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Escherichia coli YafP protein modulates DNA damaging property of the nitroaromatic compounds.

Gutierrez A, Elez M, Clermont O, Denamur E, Matic I - Nucleic Acids Res. (2011)

Bottom Line: Using a murine septicaemia model, we showed that YafP activity reduced the bacterial fitness in the absence of PolIV.The YafP antimutator activity was independent of the PolIV activity.Given that YafP was annotated as a putative acetyltransferase, it could be that YafP participates in the metabolic transformation of genotoxic compounds, hence modulating the balance between their mutagenicity and cytotoxicity.

View Article: PubMed Central - PubMed

Affiliation: Faculté de Médecine Paris Descartes, Inserm U1001, Université Paris Descartes, Paris, France.

ABSTRACT
Escherichia coli SOS functions constitute a multifaceted response to DNA damage. We undertook to study the role of yafP, a SOS gene with unknown function. yafP is part of an operon also containing the dinB gene coding for DNA Polymerase IV (PolIV). Our phylogenetic analysis showed that the gene content of this operon is variable but that the dinB and the yafP genes are conserved in the majority of E. coli natural isolates. Therefore, we studied if these proteins are functionally linked. Using a murine septicaemia model, we showed that YafP activity reduced the bacterial fitness in the absence of PolIV. Similarly, YafP increased cytotoxicity of two DNA damaging nitroaromatic compounds, 4-nitroquinoline-1-oxide (NQO) and nitrofurazone, in the absence of PolIV. The fact that PolIV counterbalances YafP-induced cytotoxicity could explain why these two genes are transcriptionally linked. We also studied the involvement of YafP in genotoxic-stress induced mutagenesis and found that PolIV and YafP reduced NQO-induced mutagenicity. The YafP antimutator activity was independent of the PolIV activity. Given that YafP was annotated as a putative acetyltransferase, it could be that YafP participates in the metabolic transformation of genotoxic compounds, hence modulating the balance between their mutagenicity and cytotoxicity.

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Sensitivity of dinB yafP operon mutants to different DNA damaging agents. (A) Sensitivity to 1 µM MMS, 10 µM NFZ and 10 µM NQO was estimated by spotting 7 µl of 5-fold serial dilutions of overnight cultures of the WT, dinB, yafP and dinB yafP strains onto LB plates containing DNA damaging agents. Strains carried the pGB2 plasmid or the pGB2 coding for the functional dinB gene. Experiments were repeated three times. Representative results are shown. (B and C) Effect of NQO on growth of WT, dinB, yafP and dinB yafP strains in liquid LB (B) or LB with 20 µM NQO (C). Experiments were repeated three times. Representative growth curves are shown.
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Figure 4: Sensitivity of dinB yafP operon mutants to different DNA damaging agents. (A) Sensitivity to 1 µM MMS, 10 µM NFZ and 10 µM NQO was estimated by spotting 7 µl of 5-fold serial dilutions of overnight cultures of the WT, dinB, yafP and dinB yafP strains onto LB plates containing DNA damaging agents. Strains carried the pGB2 plasmid or the pGB2 coding for the functional dinB gene. Experiments were repeated three times. Representative results are shown. (B and C) Effect of NQO on growth of WT, dinB, yafP and dinB yafP strains in liquid LB (B) or LB with 20 µM NQO (C). Experiments were repeated three times. Representative growth curves are shown.

Mentions: Because we observed that the presence of YafP is deleterious for bacteria in the absence of Pol IV in an in vivo stressful condition (Figure 3C), we hypothesized that YafP activity could be involved in the generation or in the processing of DNA lesions that are substrates for Pol IV. Thus, we tested the toxicity of several well known DNA damaging agents by spotting serial dilutions of the different CFT073 mutants having either an empty plasmid or plasmid expressing the functional dinB on LB plates containing DNA damaging agents (Figure 4A). Because we showed that YafP protein is synthesized upon treatment of cells with SOS inducing MMC, we tested resistance of WT strain and the different mutants to this DNA damaging agent and found no difference (data not shown). This suggests that neither YafP protein nor DinB are involved, directly or indirectly, in SOS induction via MMC or in the repair of DNA crosslinks.Figure 4.


Escherichia coli YafP protein modulates DNA damaging property of the nitroaromatic compounds.

Gutierrez A, Elez M, Clermont O, Denamur E, Matic I - Nucleic Acids Res. (2011)

Sensitivity of dinB yafP operon mutants to different DNA damaging agents. (A) Sensitivity to 1 µM MMS, 10 µM NFZ and 10 µM NQO was estimated by spotting 7 µl of 5-fold serial dilutions of overnight cultures of the WT, dinB, yafP and dinB yafP strains onto LB plates containing DNA damaging agents. Strains carried the pGB2 plasmid or the pGB2 coding for the functional dinB gene. Experiments were repeated three times. Representative results are shown. (B and C) Effect of NQO on growth of WT, dinB, yafP and dinB yafP strains in liquid LB (B) or LB with 20 µM NQO (C). Experiments were repeated three times. Representative growth curves are shown.
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Related In: Results  -  Collection

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Figure 4: Sensitivity of dinB yafP operon mutants to different DNA damaging agents. (A) Sensitivity to 1 µM MMS, 10 µM NFZ and 10 µM NQO was estimated by spotting 7 µl of 5-fold serial dilutions of overnight cultures of the WT, dinB, yafP and dinB yafP strains onto LB plates containing DNA damaging agents. Strains carried the pGB2 plasmid or the pGB2 coding for the functional dinB gene. Experiments were repeated three times. Representative results are shown. (B and C) Effect of NQO on growth of WT, dinB, yafP and dinB yafP strains in liquid LB (B) or LB with 20 µM NQO (C). Experiments were repeated three times. Representative growth curves are shown.
Mentions: Because we observed that the presence of YafP is deleterious for bacteria in the absence of Pol IV in an in vivo stressful condition (Figure 3C), we hypothesized that YafP activity could be involved in the generation or in the processing of DNA lesions that are substrates for Pol IV. Thus, we tested the toxicity of several well known DNA damaging agents by spotting serial dilutions of the different CFT073 mutants having either an empty plasmid or plasmid expressing the functional dinB on LB plates containing DNA damaging agents (Figure 4A). Because we showed that YafP protein is synthesized upon treatment of cells with SOS inducing MMC, we tested resistance of WT strain and the different mutants to this DNA damaging agent and found no difference (data not shown). This suggests that neither YafP protein nor DinB are involved, directly or indirectly, in SOS induction via MMC or in the repair of DNA crosslinks.Figure 4.

Bottom Line: Using a murine septicaemia model, we showed that YafP activity reduced the bacterial fitness in the absence of PolIV.The YafP antimutator activity was independent of the PolIV activity.Given that YafP was annotated as a putative acetyltransferase, it could be that YafP participates in the metabolic transformation of genotoxic compounds, hence modulating the balance between their mutagenicity and cytotoxicity.

View Article: PubMed Central - PubMed

Affiliation: Faculté de Médecine Paris Descartes, Inserm U1001, Université Paris Descartes, Paris, France.

ABSTRACT
Escherichia coli SOS functions constitute a multifaceted response to DNA damage. We undertook to study the role of yafP, a SOS gene with unknown function. yafP is part of an operon also containing the dinB gene coding for DNA Polymerase IV (PolIV). Our phylogenetic analysis showed that the gene content of this operon is variable but that the dinB and the yafP genes are conserved in the majority of E. coli natural isolates. Therefore, we studied if these proteins are functionally linked. Using a murine septicaemia model, we showed that YafP activity reduced the bacterial fitness in the absence of PolIV. Similarly, YafP increased cytotoxicity of two DNA damaging nitroaromatic compounds, 4-nitroquinoline-1-oxide (NQO) and nitrofurazone, in the absence of PolIV. The fact that PolIV counterbalances YafP-induced cytotoxicity could explain why these two genes are transcriptionally linked. We also studied the involvement of YafP in genotoxic-stress induced mutagenesis and found that PolIV and YafP reduced NQO-induced mutagenicity. The YafP antimutator activity was independent of the PolIV activity. Given that YafP was annotated as a putative acetyltransferase, it could be that YafP participates in the metabolic transformation of genotoxic compounds, hence modulating the balance between their mutagenicity and cytotoxicity.

Show MeSH
Related in: MedlinePlus