Functional studies of E. faecalis RNase J2 and its role in virulence and fitness

Gao, Peng; Pinkston, Kenneth L.; Bourgogne, Agathe; Murray, Barbara E.; Hoof, Ambro van; Harvey, Barrett R.

doi:10.1371/journal.pone.0175212

Cited by 16 publications

(10 citation statements)

References 39 publications

(64 reference statements)

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“…AhrC activates expression of both ebpR and ace , 24 and the ribonuclease activity of RNase J2 regulates pili and other virulence genes but does not affect ace . 41,67 Here, we showed that a bph mutant has reduced expression of ebpA and ebpR but wild-type levels of Ace and postulate that the overall decrease in pili production in this mutant is through reduced ebpR expression. Although it is unlikely that Bph regulates AhrC due to disparate Ace expression in these strains, both gene products are linked to arginine catabolism in addition to pili.…”

Section: Discussionmentioning

confidence: 66%

Exploiting biofilm phenotypes for functional characterization of hypothetical genes in Enterococcus faecalis

Willett

Dunny

2019

npj Biofilms Microbiomes

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Enterococcus faecalis is a commensal organism as well as an important nosocomial pathogen, and its infections are typically linked to biofilm formation. Nearly 25% of the E. faecalis OG1RF genome encodes hypothetical genes or genes of unknown function. Elucidating their function and how these gene products influence biofilm formation is critical for understanding E. faecalis biology. To identify uncharacterized early biofilm determinants, we performed a genetic screen using an arrayed transposon (Tn) library containing ~2000 mutants in hypothetical genes/intergenic regions and identified eight uncharacterized predicted protein-coding genes required for biofilm formation. We demonstrate that OG1RF_10435 encodes a phosphatase that modulates global protein expression and arginine catabolism and propose renaming this gene bph (biofilm phosphatase). We present a workflow for combining phenotype-driven experimental and computational evaluation of hypothetical gene products in E. faecalis, which can be used to study hypothetical genes required for biofilm formation and other phenotypes of diverse bacteria.

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Section: Discussionmentioning

confidence: 66%

Exploiting biofilm phenotypes for functional characterization of hypothetical genes in Enterococcus faecalis

Willett

Dunny

2019

npj Biofilms Microbiomes

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“…The available evidence is consistent with a posttranscriptional mechanism, potentially involving a negative regulatory sRNA targeting ebp transcripts and degraded by RNase J2. Since the deletion of rnjB does not alter ace expression (33), this mechanism is probably not related to the ArgR-dependent circuit described here.…”

Section: Discussionmentioning

confidence: 69%

“…However, these other regulatory mechanisms are posttranscriptional and likely to be unrelated to our results. An interesting mechanism for the control of pilus expression has been reported by Harvey and colleagues (33,34), who found that the deletion of the rnjB gene encoding E. faecalis RNase J2 dramatically reduces the levels of pilin gene transcripts and proteins. The available evidence is consistent with a posttranscriptional mechanism, potentially involving a negative regulatory sRNA targeting ebp transcripts and degraded by RNase J2.…”

Section: Discussionmentioning

confidence: 97%

Expression of Adhesive Pili and the Collagen-Binding Adhesin Ace Is Activated by ArgR Family Transcription Factors in Enterococcus faecalis

Manias

Dunny

2018

J Bacteriol

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It was shown previously that the disruption of the gene encoding a predicted ArgR family transcription factor results in a severe defect in biofilm formation, as well as a significant attenuation of virulence of strain OG1RF in multiple experimental infection models. Using transcriptome sequencing (RNA-seq), we observed-dependent changes in the expression of more than 20 genes. AhrC-repressed genes included predicted determinants of arginine catabolism and several other metabolic genes and predicted transporters, while AhrC-activated genes included determinants involved in the production of surface protein adhesins. Most notably, the structural and regulatory genes of the locus encoding adhesive pili were positively regulated, as well as the gene, encoding a collagen-binding adhesin. Using transcription reporter fusions, we determined that and a second transcription factor gene,, both function to activate the expression of , which directly activates the transcription of the pilus structural genes. Our data suggest that in the wild-type, the low levels of EbpR limit the expression of pili and that biofilm biomass is also limited by the amount of pili expressed by the bacteria. The expression of is similarly enhanced by AhrC and ArgR2, but expression is not dependent on EbpR. Our results demonstrate the existence of novel regulatory cascades controlled by a pair of ArgR family transcription factors that might function as a heteromeric protein complex. Cell surface adhesins play critical roles in the formation of biofilms, host colonization, and the pathogenesis of opportunistic infections by Here, we present new results showing that the expression of two major enterococcal surface adhesins, pili, and the collagen-binding protein Ace is positively regulated at the transcription level by two family transcription factors, AhrC and ArgR2. In the case of pili, the direct target of regulation is the gene, previously shown to activate the transcription of the pilus structural genes, while the activation of transcription appears to be directly impacted by the two ArgR proteins. These transcription factors may represent new targets for blocking enterococcal infections.

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“…Here we add another environmental signal, iron abundance, that can influence Ebp expression. Several intrinsic factors can impact ebp gene expression including transcriptional regulators, EbpR 47 , AhrC (also annotated as ArgR3) and ArgR2 48 , the FsrB quorum sensing peptide 49 and the RNA processing enzyme RNase J2 50, 51 . While none of these factors were differentially regulated upon iron supplementation which may suggest uncharacterized regulatory pathways are responsible for iron-regulated ebp transcription, we cannot rule out the possibility that excess iron impacts the function of these known regulators of ebp expression.…”

Section: Discussionmentioning

confidence: 99%

Sortase-assembled pili promote extracellular electron transfer and iron acquisition inEnterococcus faecalisbiofilm

Lam

Wong

Matysik

et al. 2019

Preprint

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24Enterococcus faecalis is an opportunistic human pathogen and the cause of biofilm-25 associated infections of the heart, catheterized urinary tract, wounds, and the dysbiotic gut 26 where it can expand to high numbers upon microbiome perturbations. The E. faecalis sortase-27 assembled endocarditis and biofilm associated pilus (Ebp) is involved in adhesion and 28 biofilm formation in vitro and in vivo. Extracellular electron transfer (EET) also promotes E. 29 faecalis biofilm formation in iron-rich environments, however neither the mechanism 30 underlying EET nor its role in virulence was previously known. Here we show that iron 31 associated with Ebp serve as a terminal electron acceptor for EET, leading to extracellular 32 iron reduction and intracellular iron accumulation. We found that a MIDAS motif within the 33 EbpA tip adhesin is required for interaction with iron, EET, and FeoB-mediated iron uptake. 34 We demonstrate that MenB and Ndh3, essential components of the aerobic respiratory chain 35 and a specialized flavin-mediated electron transport chain, respectively, are required for iron-36 mediated EET. In addition, using a mouse gastrointestinal (GI) colonization model, we show 37 that EET is essential for colonization of the GI tract, and Ebp is essential for augmented E. 38 faecalis GI colonization when dietary iron is in excess. Taken together, our findings show 39 that pilus mediated capture of iron within biofilms enables EET-mediated iron acquisition in 40 E. faecalis, and that these processes plays an important role in E. faecalis expansion in the GI 41 tract. 42 43 Significance 44 Understanding enterococcal biofilm development is the first step towards improved 45 therapeutics for the often antimicrobial resistant infections caused by these bacteria. Here we 46 report a role for Enterococcus faecalis endocarditis and biofilm associated pili (Ebp) in 47 (EET) which in turn promotes iron acquisition. Furthermore, we characterize the mechanisms 49 underlying electron transfer in the E. faecalis biofilm. Our findings support a model in which 50 E. faecalis use EET to drive the reduction of pilus-associated ferric iron, leading to iron 51 acquisition in E. faecalis biofilm, and contributing to enterococcal virulence in the GI tract.52 53 Introduction 54 Enterococcus faecalis is an important human opportunistic pathogen that causes a variety of 55 diseases including endocarditis, urinary tract infections (UTI), bacteremia, wound infection, 56 and medical device-associated infections 1 . Many of these infections are polymicrobial and 57 biofilm-associated, rendering them more tolerant to antimicrobial and immune clearance, and 58 contributing to their persistent nature 1 . Therefore, a detailed understanding of enterococcal 59 biofilm development is a critical step towards advancing new therapeutics for treatment of 60 these often antimicrobial resistant infections 1,2 . 61 62One important factor contributing to E. faecalis biofilm formation and virulence is the 63 sortase-assembled endo...

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Functional studies of E. faecalis RNase J2 and its role in virulence and fitness

Cited by 16 publications

References 39 publications

Exploiting biofilm phenotypes for functional characterization of hypothetical genes in Enterococcus faecalis

Exploiting biofilm phenotypes for functional characterization of hypothetical genes in Enterococcus faecalis

Expression of Adhesive Pili and the Collagen-Binding Adhesin Ace Is Activated by ArgR Family Transcription Factors in Enterococcus faecalis

Sortase-assembled pili promote extracellular electron transfer and iron acquisition inEnterococcus faecalisbiofilm

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