Limits...
Oxidative stress enhances the expression of sulfur assimilation genes: preliminary insights on the Enterococcus faecalis iron-sulfur cluster machinery regulation.

Riboldi GP, Bierhals CG, Mattos EP, Frazzon AP, d'Azevedo PA, Frazzon J - Mem. Inst. Oswaldo Cruz (2014)

Bottom Line: In Firmicutes species, a unique operon composed of the sufCDSUB genes is responsible for [Fe-S] cluster biogenesis.The aim of this study was to investigate the potential of the E. faecalis sufCDSUB system in the [Fe-S] cluster assembly using oxidative stress and iron depletion as adverse growth conditions.Likewise, strong expression of the sufCDSUB genes was observed in low concentrations of hydrogen peroxide, indicating that the lowest concentration of oxygen free radicals inside cells, known to be highly damaging to [Fe-S] clusters, is sufficient to trigger the transcriptional machinery for prompt replacement of [Fe-S] clusters.

View Article: PubMed Central - PubMed

Affiliation: Laboratório de Cocos Gram-positivos e Microbiologia Molecular, Departamento de Microbiologia, Universidade Federal de Ciências da Saúde de Porto Alegre, Porto Alegre, RS, Brasil.

ABSTRACT
The Firmicutes bacteria participate extensively in virulence and pathological processes. Enterococcus faecalis is a commensal microorganism; however, it is also a pathogenic bacterium mainly associated with nosocomial infections in immunocompromised patients. Iron-sulfur [Fe-S] clusters are inorganic prosthetic groups involved in diverse biological processes, whose in vivo formation requires several specific protein machineries. Escherichia coli is one of the most frequently studied microorganisms regarding [Fe-S] cluster biogenesis and encodes the iron-sulfur cluster and sulfur assimilation systems. In Firmicutes species, a unique operon composed of the sufCDSUB genes is responsible for [Fe-S] cluster biogenesis. The aim of this study was to investigate the potential of the E. faecalis sufCDSUB system in the [Fe-S] cluster assembly using oxidative stress and iron depletion as adverse growth conditions. Quantitative real-time polymerase chain reaction demonstrated, for the first time, that Gram-positive bacteria possess an OxyR component responsive to oxidative stress conditions, as fully described for E. coli models. Likewise, strong expression of the sufCDSUB genes was observed in low concentrations of hydrogen peroxide, indicating that the lowest concentration of oxygen free radicals inside cells, known to be highly damaging to [Fe-S] clusters, is sufficient to trigger the transcriptional machinery for prompt replacement of [Fe-S] clusters.

Show MeSH

Related in: MedlinePlus

: critical threshold (Ct) values for quantitative polymerase chain reactionof gyrase beta chain (gyrB), RNA polymerase beta chain(rpoB), elongation factor for transporter RNA(tuf) and 23S ribosomal RNA (23SrRNA)genes of Enterococcus faecalis strain X1 under differentsimulations of iron depletion (left panel) and oxidative stress (right panel).DIP: 2’,2’-dipyridyl; H2O2: hydrogen peroxide.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4155840&req=5

f02: : critical threshold (Ct) values for quantitative polymerase chain reactionof gyrase beta chain (gyrB), RNA polymerase beta chain(rpoB), elongation factor for transporter RNA(tuf) and 23S ribosomal RNA (23SrRNA)genes of Enterococcus faecalis strain X1 under differentsimulations of iron depletion (left panel) and oxidative stress (right panel).DIP: 2’,2’-dipyridyl; H2O2: hydrogen peroxide.

Mentions: Relative expression of the SUF operon: finding appropriate housekeeping genesfor data normalisation - In an attempt to find good constitutive genecandidates for our qPCR measurements, we determined the expression profile ofrpoB, gyrB, tuf and 23SrRNA under the differentgrowth conditions studied here. As illustrated in Fig.2, the linearity observed for the gyrB andrpoB genes was satisfactory, with a minor variation of 1 Ct amongthe different cellular growth conditions. However, tuf and23SrRNA did not exhibit a regular expression pattern, as verified bythe wide Ct ranges observed under the various conditions. Additionally, insilico analysis of these candidate constitutive genes using the algorithmsNormfinder, geNorm and BestKeeper predicted gyrB andrpoB to be the most stably expressed genes for data normalisation. Similarexperiments with E. faecalis FA22 and JH22 strains corroborated theseresults (Supplementary data).


Oxidative stress enhances the expression of sulfur assimilation genes: preliminary insights on the Enterococcus faecalis iron-sulfur cluster machinery regulation.

Riboldi GP, Bierhals CG, Mattos EP, Frazzon AP, d'Azevedo PA, Frazzon J - Mem. Inst. Oswaldo Cruz (2014)

: critical threshold (Ct) values for quantitative polymerase chain reactionof gyrase beta chain (gyrB), RNA polymerase beta chain(rpoB), elongation factor for transporter RNA(tuf) and 23S ribosomal RNA (23SrRNA)genes of Enterococcus faecalis strain X1 under differentsimulations of iron depletion (left panel) and oxidative stress (right panel).DIP: 2’,2’-dipyridyl; H2O2: hydrogen peroxide.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4155840&req=5

f02: : critical threshold (Ct) values for quantitative polymerase chain reactionof gyrase beta chain (gyrB), RNA polymerase beta chain(rpoB), elongation factor for transporter RNA(tuf) and 23S ribosomal RNA (23SrRNA)genes of Enterococcus faecalis strain X1 under differentsimulations of iron depletion (left panel) and oxidative stress (right panel).DIP: 2’,2’-dipyridyl; H2O2: hydrogen peroxide.
Mentions: Relative expression of the SUF operon: finding appropriate housekeeping genesfor data normalisation - In an attempt to find good constitutive genecandidates for our qPCR measurements, we determined the expression profile ofrpoB, gyrB, tuf and 23SrRNA under the differentgrowth conditions studied here. As illustrated in Fig.2, the linearity observed for the gyrB andrpoB genes was satisfactory, with a minor variation of 1 Ct amongthe different cellular growth conditions. However, tuf and23SrRNA did not exhibit a regular expression pattern, as verified bythe wide Ct ranges observed under the various conditions. Additionally, insilico analysis of these candidate constitutive genes using the algorithmsNormfinder, geNorm and BestKeeper predicted gyrB andrpoB to be the most stably expressed genes for data normalisation. Similarexperiments with E. faecalis FA22 and JH22 strains corroborated theseresults (Supplementary data).

Bottom Line: In Firmicutes species, a unique operon composed of the sufCDSUB genes is responsible for [Fe-S] cluster biogenesis.The aim of this study was to investigate the potential of the E. faecalis sufCDSUB system in the [Fe-S] cluster assembly using oxidative stress and iron depletion as adverse growth conditions.Likewise, strong expression of the sufCDSUB genes was observed in low concentrations of hydrogen peroxide, indicating that the lowest concentration of oxygen free radicals inside cells, known to be highly damaging to [Fe-S] clusters, is sufficient to trigger the transcriptional machinery for prompt replacement of [Fe-S] clusters.

View Article: PubMed Central - PubMed

Affiliation: Laboratório de Cocos Gram-positivos e Microbiologia Molecular, Departamento de Microbiologia, Universidade Federal de Ciências da Saúde de Porto Alegre, Porto Alegre, RS, Brasil.

ABSTRACT
The Firmicutes bacteria participate extensively in virulence and pathological processes. Enterococcus faecalis is a commensal microorganism; however, it is also a pathogenic bacterium mainly associated with nosocomial infections in immunocompromised patients. Iron-sulfur [Fe-S] clusters are inorganic prosthetic groups involved in diverse biological processes, whose in vivo formation requires several specific protein machineries. Escherichia coli is one of the most frequently studied microorganisms regarding [Fe-S] cluster biogenesis and encodes the iron-sulfur cluster and sulfur assimilation systems. In Firmicutes species, a unique operon composed of the sufCDSUB genes is responsible for [Fe-S] cluster biogenesis. The aim of this study was to investigate the potential of the E. faecalis sufCDSUB system in the [Fe-S] cluster assembly using oxidative stress and iron depletion as adverse growth conditions. Quantitative real-time polymerase chain reaction demonstrated, for the first time, that Gram-positive bacteria possess an OxyR component responsive to oxidative stress conditions, as fully described for E. coli models. Likewise, strong expression of the sufCDSUB genes was observed in low concentrations of hydrogen peroxide, indicating that the lowest concentration of oxygen free radicals inside cells, known to be highly damaging to [Fe-S] clusters, is sufficient to trigger the transcriptional machinery for prompt replacement of [Fe-S] clusters.

Show MeSH
Related in: MedlinePlus