MglA/SspA Complex Interactions Are Modulated by Inorganic Polyphosphate

Wrench, Algevis; Gardner, C. L.; Siegel, Sara D.; Pagliai, Fernando A.; Malekiha, Mahsa; González, Claudio F.; Lorca, Graciela L.

doi:10.1371/journal.pone.0076428

Cited by 16 publications

(17 citation statements)

References 53 publications

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“…Previous work suggested that ppGpp regulates the same set of genes as that regulated by MglA, SspA, and PigR and promotes the interaction between the MglASspA complex and PigR in F. tularensis (21). Another recent study found that polyphosphate binds to the MglA-SspA complex in vitro (33). It could be that ppGpp or polyphosphate interacts directly with the MglA-SspA complex to promote the interaction with PigR and that one or more of the mutants is defective for binding one of these molecules.…”

Section: Discussionmentioning

confidence: 97%

“…MglA(Y11A), MglA(T47A), MglA(P48S), MglA(Y63A), MglA(R64A), MglA(K101E), SspA (K65E), SspA(V105E), and SspA(L130S) were defective for the interaction with PigR in the E. coli bridge-hybrid assay; therefore, if these substitutions do disrupt binding of another transcription factor, this factor must be conserved in E. coli. The molecules guanosine tetraphosphate (ppGpp) and polyphosphate have been proposed to play a role in virulence gene regulation along with MglA, SspA, and PigR (21,33,34). Previous work suggested that ppGpp regulates the same set of genes as that regulated by MglA, SspA, and PigR and promotes the interaction between the MglASspA complex and PigR in F. tularensis (21).…”

Section: Discussionmentioning

confidence: 99%

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Coordinate Control of Virulence Gene Expression in Francisella tularensis Involves Direct Interaction between Key Regulators

Rohlfing

Dove

2014

J Bacteriol

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In Francisella tularensis, the putative DNA-binding protein PigR works in concert with the SspA protein family members MglA and SspA to control the expression of genes that are essential for the intramacrophage growth and survival of the organism. MglA and SspA form a complex that interacts with RNA polymerase (RNAP), and this interaction between the MglA-SspA complex and RNAP is thought to be critical to its regulatory function. How PigR works in concert with the MglA-SspA complex is not known; previously published findings differ over whether PigR interacts with the MglA-SspA complex, leading to disparate models for how PigR and the MglA-SspA complex exert their regulatory effects. Here, using a combination of genetic assays, we identify mutants of MglA and SspA that are specifically defective for interaction with PigR. Analysis of the MglA and SspA mutants in F. tularensis reveals that interaction between PigR and the MglA-SspA complex is essential in order for PigR to work coordinately with MglA and SspA to positively regulate the expression of virulence genes. Our findings uncover a surface of the MglASspA complex that is important for interaction with PigR and support the idea that PigR exerts its regulatory effects through an interaction with the RNAP-associated MglA-SspA complex.

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Section: Discussionmentioning

confidence: 97%

Section: Discussionmentioning

confidence: 99%

Coordinate Control of Virulence Gene Expression in Francisella tularensis Involves Direct Interaction between Key Regulators

Rohlfing

Dove

2014

J Bacteriol

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“…polyphosphate hexametaphosphate (hexaMP), binds the MglA-SspA complex (Wrench et al 2013b). However, we found that hexaMP did not compete with ppGpp binding to MglA-SspA (Supplemental Fig.…”

Section: Mgla-sspa Interacts Directly and Specifically With Ppgppmentioning

confidence: 99%

Dissection of the molecular circuitry controlling virulence in Francisella tularensis

et al. 2017

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Francisella tularensis, the etiological agent of tularemia, is one of the most infectious bacteria known. Because of its extreme pathogenicity, F. tularensis is classified as a category A bioweapon by the US government. F. tularensis virulence stems from genes encoded on the Francisella pathogenicity island (FPI). An unusual set of Francisella regulators—the heteromeric macrophage growth locus protein A (MglA)–stringent starvation protein A (SspA) complex and the DNA-binding protein pathogenicity island gene regulator (PigR)—activates FPI transcription and thus is essential for virulence. Intriguingly, the second messenger, guanosine–tetraphosphate (ppGpp), which is produced during infection, is also involved in coordinating Francisella virulence; however, its role has been unclear. Here we identify MglA–SspA as a novel ppGpp-binding complex and describe structures of apo- and ppGpp-bound MglA–SspA. We demonstrate that MglA–SspA, which binds RNA polymerase (RNAP), also interacts with the C-terminal domain of PigR, thus anchoring the (MglA–SspA)–RNAP complex to the FPI promoter. Furthermore, we show that MglA–SspA must be bound to ppGpp to mediate high-affinity interactions with PigR. Thus, these studies unveil a novel pathway different from those described previously for regulation of transcription by ppGpp. The data also indicate that F. tularensis pathogenesis is controlled by a highly interconnected molecular circuitry in which the virulence machinery directly senses infection via a small molecule stress signal.

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“…(p)ppGpp promotes the interaction between PigR and the MglA-SspA complex in E. coli. Prior work suggested that in F. tularensis (p)ppGpp exerts its effects on virulence gene expression by promoting the accumulation of polyphosphate, which in turn, is required for MglA and SspA to form a complex (50). To test whether or not the interaction between MglA and SspA depends (directly or indirectly) on the ability of cells to synthesize (p)ppGpp, we took advantage of a two-hybrid assay we had used previously to detect the interaction between MglA and SspA in E. coli (11,16,51).…”

Section: Resultsmentioning

confidence: 99%

“…Studies in F. tularensis suggest that (p)ppGpp promotes the interaction between PigR and the RNAP-associated MglA-SspA complex (13), and recent work has revealed that ppGpp can bind directly to the MglA-SspA complex to mediate its effects (15). However, other recent findings suggested that (p)ppGpp might exert its effects on virulence gene expression indirectly by promoting the accumulation of polyphosphate, which in turn was required for MglA and SspA to form a complex (50). Here we present evidence that polyphosphate is not required in order for MglA and SspA to interact.…”

mentioning

confidence: 99%

Polyphosphate Kinase Antagonizes Virulence Gene Expression in Francisella tularensis

2018

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The alarmone ppGpp is a critical regulator of virulence gene expression in In this intracellular pathogen, ppGpp is thought to work in concert with the putative DNA-binding protein PigR and the SspA protein family members MglA and SspA to control a common set of genes. MglA and SspA form a complex that interacts with RNA polymerase (RNAP), and PigR functions by interacting with the RNAP-associated MglA-SspA complex. Prior work suggested that ppGpp indirectly exerts its regulatory effects in by promoting the accumulation of polyphosphate in the cell, which in turn was required for formation of the MglA-SspA complex. Here we show that in , neither polyphosphate nor ppGpp is required for formation of the MglA-SspA complex but that ppGpp promotes the interaction between PigR and the MglA-SspA complex. Moreover, we show that polyphosphate kinase, the enzyme responsible for the synthesis of polyphosphate, antagonizes virulence gene expression in, a finding that is inconsistent with the notion that polyphosphate accumulation promotes virulence gene expression in this organism. Our findings identify polyphosphate kinase as a novel negative regulator of virulence gene expression in and support a model in which ppGpp exerts its positive regulatory effects by promoting the interaction between PigR and the MglA-SspA complex. In , MglA and SspA form a complex that associates with RNA polymerase to positively control the expression of key virulence genes. The MglA-SspA complex works together with the putative DNA-binding protein PigR and the alarmone ppGpp. PigR functions by interacting directly with the MglA-SspA complex, but how ppGpp exerts its effects was unclear. Prior work indicated that ppGpp acts by promoting the accumulation of polyphosphate, which is required for MglA and SspA to interact. Here we show that formation of the MglA-SspA complex does not require polyphosphate. Furthermore, we find that polyphosphate antagonizes the expression of virulence genes in Thus, ppGpp does not promote virulence gene expression in this organism through an effect on polyphosphate.

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MglA/SspA Complex Interactions Are Modulated by Inorganic Polyphosphate

Cited by 16 publications

References 53 publications

Coordinate Control of Virulence Gene Expression in Francisella tularensis Involves Direct Interaction between Key Regulators

Coordinate Control of Virulence Gene Expression in Francisella tularensis Involves Direct Interaction between Key Regulators

Dissection of the molecular circuitry controlling virulence in Francisella tularensis

Polyphosphate Kinase Antagonizes Virulence Gene Expression in Francisella tularensis

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