摘要
Arsenic is an environmental contaminant with known cytotoxic and carcinogenic properties, but the cellular mechanisms of its action are not fully known. As retrotransposition consists a potent mutagenic factor affecting genome stability, we investigated the effect of arsenic on retrotransposition of an enhanced green fluorescent protein (EGFP)tagged nonautonomous long terminal repeat (LTR)-retrotransposon viral-like 30 (VL30) in a mouse NIH3T3 cell culture-retrotransposition assay. Flow cytometry analysis of assay cells treated with 2.520M sodium arsenite revealed induction of retrotransposition events in a dose- and time-dependent manner, which was further confirmed as genomic integrations by PCR analysis and appearance of EGFP-positive cells by UV microscopy. Specifically, 20M sodium arsenite strongly induced the VL30 retrotransposition frequency, which was similar to 90,000-fold higher than the natural one and also VL30 RNA expression was similar to 6.6-fold. Inhibition of the activity of endogenous reverse transcriptases by efavirenz at 15M or nevirapine at 375M suppressed the arsenite-induced VL30 retrotransposition by 71.16 or 79.88%, respectively. In addition, the antioxidant N-acetyl-cysteine reduced the level of arsenite-induced retrotransposition, which correlated with the rescue of arsenite-induced G2/M cell cycle arrest and cell toxicity. Treatment of assay cells ectopically overexpressing the human heat-shock protein 70 (Hsp70) with 15M sodium arsenite resulted in an additional similar to 4.5-fold induction of retrotransposition compared with normal assay cells, whereas treatment with 20M produced a massive cell death. Our results show for the first time that arsenic both as an oxidative and heat-shock mimicking agent is a potent inducer of VL30 retrotransposition in mouse cells. The impact of arsenic-induced retrotransposition, as a cellular response, on contribution to or explanation of the arsenic-associated toxicity and carcinogenicity is discussed.
- 出版日期2013-8
