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Growth-dependent heterogeneity in the DNA damage response in Escherichia coli

ABSTRACT: Two datasets have been collected in order to quantify the effect of varying the growth rate (modulated by changing the nutrient quality) on the SOS response in Escherichia coli K-12 MG1655. The first dataset consists of snapshot images taken for cells mounted on agar-pads after being grown in different conditions. The second dataset consists of timelapse images of cells grown in the mother machine microfluidic device under different conditions. Apart from the wild type (WT) strain, several derivative strains were also used. Of these, the "2-pal" strain was focussed on as it constitutes a reliable and controlled source of chronic, replication-dependent DNA damage. Data was collected for three growth conditions separated by growth medium corresponding to 'slow', 'medium', and 'fast' growth rates. Some datasets additionally include ciprofloxacin exposure as an alternative DNA-damaging agent. Images were captured in brightfield, mCherry (red), and mGFP (green) fluorescence channels. The mKate2 fluorescent marker is linked to constitutive gene expression (Ptet01-mKate2) and primarily used for cell segmentation; while mGFP fluorescent marker is linked to SOS regulon expression (PsulA-mGFP).

ORGANISM(S): Escherichia coli

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PROVIDER: S-BSST833 | bioimages |

REPOSITORIES: bioimages

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Project description:UNLABELLED:BK polyomavirus (BKPyV) reactivation is associated with severe human disease in kidney and bone marrow transplant patients. The interplay between viral and host factors that regulates the productive infection process remains poorly understood. We have previously reported that the cellular DNA damage response (DDR) is activated upon lytic BKPyV infection and that its activation is required for optimal viral replication in primary kidney epithelial cells. In this report, we set out to determine what viral components are responsible for activating the two major phosphatidylinositol 3-kinase-like kinases (PI3KKs) involved in the DDR: ataxia telangiectasia mutated (ATM) kinase and ATM and Rad3-related (ATR) kinase. Using a combination of UV treatment, lentivirus transduction, and mutant virus infection experiments, our results demonstrate that neither the input virus nor the expression of large T antigen (TAg) alone is sufficient to trigger the activation of ATM or ATR in our primary culture model. Instead, our data suggest that the activation of both the ATM- and ATR-mediated DDR pathways is linked to viral DNA replication. Intriguingly, a TAg mutant virus that is unable to activate the DDR causes substantial host DNA damage. Our study provides insight into how DDRs are activated by polyomaviruses in primary cells with intact cell cycle checkpoints and how the activation might be linked to the maintenance of host genome stability. IMPORTANCE:Polyomaviruses are opportunistic pathogens that are associated with several human diseases under immunosuppressed conditions. BK polyomavirus (BKPyV) affects mostly kidney and bone marrow transplant patients. The detailed replication mechanism of these viruses remains to be determined. We have previously reported that BKPyV activates the host DNA damage response (DDR), a response normally used by the host cell to combat genotoxic stress, to aid its own replication. In this study, we identified that the trigger for DDR activation is viral replication. Furthermore, we show that the virus is able to cause host DNA damage in the absence of viral replication and DDR activation. These results suggest an intricate relationship between viral replication, DDR activation, and host genome instability.

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Growth-dependent heterogeneity in the DNA damage response in Escherichia coli

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