Role of the (p)ppGpp Synthase RSH, a RelA/SpoT Homolog, in Stringent Response and Virulence of<i>Staphylococcus aureus</i>

Geiger, Tobias; Goerke, Christiane; Fritz, Michaela; Schäfer, Tina; Ohlsen, Knut; Liebeke, Manuel; Lalk, Michael; Wolz, Christiane

doi:10.1128/iai.01439-09

Cited by 128 publications

(213 citation statements)

References 58 publications

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“…However, loss of Rel did not lower persistence rates and, depending on the drug target, the ⌬rel strain, which has ϳ4-foldhigher basal levels of ppGpp due to constitutive alarmone synthesis by RelQ (54), produced a significantly higher number of persisters than the parent strain (45). This finding is a departure from the general concept that the SR mediates bacterial persistence, as activation of the SR in E. faecalis and Firmicutes in general is dependent on the Rel enzyme (43,45,66,74,(99)(100)(101). Resolution of this notion will require systematic studies on basal (p)ppGpp elevation in other related species.…”

Section: Figcontrasting

confidence: 55%

“…Specifically, reduction in cellular pools of the coeffector GTP by (p)ppGpp results in less stable CodY-DNA interactions, thereby alleviating CodY regulation (71). Importantly, the (p)ppGpp/CodY association is critical for nutrient stress tolerance and virulence in several bacterial pathogens (72)(73)(74)(75)(76)(77)). Yet, the relative contributions of GTP-and CodY-dependent mechanisms for stress tolerance and virulence controlled by (p)ppGpp are not entirely clear, as GTP depletion can also affect stress survival in a CodY-independent manner (67,68).…”

Section: (P)ppgpp and Codymentioning

confidence: 99%

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Many Means to a Common End: the Intricacies of (p)ppGpp Metabolism and Its Control of Bacterial Homeostasis

2015

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In nearly all bacterial species examined so far, amino acid starvation triggers the rapid accumulation of the nucleotide second messenger (p)ppGpp, the effector of the stringent response. While for years the enzymes involved in (p)ppGpp metabolism and the significance of (p)ppGpp accumulation to stress survival were considered well defined, a recent surge of interest in the field has uncovered an unanticipated level of diversity in how bacteria metabolize and utilize (p)ppGpp to rapidly synchronize a variety of biological processes important for growth and stress survival. In addition to the classic activation of the stringent response, it has become evident that (p)ppGpp exerts differential effects on cell physiology in an incremental manner rather than simply acting as a biphasic switch that controls growth or stasis. Of particular interest is the intimate relationship of (p)ppGpp with persister cell formation and virulence, which has spurred the pursuit of (p)ppGpp inhibitors as a means to control recalcitrant infections. Here, we present an overview of the enzymes responsible for (p)ppGpp metabolism, elaborate on the intricacies that link basal production of (p)ppGpp to bacterial homeostasis, and discuss the implications of targeting (p)ppGpp synthesis as a means to disrupt long-term bacterial survival strategies. While analyzing nucleotide extracts of Escherichia coli, Cashel and Gallant visualized two spots by thin-layer chromatography that could be implicated in the inhibition of stable RNA accumulation provoked by amino acid starvation, which they dubbed "magic spots" (1). These magic spots were later identified as the hyperphosphorylated guanosine derivatives ppGpp (GDP, 3=-diphosphate) and pppGpp (GTP, 3=-diphosphate), collectively referred to as (p)ppGpp, or alarmones (2, 3). Subsequent studies revealed that (p)ppGpp is responsible for activation of the stringent response (SR), a highly conserved stress response to nutrient starvation (4, 5). Generally speaking, accumulation of (p)ppGpp induces large-scale transcriptional alterations leading to general repression of genes required for rapid growth, such as rRNA genes, and concomitant activation of genes involved in amino acid biosynthesis, nutrient acquisition, and stress survival. In addition to transcriptional control, (p)ppGpp has been shown to directly inhibit the activity of several enzymes, including DNA primase, translation factors, and enzymes involved in GTP biosynthesis (6) (Fig. 1). Ultimately, the SR reallocates cellular resources toward adaptation to a semidormant state, facilitating survival under unfavorable conditions (5, 7). Although initially defined as a response to amino acid and carbon starvation, the term SR has since been expanded to include any regulatory effect exerted by cellular (p)ppGpp accumulation irrespective of the triggering mechanism (4).The broad physiological alterations induced by (p)ppGpp accumulation rely heavily upon transcriptional alterations. In Gammaproteobacteria, such as the Gram-negative paradigmatic orga...

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Section: Figcontrasting

confidence: 55%

Section: (P)ppgpp and Codymentioning

confidence: 99%

Many Means to a Common End: the Intricacies of (p)ppGpp Metabolism and Its Control of Bacterial Homeostasis

2015

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“…Mutations in sarA decrease S. aureus biofilm and increase antibiotic susceptibility both in vitro and in vivo (31,57), whereas mutations in mecA, which encodes penicillin binding protein 2a, impede resistance to ␤-lactam antibiotics (58). Several other genes identified in our screen have also been associated with biofilm formation (codY), ␤-lactam resistance (fmtC and vraR), or both processes (rsbU, rsbW, fmtA, and rsh) (4,10,16,(33)(34)(35)(36)(37)(38)(39). In addition, two genes (ilvE and fmtA) were previously shown to be induced by cell wall stress (36), and four genes (sarA, atl, codY, and rsbU) were specifically shown to limit biofilm in strain USA300 (38).…”

Section: Discussionmentioning

confidence: 99%

“…6C). Several of these genes have previously been shown to play a role in S. aureus ␤-lactam resistance (fmtA, fmtC, rsh, and vraR), biofilm formation (codY, rsbU, and rsbW), or both processes (atl, mecA, and sarA) (4,10,16,(33)(34)(35)(36)(37)(38)(39). Two genes (ilvE and fmtA) were previously shown to be induced by cell wall stress (36), and one gene (atl) was previously shown to be required for induction of biofilm by sub-MIC methicillin in strain LAC (16).…”

Section: Resultsmentioning

confidence: 99%

Effects of Low-Dose Amoxicillin on Staphylococcus aureus USA300 Biofilms

Mlynek

Callahan

Shimkevitch

et al. 2016

Antimicrob Agents Chemother

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cPrevious studies showed that sub-MIC levels of ␤-lactam antibiotics stimulate biofilm formation in most methicillin-resistant Staphylococcus aureus (MRSA) strains. Here, we investigated this process by measuring the effects of sub-MIC amoxicillin on biofilm formation by the epidemic community-associated MRSA strain USA300. We found that sub-MIC amoxicillin increased the ability of USA300 cells to attach to surfaces and form biofilms under both static and flow conditions. We also found that USA300 biofilms cultured in sub-MIC amoxicillin were thicker, contained more pillar and channel structures, and were less porous than biofilms cultured without antibiotic. Biofilm formation in sub-MIC amoxicillin correlated with the production of extracellular DNA (eDNA). However, eDNA released by amoxicillin-induced cell lysis alone was evidently not sufficient to stimulate biofilm. Sub-MIC levels of two other cell wall-active agents with different mechanisms of action-D-cycloserine and fosfomycin-also stimulated eDNA-dependent biofilm, suggesting that biofilm formation may be a mechanistic adaptation to cell wall stress. Screening a USA300 mariner transposon library for mutants deficient in biofilm formation in sub-MIC amoxicillin identified numerous known mediators of S. aureus ␤-lactam resistance and biofilm formation, as well as novel genes not previously associated with these phenotypes. Our results link cell wall stress and biofilm formation in MRSA and suggest that eDNA-dependent biofilm formation by strain USA300 in low-dose amoxicillin is an inducible phenotype that can be used to identify novel genes impacting MRSA ␤-lactam resistance and biofilm formation.

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“…The genome of S. aureus also encodes two other monofunctional synthetases, RelP and RelQ, and transcription of these genes increases when cells are exposed to cell wall-targeting antimicrobials (20,21). Recent work on S. aureus has shown that the ability to switch on the stringent response is essential for its virulence and is required for the organism to cause chronic infections (22)(23)(24)(25).…”

mentioning

confidence: 99%

ppGpp negatively impacts ribosome assembly affecting growth and antimicrobial tolerance in Gram-positive bacteria

Corrigan

Bellows

Wood

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

185

227

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The stringent response is a survival mechanism used by bacteria to deal with stress. It is coordinated by the nucleotides guanosine tetraphosphate and pentaphosphate [(p)ppGpp], which interact with target proteins to promote bacterial survival. Although this response has been well characterized in proteobacteria, very little is known about the effectors of this signaling system in Grampositive species. Here, we report on the identification of seven target proteins for the stringent response nucleotides in the Grampositive bacterium Staphylococcus aureus. We demonstrate that the GTP synthesis enzymes HprT and Gmk bind with a high affinity, leading to an inhibition of GTP production. In addition, we identified five putative GTPases-RsgA, RbgA, Era, HflX, and ObgE-as (p)ppGpp target proteins. We show that RsgA, RbgA, Era, and HflX are functional GTPases and that their activity is promoted in the presence of ribosomes but strongly inhibited by the stringent response nucleotides. By characterizing the function of RsgA in vivo, we ascertain that this protein is involved in ribosome assembly, with an rsgA deletion strain, or a strain inactivated for GTPase activity, displaying decreased growth, a decrease in the amount of mature 70S ribosomes, and an increased level of tolerance to antimicrobials. We additionally demonstrate that the interaction of ppGpp with cellular GTPases is not unique to the staphylococci, as homologs from Bacillus subtilis and Enterococcus faecalis retain this ability. Taken together, this study reveals ribosome inactivation as a previously unidentified mechanism through which the stringent response functions in Gram-positive bacteria.ribosome | stringent response | tolerance | ppGpp | Staphylococcus aureus

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Role of the (p)ppGpp Synthase RSH, a RelA/SpoT Homolog, in Stringent Response and Virulence ofStaphylococcus aureus

Cited by 128 publications

References 58 publications

Many Means to a Common End: the Intricacies of (p)ppGpp Metabolism and Its Control of Bacterial Homeostasis

Many Means to a Common End: the Intricacies of (p)ppGpp Metabolism and Its Control of Bacterial Homeostasis

Effects of Low-Dose Amoxicillin on Staphylococcus aureus USA300 Biofilms

ppGpp negatively impacts ribosome assembly affecting growth and antimicrobial tolerance in Gram-positive bacteria

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