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The Campylobacter jejuni MarR-like transcriptional regulators RrpA and RrpB both influence bacterial responses to oxidative and aerobic stresses.

Gundogdu O, da Silva DT, Mohammad B, Elmi A, Mills DC, Wren BW, Dorrell N - Front Microbiol (2015)

Bottom Line: Mutation of either rrpA or rrpB reduces catalase (KatA) expression.Mutation of either rrpA or rrpB also results in a reduction in the level of katA expression, but this reduction was not observed in the rrpAB double mutant.Together these data indicate a role for both RrpA and RrpB in the C. jejuni peroxide oxidative and aerobic (O2) stress responses, enhancing bacterial survival in vivo and in the environment.

View Article: PubMed Central - PubMed

Affiliation: Faculty of Infectious and Tropical Diseases, London School of Hygiene and Tropical Medicine , London, UK.

ABSTRACT
The ability of the human intestinal pathogen Campylobacter jejuni to respond to oxidative stress is central to bacterial survival both in vivo during infection and in the environment. Re-annotation of the C. jejuni NCTC11168 genome revealed the presence of two MarR-type transcriptional regulators Cj1546 and Cj1556, originally annotated as hypothetical proteins, which we have designated RrpA and RrpB (regulator of response to peroxide) respectively. Previously we demonstrated a role for RrpB in both oxidative and aerobic (O2) stress and that RrpB was a DNA binding protein with auto-regulatory activity, typical of MarR-type transcriptional regulators. In this study, we show that RrpA is also a DNA binding protein and that a rrpA mutant in strain 11168H exhibits increased sensitivity to hydrogen peroxide oxidative stress. Mutation of either rrpA or rrpB reduces catalase (KatA) expression. However, a rrpAB double mutant exhibits higher levels of resistance to hydrogen peroxide oxidative stress, with levels of KatA expression similar to the wild-type strain. Mutation of either rrpA or rrpB also results in a reduction in the level of katA expression, but this reduction was not observed in the rrpAB double mutant. Neither the rrpA nor rrpB mutant exhibits any significant difference in sensitivity to either cumene hydroperoxide or menadione oxidative stresses, but both mutants exhibit a reduced ability to survive aerobic (O2) stress, enhanced biofilm formation and reduced virulence in the Galleria mellonella infection model. The rrpAB double mutant exhibits wild-type levels of biofilm formation and wild-type levels of virulence in the G mellonella infection model. Together these data indicate a role for both RrpA and RrpB in the C. jejuni peroxide oxidative and aerobic (O2) stress responses, enhancing bacterial survival in vivo and in the environment.

No MeSH data available.


Related in: MedlinePlus

(A) Effect of oxidative stress on the survival of C. jejuni 11168H wild-type strain, rrpA, rrpA complement strain (rrpA comp), rrpB, rrpB complement strain (rrpB comp), katA and perR mutants. C. jejuni strains were incubated with 25, 50, or 100 mM H2O2 for 15 min at 37°C then bacterial survival assessed. (B) Catalase activity assays were performed on bacterial whole cell lysates from the 11168H wild-type strain, rrpA, rrpA comp, rrpB, rrpB comp, katA and perR mutants for 1 min. Asterisks denote a statistically significant difference (*p < 0.05, **p < 0.01, ***p < 0.001).
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Figure 2: (A) Effect of oxidative stress on the survival of C. jejuni 11168H wild-type strain, rrpA, rrpA complement strain (rrpA comp), rrpB, rrpB complement strain (rrpB comp), katA and perR mutants. C. jejuni strains were incubated with 25, 50, or 100 mM H2O2 for 15 min at 37°C then bacterial survival assessed. (B) Catalase activity assays were performed on bacterial whole cell lysates from the 11168H wild-type strain, rrpA, rrpA comp, rrpB, rrpB comp, katA and perR mutants for 1 min. Asterisks denote a statistically significant difference (*p < 0.05, **p < 0.01, ***p < 0.001).

Mentions: The rrpA mutant and rrpA complement were tested for sensitivity to H2O2 (25, 50, and 100 mM). The rrpA mutant exhibited increased sensitivity to 25 mM H2O2 stress, whilst the rrpA complement exhibited survival similar to the wild-type strain (Figure 2A). As controls, H2O2 stress assays were also performed on 11168H katA and perR mutants. The perR mutant displayed a high level of resistance to H2O2 stress, in confirmation of a previous study which demonstrated that a C. jejuni NCTC11168 perR mutant constitutively expressed both katA and ahpC, resulting in enhanced resistance to peroxide stress compared to the wild-type strain (van Vliet et al., 1999). The katA mutant does not express catalase and did not survive exposure to any concentration of H2O2. Catalase activity assays were performed on bacterial whole cell lysates. Both the rrpA and rrpB mutants exhibited reduced catalase activity compared to the wild-type strain, whilst the katA mutant had no catalase activity and the perR mutant exhibited significantly increased catalase activity (Figure 2B). Catalase activity was restored to wild-type levels in both the rrpA and rrpB complements (Figure 2).


The Campylobacter jejuni MarR-like transcriptional regulators RrpA and RrpB both influence bacterial responses to oxidative and aerobic stresses.

Gundogdu O, da Silva DT, Mohammad B, Elmi A, Mills DC, Wren BW, Dorrell N - Front Microbiol (2015)

(A) Effect of oxidative stress on the survival of C. jejuni 11168H wild-type strain, rrpA, rrpA complement strain (rrpA comp), rrpB, rrpB complement strain (rrpB comp), katA and perR mutants. C. jejuni strains were incubated with 25, 50, or 100 mM H2O2 for 15 min at 37°C then bacterial survival assessed. (B) Catalase activity assays were performed on bacterial whole cell lysates from the 11168H wild-type strain, rrpA, rrpA comp, rrpB, rrpB comp, katA and perR mutants for 1 min. Asterisks denote a statistically significant difference (*p < 0.05, **p < 0.01, ***p < 0.001).
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Related In: Results  -  Collection

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Figure 2: (A) Effect of oxidative stress on the survival of C. jejuni 11168H wild-type strain, rrpA, rrpA complement strain (rrpA comp), rrpB, rrpB complement strain (rrpB comp), katA and perR mutants. C. jejuni strains were incubated with 25, 50, or 100 mM H2O2 for 15 min at 37°C then bacterial survival assessed. (B) Catalase activity assays were performed on bacterial whole cell lysates from the 11168H wild-type strain, rrpA, rrpA comp, rrpB, rrpB comp, katA and perR mutants for 1 min. Asterisks denote a statistically significant difference (*p < 0.05, **p < 0.01, ***p < 0.001).
Mentions: The rrpA mutant and rrpA complement were tested for sensitivity to H2O2 (25, 50, and 100 mM). The rrpA mutant exhibited increased sensitivity to 25 mM H2O2 stress, whilst the rrpA complement exhibited survival similar to the wild-type strain (Figure 2A). As controls, H2O2 stress assays were also performed on 11168H katA and perR mutants. The perR mutant displayed a high level of resistance to H2O2 stress, in confirmation of a previous study which demonstrated that a C. jejuni NCTC11168 perR mutant constitutively expressed both katA and ahpC, resulting in enhanced resistance to peroxide stress compared to the wild-type strain (van Vliet et al., 1999). The katA mutant does not express catalase and did not survive exposure to any concentration of H2O2. Catalase activity assays were performed on bacterial whole cell lysates. Both the rrpA and rrpB mutants exhibited reduced catalase activity compared to the wild-type strain, whilst the katA mutant had no catalase activity and the perR mutant exhibited significantly increased catalase activity (Figure 2B). Catalase activity was restored to wild-type levels in both the rrpA and rrpB complements (Figure 2).

Bottom Line: Mutation of either rrpA or rrpB reduces catalase (KatA) expression.Mutation of either rrpA or rrpB also results in a reduction in the level of katA expression, but this reduction was not observed in the rrpAB double mutant.Together these data indicate a role for both RrpA and RrpB in the C. jejuni peroxide oxidative and aerobic (O2) stress responses, enhancing bacterial survival in vivo and in the environment.

View Article: PubMed Central - PubMed

Affiliation: Faculty of Infectious and Tropical Diseases, London School of Hygiene and Tropical Medicine , London, UK.

ABSTRACT
The ability of the human intestinal pathogen Campylobacter jejuni to respond to oxidative stress is central to bacterial survival both in vivo during infection and in the environment. Re-annotation of the C. jejuni NCTC11168 genome revealed the presence of two MarR-type transcriptional regulators Cj1546 and Cj1556, originally annotated as hypothetical proteins, which we have designated RrpA and RrpB (regulator of response to peroxide) respectively. Previously we demonstrated a role for RrpB in both oxidative and aerobic (O2) stress and that RrpB was a DNA binding protein with auto-regulatory activity, typical of MarR-type transcriptional regulators. In this study, we show that RrpA is also a DNA binding protein and that a rrpA mutant in strain 11168H exhibits increased sensitivity to hydrogen peroxide oxidative stress. Mutation of either rrpA or rrpB reduces catalase (KatA) expression. However, a rrpAB double mutant exhibits higher levels of resistance to hydrogen peroxide oxidative stress, with levels of KatA expression similar to the wild-type strain. Mutation of either rrpA or rrpB also results in a reduction in the level of katA expression, but this reduction was not observed in the rrpAB double mutant. Neither the rrpA nor rrpB mutant exhibits any significant difference in sensitivity to either cumene hydroperoxide or menadione oxidative stresses, but both mutants exhibit a reduced ability to survive aerobic (O2) stress, enhanced biofilm formation and reduced virulence in the Galleria mellonella infection model. The rrpAB double mutant exhibits wild-type levels of biofilm formation and wild-type levels of virulence in the G mellonella infection model. Together these data indicate a role for both RrpA and RrpB in the C. jejuni peroxide oxidative and aerobic (O2) stress responses, enhancing bacterial survival in vivo and in the environment.

No MeSH data available.


Related in: MedlinePlus