Abstract
BackgroundPrevious reports showed that mutagenesis in nutrient-limiting conditions is dependent on Mfd in Bacillus subtilis. Mfd initiates one type of transcription-coupled repair (TCR); this type of repair is known to target bulky lesions, like those associated with UV exposure. Interestingly, the roles of Mfd in repair of oxidative-promoted DNA damage and regulation of transcription differ. Here, we used a genetic approach to test whether Mfd protected B. subtilis from exposure to two different oxidants.ResultsWild-type cells survived tert-butyl hydroperoxide (t-BHP) exposure significantly better than Mfd-deficient cells. This protective effect was independent of UvrA, a component of the canonical TCR/nucleotide excision repair (NER) pathway. Further, our results suggest that Mfd and MutY, a DNA glycosylase that processes 8-oxoG DNA mismatches, work together to protect cells from lesions generated by oxidative damage. We also tested the role of Mfd in mutagenesis in starved cells exposed to t-BHP. In conditions of oxidative stress, Mfd and MutY may work together in the formation of mutations. Unexpectedly, Mfd increased survival when cells were exposed to the protein oxidant diamide. Under this type of oxidative stress, cells survival was not affected by MutY or UvrA.ConclusionsThese results are significant because they show that Mfd mediates error-prone repair of DNA and protects cells against oxidation of proteins by affecting gene expression; Mfd deficiency resulted in increased gene expression of the OhrR repressor which controls the cellular response to organic peroxide exposure. These observations point to Mfd functioning beyond a DNA repair factor in cells experiencing oxidative stress.
Highlights
Previous reports showed that mutagenesis in nutrient-limiting conditions is dependent on Mfd in Bacillus subtilis
Our research indicates that Mfd promotes repair of oxidative damage in B. subtilis and suggests that oxidative damage is a precursor to stationary-phase mutagenesis
To determine if Mfd protects against oxidative damage in stationary-phase B. subtilis cells, we subjected YB955 (Mfd+) and YB9801 (Mfd−) to varying levels of oxidative stress by exposing cells to the oxidant tert-butyl hydroperoxide (t-BHP). t-BHP is an organic peroxide that is metabolized by cells to form multiple radicals which damage several cellular components, such as lipids and DNA [6, 24]
Summary
Previous reports showed that mutagenesis in nutrient-limiting conditions is dependent on Mfd in Bacillus subtilis. In B. subtilis, a bacterial model for cell growth and differentiation, the cytotoxic effects of oxidative stress are countered by regulons under the control of several transcription factors (e.g., PerR, OhrR, Spx, YodB, SigB, and MgsR); some of them are activated by sensing the redox state of the cell [6]. These regulons have been elucidated by experiments examining how cells respond to exposure to oxidants that include hydrogen peroxide, paraquat, organic peroxides, and diamide [6]. Genes that are transcriptionally active, directly or indirectly [7, 8], in these conditions code for factors that detoxify ROS (e.g., kat and sod, ahpC, ahpF), confer resistance to heavy metals (e.g., arsB), prevent protein misfolding (e.g., groES, dnaK), Martin et al BMC Microbiology (2019) 19:26 and maintain thiol-disulfide homeostasis in the cell (trx, cys, bsh) [9, 10]
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