Effects of the ERES Pathogenicity Region Regulator Ralp3 on Streptococcus pyogenes Serotype M49 Virulence Factor Expression

Siemens, Nikolai; Fiedler, Tomas; Normann, Jana; Klein, Johannes; Münch, Richard; Patenge, Nadja; Kreikemeyer, Bernd

doi:10.1128/jb.00227-12

Cited by 23 publications

(26 citation statements)

References 45 publications

(70 reference statements)

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“…In contrast, Siemens et al identified only 21 carbon sources with Biolog that supported the metabolism (defined as Ն10% of metabolism on glucose) of the M49 GAS "generalist" strain 591, a strain that is able to colonize both the throat and skin (48). Out of these 21 carbon sources, all but 5 (Tween 40, Tween 80, pectin, mannan, and gelatin) were in common with our studies in M1T1 (Table 3).…”

Section: Discussioncontrasting

confidence: 34%

The Phosphoenolpyruvate Phosphotransferase System in Group A Streptococcus Acts To Reduce Streptolysin S Activity and Lesion Severity during Soft Tissue Infection

Gera

Jamin

et al. 2014

Infect Immun

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bObtaining essential nutrients, such as carbohydrates, is an important process for bacterial pathogens to successfully colonize host tissues. The phosphoenolpyruvate phosphotransferase system (PTS) is the primary mechanism by which bacteria transport sugars and sense the carbon state of the cell. The group A streptococcus (GAS) is a fastidious microorganism that has adapted to a variety of niches in the human body to elicit a wide array of diseases. A ⌬ptsI mutant (enzyme I [EI] deficient) generated in three different strains of M1T1 GAS was unable to grow on multiple carbon sources (PTS and non-PTS). Complementation with ptsI expressed under its native promoter in single copy was able to rescue the growth defect of the mutant. In a mouse model of GAS soft tissue infection, all ⌬ptsI mutants exhibited a significantly larger and more severe ulcerative lesion than mice infected with the wild type. Increased transcript levels of sagA and streptolysin S (SLS) activity during exponential-phase growth was observed. We hypothesized that early onset of SLS activity would correlate with the severity of the lesions induced by the ⌬ptsI mutant. In fact, infection of mice with a ⌬ptsI sagB double mutant resulted in a lesion comparable to that of either the wild type or a sagB mutant alone. Therefore, a functional PTS is not required for subcutaneous skin infection in mice; however, it does play a role in coordinating virulence factor expression and disease progression.T he ability to obtain essential nutrients during an infection is critical for bacterial pathogens to successfully colonize and proliferate within host tissues. One such key process involves the ability to import and catabolize optimal carbon sources such as carbohydrates. Bacteria have evolved elegant regulatory pathways that detect the presence of preferred carbohydrates, repress the utilization of nonpreferred sugars, and regulate their metabolism based on this information flow (1, 2). In fact, many pathogens tightly control the genes involved in carbohydrate utilization and regulation in response to in vivo growth, and these same genes have been shown to be important to the disease process (3-8). Therefore, it is apparent that bacterial pathogens have closely linked their sugar metabolic sensing networks to virulence gene expression during infection.The primary bacterial system coupling the transport of carbohydrates across the cytoplasmic membrane with their phosphorylation is a multiprotein phosphorelay called the phosphoenolpyruvate (PEP)-dependent phosphotransferase system (PTS) (9). The PTS is composed of at least three distinct proteins: the cytosolic proteins enzyme I (EI) and Hpr, encoded by ptsI and ptsH, respectively, and the membrane-bound sugar-specific enzyme II (EII) proteins. Each EII consists of one or two integral membranebound domains (EIIC/EIID) that are necessary for sugar translocation, and these domains form complexes with two hydrophilic components (EIIA and EIIB) that are required for substrate phosphorylation (1, 10). EI initiates ...

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Section: Discussioncontrasting

confidence: 34%

The Phosphoenolpyruvate Phosphotransferase System in Group A Streptococcus Acts To Reduce Streptolysin S Activity and Lesion Severity during Soft Tissue Infection

Gera

Jamin

et al. 2014

Infect Immun

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“…Interestingly, the antiphagocytic hyaluronic acid capsule (hasA operon) was also found in our screen, but only in donor A (see Table S2 in the supplemental material). We identified the standalone regulator RofA-like protein 3 (ralp3, Spy0559), which was recently shown to have a role in GAS survival in human blood and plasma (39). The gene perR (Spy0161) encoding the peroxide resistance regulator was found conserved in all donor bloods and has been shown to be important for GAS fitness within the host and necessary for resistance to phagocytic killing in human blood (32).…”

Section: Discussionmentioning

confidence: 99%

Genome-Wide Identification of Genes Required for Fitness of Group A Streptococcus in Human Blood

et al. 2013

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The group A streptococcus (GAS) is a strict human pathogen responsible for a wide spectrum of diseases. Although GAS genome sequences are available, functional genomic analyses have been limited. We developed a mariner-based transposon, osKaR, designed to perform Transposon-Site Hybridization (TraSH) in GAS and successfully tested its use in several invasive serotypes. A complex osKaR mutant library in M1T1 GAS strain 5448 was subjected to negative selection in human blood to identify genes important for GAS fitness in this clinically relevant environment. Mutants underrepresented after growth in blood (output pool) compared to growth in rich media (input pool) were identified using DNA microarray hybridization of transposon-specific tags en masse. Using blood from three different donors, we identified 81 genes that met our criteria for reduced fitness in blood from at least two individuals. Genes known to play a role in survival of GAS in blood were found, including those encoding the virulence regulator Mga (mga), the peroxide response regulator PerR (perR), and the RofA-like regulator Ralp-3 (ralp3). We also identified genes previously reported for their contribution to sepsis in other pathogens, such as de novo nucleotide synthesis (purD, purA, pyrB, carA, carB, guaB), sugar metabolism (scrB, fruA), zinc uptake (adcC), and transcriptional regulation (cpsY). To validate our findings, independent mutants with mutations in 10 different genes identified in our screen were confirmed to be defective for survival in blood bactericidal assays. Overall, this work represents the first use of TraSH in GAS to identify potential virulence genes.

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“…Since the regulatory network of GAS M49 was published, it is known that Ralp3 regulates genes with metabolic function, especially both lac and fru operons [17]. To compare the ability of different serotypes to utilize sugars, growth experiments in CDM batch cultures with lactose and fructose as sole carbon source were performed.…”

Section: Resultsmentioning

confidence: 99%

“…The GAS serotype M1 genome sequence [23] revealed the existence of two additional Ralp regulators, Ralp3 and Ralp4 [24]. Both were partially characterized [14], [17], [25]. Kreikemeyer and colleagues exclusively found ralp3 homologues genes in the genomes of GAS serotypes M1, M4, M12, M28, and M49 [14].…”

Section: Discussionmentioning

confidence: 99%

“…The complete eno ralp3 epf sagA region was called ERES pathogenicity island [14]. Deletion of Ralp3 in GAS M49 reduced the adherence to and internalization into human keratinocytes, severely attenuated survival of the mutant in human blood and serum, reduced the plasminogen-binding capacity, decreased the secretion of the important virulence factor SpeB, and finally downregulated expression of genes encoding proteins important for metabolism of fructose and lactose [17]. Together that study suggested a rather beneficial activity of Ralp3 expression for many virulence attributes and also metabolism of the GAS M49 serotype.…”

Section: Introductionmentioning

confidence: 99%

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Heterologous Expression of Ralp3 in Streptococcus pyogenes M2 and M6 Strains Affects the Virulence Characteristics

Siemens

Kreikemeyer

2013

PLoS ONE

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BackgroundRalp3 is a transcriptional regulator present in a serotype specific fashion on the chromosome of the human pathogen Streptococcus pyogenes (group A streptococci, GAS). In serotypes harbouring the ralp3 gene either positive or negative effects on important metabolic and virulence genes involved in colonization and immune evasion in the human host were observed. A previous study revealed that deletion of ralp3 in a GAS M49 serotype significantly attenuated many virulence traits and caused metabolic disadvantages. This leads to two questions: (i) which kind of consequences could Ralp3 expression have in GAS serotypes naturally lacking this gene, and (ii) is Ralp3 actively lost during evolution in these serotypes.Methodology/Principal FindingsWe investigated the role of Ralp3 in GAS M2 and M6 pathogenesis. Both serotypes lack ralp3 on their chromosome. The heterologous expression of ralp3 in both serotypes resulted in reduced attachment to and internalization into the majority of tested epithelial cells. Both ralp3 expression strains showed a decreased ability to survive in human blood and exclusively M2::ralp3 showed decreased survival in human serum. Both mutants secreted more active SpeB in the supernatant, resulting in a higher activity compared to wild type strains. The respective M2 and M6 wild type strains outcompeted the ralp3 expression strains in direct metabolic competition assays. The phenotypic changes observed in the M2:ralp3 and M6:ralp3 were verified on the transcriptional level. Consistent with the virulence data, tested genes showed transcript level changes in the same direction.Conclusions/SignificanceTogether these data suggest that Ralp3 can take over transcriptional control of virulence genes in serotypes lacking the ralp3 gene. Those serotypes most likely lost Ralp3 during evolution since obviously expression of this gene is disadvantageous for metabolism and pathogenesis.

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Effects of the ERES Pathogenicity Region Regulator Ralp3 on Streptococcus pyogenes Serotype M49 Virulence Factor Expression

Cited by 23 publications

References 45 publications

The Phosphoenolpyruvate Phosphotransferase System in Group A Streptococcus Acts To Reduce Streptolysin S Activity and Lesion Severity during Soft Tissue Infection

The Phosphoenolpyruvate Phosphotransferase System in Group A Streptococcus Acts To Reduce Streptolysin S Activity and Lesion Severity during Soft Tissue Infection

Genome-Wide Identification of Genes Required for Fitness of Group A Streptococcus in Human Blood

Heterologous Expression of Ralp3 in Streptococcus pyogenes M2 and M6 Strains Affects the Virulence Characteristics

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