Oxidative Stress and Replication-Independent DNA Breakage Induced by Arsenic in Saccharomyces cerevisiae

Abstract

Arsenic is a well-established human carcinogen of poorly understood mechanism of genotoxicity. It is generally accepted that arsenic acts indirectly by generating oxidative DNA damage that can be converted to replication-dependent DNA double-strand breaks (DSBs), as well as by interfering with DNA repair pathways and DNA methylation. Here we show that in budding yeast arsenic also causes replication and transcription-independent DSBs in all phases of the cell cycle, suggesting a direct genotoxic mode of arsenic action. This is accompanied by DNA damage checkpoint activation resulting in cell cycle delays in S and G2/M phases in wild type cells. In G1 phase, arsenic activates DNA damage response only in the absence of the Yku70–Yku80 complex which normally binds to DNA ends and inhibits resection of DSBs. This strongly indicates that DSBs are produced by arsenic in G1 but DNA ends are protected by Yku70–Yku80 and thus invisible for the checkpoint response. Arsenic-induced DSBs are processed by homologous recombination (HR), as shown by Rfa1 and Rad52 nuclear foci formation and requirement of HR proteins for cell survival during arsenic exposure. We show further that arsenic greatly sensitizes yeast to phleomycin as simultaneous treatment results in profound accumulation of DSBs. Importantly, we observed a similar response in fission yeast Schizosaccharomyces pombe, suggesting that the mechanisms of As(III) genotoxicity may be conserved in other organisms.

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“…500 nM As þ3 ) result in oxidative stress that is associated with more DNA damage and double strand breaks. The findings are in agreement with those obtained by other groups (Litwin et al, 2013;Qin et al, 2012). There is also evidence showing that PARP contributes to XPA repair of double strand breaks (King et al, 2012).…”

Global Fitness Profiling Identifies Arsenic and Cadmium Tolerance Mechanisms in Fission Yeast

“…Thus arsenite concentrations that modestly inhibit growth of wild-type cells do not cause levels of DNA damage that will lead to cell death in the absence of core DNA damage responses. However, we note that the arsenite concentrations used in our screen (100–200 μΜ NaAsO 2 ) were much lower than those used by Litwin and colleagues (500–4000 μΜ NaAsO 2 ) in their studies with S. cerevisiae and S. pombe ( Litwin et al 2013 ). Therefore, it appears that DNA damage responses can be important for coping with very high concentrations of arsenite in fission yeast, or tolerating simultaneous exposure to arsenite and other toxins, but at lower arsenite concentrations other cellular activities are much more important, particularly those involved in arsenite detoxification.…”

“…Arsenic has carcinogenic properties yet it remains unclear whether it causes genome instability, and if so, whether arsenic directly or indirectly creates DNA damage. A recent study with S. cerevisiae indicated that arsenite causes replication and transcription-independent double-strand breaks (DSBs) in all phases of the cell cycle, triggering DNA damage checkpoint responses and homology dependent repair (HDR) requiring members of the Rad52 epistasis group ( Litwin et al 2013 ). These studies further indicated that arsenite has similar effects in fission yeast.…”

Construction of Novel Saccharomyces cerevisiae Strains for Bioethanol Active Dry Yeast (ADY) Production

“…ROS is injurious to biomolecules such as DNA and protein and can induce lipid peroxidation in yeast cells [37,38]. Under regular conditions, low levels of MDA resulting from lipid peroxidation formed in both ZTW1 and Z3-86.…”