The Role of Mfd in Stationary-Phase Oxidative Damage Repair in Bacillus subtilis

Since the 1950’s it has been shown that bacterial cells accumulate mutations even in non- dividing conditions, but how this type of mutation occurs is still highly debated. In Bacillus subtilis, Mfd, a precursor of the nucleotide excision repair (NER) system, mediates the formation of mutations in stationary-phase or non-replicating cells. In growing cells, Mfd recruits repair when RNA polymerase is stalled during transcription; it then recruits proteins from NER to repair damage. Here, we examine the hypothesis that Mfd mediates the formation of mutations by interacting with cellular components that repair reactive oxygen species (ROS), a natural byproduct of bacteria cell respiration. Utilizing two oxidants, we tested the hypothesis that Mfd protects cell viability and mutation development after exposure to ROS in stationary-phase. Our data showed that Mfd mitigated damage caused by reactive oxygen species and that such effect is independent of the NER system. Also, Mfd, MutY and reactive oxygen species mediated the formation of mutations in stationary-phase cells

Mitsui-7, heat-treated, and nitrogen-doped multi-walled carbon nanotubes elicit genotoxicity in human lung epithelial cells.

Background: The unique physicochemical properties of multi-walled carbon nanotubes (MWCNT) have led to many industrial applications. Due to their low density and small size, MWCNT are easily aerosolized in the workplace making respiratory exposures likely in workers. The International Agency for Research on Cancer designated the pristine Mitsui-7 MWCNT (MWCNT-7) as a Group 2B carcinogen, but there was insufficient data to classify all other MWCNT. Previously, MWCNT exposed to high temperature (MWCNT-HT) or synthesized with nitrogen (MWCNT-ND) have been found to elicit attenuated toxicity; however, their genotoxic and carcinogenic potential are not known. Our aim was to measure the genotoxicity of MWCNT-7 compared to these two physicochemically-altered MWCNTs in human lung epithelial cells (BEAS-2B & SAEC). Results: Dose-dependent partitioning of individual nanotubes in the cell nuclei was observed for each MWCNT material and was greatest for MWCNT-7. Exposure to each MWCNT led to significantly increased mitotic aberrations with multi- and monopolar spindle morphologies and fragmented centrosomes. Quantitative analysis of the spindle pole demonstrated significantly increased centrosome fragmentation from 0.024-2.4 [mu]g/mL of each MWCNT. Significant aneuploidy was measured in a dose-response from each MWCNT-7, HT, and ND; the highest dose of 24 [mu]g/mL produced 67, 61, and 55%, respectively. Chromosome analysis demonstrated significantly increased centromere fragmentation and translocations from each MWCNT at each dose. Following 24 h of exposure to MWCNT-7, ND and/or HT in BEAS-2B a significant arrest in the G1/S phase in the cell cycle occurred, whereas the MWCNT-ND also induced a G2 arrest. Primary SAEC exposed for 24 h to each MWCNT elicited a significantly greater arrest in the G1 and G2 phases. However, SAEC arrested in the G1/S phase after 72 h of exposure. Lastly, a significant increase in clonal growth was observed one month after exposure to 0.024 [mu]g/mL MWCNT-HT & ND. Conclusions: Although MWCNT-HT & ND cause a lower incidence of genotoxicity, all three MWCNTs cause the same type of mitotic and chromosomal disruptions. Chromosomal fragmentation and translocations have not been observed with other nanomaterials. Because in vitro genotoxicity is correlated with in vivo genotoxic response, these studies in primary human lung cells may predict the genotoxic potency in exposed human populations