A Transcriptional Regulator and ABC Transporters Link Stress Tolerance, (p)ppGpp, and Genetic Competence in
            <i>Streptococcus mutans</i>

Seaton, Kinda; Ahn, Sang‐Joon; Sagstetter, Ann M.; Burne, Robert A.

doi:10.1128/jb.01257-10

Cited by 65 publications

(111 citation statements)

References 47 publications

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“…Reporter strains containing fusions of the promoter regions of the fructosyltransferase gene (ftf) and the nonlantibiotic mutacin D gene (nlmD) were constructed by amplifying the promoter region and the ribosome-binding site (RBS) of each gene from S. mutans UA159 and cloning them in front of the staphylococcal chloramphenicol acetyltransferase gene (cat) lacking its intrinsic promoter and RBS, resulting in strains with Pftf-cat and PnlmD-cat reporter fusions, respectively. The PnlmD-cat fusion was introduced as a single copy into the mtlA-phn region of the chromosome of S. mutans UA159 using the integration vector pJL105 (50). For the Pftf-cat reporter fusion, plasmid pBGK was used as the integration vector to introduce the fusion as a single copy in the gtfA gene.…”

Section: Methodsmentioning

confidence: 99%

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Effects of Arginine on Streptococcus mutans Growth, Virulence Gene Expression, and Stress Tolerance

Chakraborty

Burne

2017

Appl Environ Microbiol

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Streptococcus mutans is a common constituent of oral biofilms and a primary etiologic agent of human dental caries. The bacteria associated with dental caries have potent abilities to produce organic acids from dietary carbohydrates and to grow and metabolize in acidic conditions. By contrast, many commensal bacteria produce ammonia through the arginine deiminase system (ADS), which moderates the pH of oral biofilms. Arginine metabolism by the ADS is a significant deterrent to the initiation and progression of dental caries. In this study, we observed how exogenously provided L-arginine affects the growth, the virulence properties, and the tolerance of environmental stresses of S. mutans. Supplementation with 1.5% arginine (final concentration) had an inhibitory effect on the growth of S. mutans in complex and chemically defined media, particularly when cells were exposed to acid or oxidative stress. The genes encoding virulence factors required for attachment/accumulation (gtfB and spaP), bacteriocins (nlmA, nlmB, nlmD, and cipB), and the sigma factor required for competence development (comX) were downregulated during growth with 1.5% arginine. Deep sequencing of RNA (RNA-Seq) comparing the transcriptomes of S. mutans growing in chemically defined media with and without 1.5% arginine revealed differential expression of genes encoding ATP-binding cassette transporters, metal transporters, and constituents required for survival, metabolism, and biofilm formation. Therefore, the mechanisms of action by which arginine inhibits dental caries include direct adverse effects on multiple virulence-related properties of the most common human dental caries pathogen.IMPORTANCE Metabolism of the amino acid arginine by the arginine deiminase system (ADS) of certain oral bacteria raises the pH of dental plaque and provides a selective advantage to health-associated bacteria, thereby protecting the host from dental caries (cavities). Here, we examine the effects of arginine on the cavitycausing bacterium Streptococcus mutans. We find that arginine negatively impacts the growth, the pathogenic potential, and the tolerance of environmental stresses in a way that is likely to compromise the ability of S. mutans to cause disease. Using genetic and genomic techniques, multiple mechanisms by which arginine exerts its influence on virulence-related properties of S. mutans are discovered. This report demonstrates that a primary mechanism of action by which arginine inhibits the initiation and progression of dental caries may be by reducing the pathogenic potential of S. mutans.

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Section: Methodsmentioning

confidence: 99%

“…Colonies were counted after 48 h of incubation at 37°C in a 5% CO 2 aerobic atmosphere. Transformation efficiency was expressed as the number of transformants divided by the total number of viable cells; this number was then multiplied by 100 to obtain percentage of transformants (50).…”

Section: Methodsmentioning

confidence: 99%

Effects of Arginine on Streptococcus mutans Growth, Virulence Gene Expression, and Stress Tolerance

Chakraborty

Burne

2017

Appl Environ Microbiol

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“…This indicates that RelRS is involved in the regulation of (p)ppGpp metabolism (127). The environmental signals to which RelRS responds have not been determined, although it has been suggested that RelRS may sense oxidative stressors or by-products of oxidative metabolism (131). Expression of relPRS is regulated by the MarR family transcriptional regulator RcrR, which is involved in stress tolerance and competence, although the signals to which RcrR responds remain unidentified (131,132).…”

Section: The Stringent Response: the Ribosome As A Sensormentioning

confidence: 99%

Stress Physiology of Lactic Acid Bacteria

Papadimitriou

Alegría

Bron

et al. 2016

Microbiol Mol Biol Rev

515

404

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SUMMARYLactic acid bacteria (LAB) are important starter, commensal, or pathogenic microorganisms. The stress physiology of LAB has been studied in depth for over 2 decades, fueled mostly by the technological implications of LAB robustness in the food industry. Survival of probiotic LAB in the host and the potential relatedness of LAB virulence to their stress resilience have intensified interest in the field. Thus, a wealth of information concerning stress responses exists today for strains as diverse as starter (e.g.,Lactococcus lactis), probiotic (e.g., severalLactobacillusspp.), and pathogenic (e.g.,EnterococcusandStreptococcusspp.) LAB. Here we present the state of the art for LAB stress behavior. We describe the multitude of stresses that LAB are confronted with, and we present the experimental context used to study the stress responses of LAB, focusing on adaptation, habituation, and cross-protection as well as on self-induced multistress resistance in stationary phase, biofilms, and dormancy. We also consider stress responses at the population and single-cell levels. Subsequently, we concentrate on the stress defense mechanisms that have been reported to date, grouping them according to their direct participation in preserving cell energy, defending macromolecules, and protecting the cell envelope. Stress-induced responses of probiotic LAB and commensal/pathogenic LAB are highlighted separately due to the complexity of the peculiar multistress conditions to which these bacteria are subjected in their hosts. Induction of prophages under environmental stresses is then discussed. Finally, we present systems-based strategies to characterize the “stressome” of LAB and to engineer new food-related and probiotic LAB with improved stress tolerance.

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“…For example, SASs might respond to different internal or external cues than the Rel enzyme. In support, there is evidence of transcriptional induction of the Bacillus subtilis, Staphylococcus aureus, and Streptococcus mutans SASs under alkaline, cell wall, or oxygen stresses, respectively (22,24,30). Alternatively, they might differ from long RSH enzymes because their constitutive synthetase activity ensures persistent basal (p)ppGpp production (21,28).…”

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confidence: 99%

From (p)ppGpp to (pp)pGpp: Characterization of Regulatory Effects of pGpp Synthesized by the Small Alarmone Synthetase of Enterococcus faecalis

et al. 2015

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The bacterial stringent response (SR) is a conserved stress tolerance mechanism that orchestrates physiological alterations to enhance cell survival. This response is mediated by the intracellular accumulation of the alarmones pppGpp and ppGpp, collectively called (p)ppGpp. In Enterococcus faecalis, (p)ppGpp metabolism is carried out by the bifunctional synthetase/hydrolase E. faecalis Rel (Rel Ef ) and the small alarmone synthetase (SAS) RelQ Ef . Although Rel is the main enzyme responsible for SR activation in Firmicutes, there is emerging evidence that SASs can make important contributions to bacterial homeostasis. Here, we showed that RelQ Ef synthesizes ppGpp more efficiently than pppGpp without the need for ribosomes, tRNA, or mRNA. In addition to (p)ppGpp synthesis from GDP and GTP, RelQ Ef also efficiently utilized GMP to form GMP 3=-diphosphate (pGpp). Based on this observation, we sought to determine if pGpp exerts regulatory effects on cellular processes affected by (p)ppGpp. We found that pGpp, like (p)ppGpp, strongly inhibits the activity of E. faecalis enzymes involved in GTP biosynthesis and, to a lesser extent, transcription of rrnB by Escherichia coli RNA polymerase. Activation of E. coli RelA synthetase activity was observed in the presence of both pGpp and ppGpp, while RelQ Ef was activated only by ppGpp. Furthermore, enzymatic activity of RelQ Ef is insensitive to relacin, a (p)ppGpp analog developed as an inhibitor of "long" RelA/SpoT homolog (RSH) enzymes. We conclude that pGpp can likely function as a bacterial alarmone with target-specific regulatory effects that are similar to what has been observed for (p)ppGpp. IMPORTANCEAccumulation of the nucleotide second messengers (p)ppGpp in bacteria is an important signal regulating genetic and physiological networks contributing to stress tolerance, antibiotic persistence, and virulence. Understanding the function and regulation of the enzymes involved in (p)ppGpp turnover is therefore critical for designing strategies to eliminate the protective effects of this molecule. While characterizing the (p)ppGpp synthetase RelQ of Enterococcus faecalis (RelQ Ef ), we found that, in addition to (p)ppGpp, RelQ Ef is an efficient producer of pGpp (GMP 3=-diphosphate). In vitro analysis revealed that pGpp exerts complex, target-specific effects on processes known to be modulated by (p)ppGpp. These findings provide a new regulatory feature of RelQ Ef and suggest that pGpp may represent a new member of the (pp)pGpp family of alarmones. In order to survive under adverse environmental conditions, such as nutrient starvation, bacteria have evolved complex, interconnected regulatory networks that sense and integrate internal and external metabolic cues to activate cellular responses that enhance bacterial survival. One critical and highly conserved mechanism employed by bacteria to cope with nutritional as well as a variety of environmental and chemical stresses is the stringent response (SR). The SR is mediated by the accumulation of two guanine analog...

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A Transcriptional Regulator and ABC Transporters Link Stress Tolerance, (p)ppGpp, and Genetic Competence in Streptococcus mutans

Cited by 65 publications

References 47 publications

Effects of Arginine on Streptococcus mutans Growth, Virulence Gene Expression, and Stress Tolerance

Effects of Arginine on Streptococcus mutans Growth, Virulence Gene Expression, and Stress Tolerance

Stress Physiology of Lactic Acid Bacteria

From (p)ppGpp to (pp)pGpp: Characterization of Regulatory Effects of pGpp Synthesized by the Small Alarmone Synthetase of Enterococcus faecalis

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