Acclimation and repair of DNA damage in recombinant bioluminescent Escherichia coli

Abstract

Aims: The aim of this study is to understand different adaptive responses in bacteria caused by three different mutagens, namely, an intercalating agent, an alkylating agent and a hydroxylating agent, and the repair systems according to the type of DNA damage, that is, DNA cross-linking and delayed DNA synthesis, alkylation and hydroxylation of DNA. A recombinant bioluminescent Escherichia coli, DPD2794 with the recA promoter fused to luxCDABE originating from Vibrio fischeri, was used in this study. Methods and Results: The recombinant bioluminescent E. coli strain DPD2794, containing a recA promoter fused to luxCDABE from V. fischeri, was used to detect adaptive and repair responses to DNA damage caused by mitomycin C (MMC), and these responses were compared with those when the cells were induced with N-methyl-N-nitro-N-nitrosoguanidine (MNNG) and hydrogen peroxide (H 2 O 2 ). The response ratio between the induced samples and that of the controls decreased suddenly when the induced culture was used in further inductions, indicating a possible adaptive response to DNA damage. DNA damage, or the proteins produced, because of MMC addition does not appear to be completely resolved until the seventh sub-culture after the initial induction, whereas simple damage, such as the base modification caused by MNNG and H 2 O 2 , appears to be repaired rapidly as evidenced by the quick recovery of sensitivity. Conclusions: These results suggest that it takes more time to completely repair DNA damage caused by MMC, as compared with a simple repair such as that required for the damage caused by MNNG and H 2 O 2 . Therefore, repair of the damage caused by these three mutagens is controlled by different regulons, even though they all induced the recA promoter. Significance and Impact of the Study: Using a bioluminescent E. coli harbouring a recA promoter-lux fusion, it was found that different adaptive responses and repair systems for DNA damage caused by several mutagens exists in E. coli.

Construction of a nrdA::luxCDABE Fusion and Its Use in Escherichia coli as a DNA Damage Biosensor

“…To characterize strain BBT NrdA , we used four different chemicals that are known DNA mutagens [22], i.e. , nalidixic acid (NDA), mitomycin C (MMC), 1-methyl-1-nitroso-N-methylguanidine (MNNG), and 4-nitroquinoline N-oxide (4-NQO).…”

Construction of a bioreporter by heterogeneously expressing a Vibrio natriegens recA::luxCDABE fusion in Escherichia coli, and genotoxicity assessments of petrochemical-contaminated groundwater in northern China

“…24 It has been widely applied for the construction of many bioreporters in E. coli and Acinetobacter baylyi in our lab 14 and across the world. 27 The use of luxCDABE as the reporter gene simplied bioluminescence detection as no substrate needed to be added. 2,28 DH5a_lux was induced by different concentrations of mitomycin C, ranging from 0.1 nM to 1 mM, and the induction and growth kinetic curves were shown in Fig.…”

A whole-cell bioreporter assay for quantitative genotoxicity evaluation of environmental samples

“…RecA is essential in the SOS response of Escherichia coli, responsible for DNA repair/maintenance via homologous recombination (Horii et al, 1980). Therefore, the recA-based whole-cell bioreporters are widely used for measuring general toxicity, capable of detecting not only the levels but also mechanisms of DNA damage (Sørensen et al, 2006;Ron, 2007), including DNA cross-linking and delayed DNA synthesis, alkylation and hydroxylation of DNA (Min and Gu, 2003;Chen et al, 2008). As most genotoxins are inducers of the SOS response (Quillardet et al, 1982), the recA-based bioreporter assay is introduced in genotoxicity assessment of environmental samples.…”