Role of BvgA phosphorylation and DNA binding affinity in control of Bvg-mediated phenotypic phase transition in Bordetella pertussis

Department of Molecular, Cellular and Developmental Biology, University of California, Santa Barbara, CA 93109-9610, USA.
Molecular Microbiology (Impact Factor: 4.42). 12/2005; 58(3):700-13. DOI: 10.1111/j.1365-2958.2005.04875.x
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To investigate the mechanism by which the Bordetella BvgAS phosphorelay controls expression of at least three distinct phenotypic phases, we isolated and characterized two B. pertussis mutants that were able to express Bvg- and Bvg(i) phase phenotypes but not Bvg+ phase phenotypes. In both cases, the mutant phenotype was due to a single nucleotide change in bvgA resulting in a single amino acid substitution in BvgA. In vitro phosphorylation assays showed that BvgA containing the T194M substitution was significantly impaired in its ability to use either BvgS or acetyl phosphate as a substrate for phosphorylation. Binding studies indicated that this mutant protein was able to bind an oligonucleotide containing a high-affinity BvgA binding site in a manner similar to wild-type BvgA, but was defective for binding the fhaB promoter in the absence of RNA polymerase (RNAP). By contrast, BvgA containing the R152H substitution had wild-type phosphorylation properties but was severely defective in its ability to bind either the high-affinity BvgA binding site-containing oligonucleotide or the fhaB promoter by itself. Both mutant BvgA proteins were able to bind the fhaB promoter in the presence of RNAP however, demonstrating the profound effect that RNAP has on stabilizing the ternary complexes between promoter DNA, BvgA and RNAP. Our results are consistent with the hypothesis that BvgAS controls expression of multiple phenotypic phases by adjusting the intracellular concentration of BvgA-P and they demonstrate the additive nature of BvgA binding site affinity and protein-protein interactions at different Bvg-regulated promoters.

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Available from: Allison M Jones, Dec 16, 2014
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    • "The expression of B. pertussis virulence factors is controlled by the two-component BvgAS sensory transduction system [1], [15], [16]. BvgAS controls the expression of a spectrum of phenotypic phases transitioning between a virulent (Bvg+) phase and a nonvirulent (Bvg-) phase, which is referred to as phenotypic modulation [17]–[20]. During the virulent Bvg+ phase, the BvgAS system controls the expression of over 100 virulence genes [21] by binding phosphorylated BvgA to the promoters of the virulence genes. "
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    ABSTRACT: Bordetella pertussis (B. pertussis) is the causative agent of whooping cough, which is a highly contagious disease in the human respiratory tract. Despite vaccination since the 1950s, pertussis remains the most prevalent vaccine-preventable disease in developed countries. A recent resurgence pertussis is associated with the expansion of B. pertussis strains with a novel allele for the pertussis toxin (ptx) promoter ptxP3 in place of resident ptxP1 strains. The recent expansion of ptxP3 strains suggests that these strains carry mutations that have increased their fitness. Compared to the ptxP1 strains, ptxP3 strains produce more Ptx, which results in increased virulence and immune suppression. In this study, we investigated the contribution of gene expression changes of various genes on the increased fitness of the ptxP3 strains. Using genome-wide gene expression profiling, we show that several virulence genes had higher expression levels in the ptxP3 strains compared to the ptxP1 strains. We provide the first evidence that wildtype ptxP3 strains are better colonizers in an intranasal mouse infection model. This study shows that the ptxP3 mutation and the genetic background of ptxP3 strains affect fitness by contributing to the ability to colonize in a mouse infection model. These results show that the genetic background of ptxP3 strains with a higher expression of virulence genes contribute to increased fitness.
    PLoS ONE 06/2013; 8(6):e66150. DOI:10.1371/journal.pone.0066150 · 3.23 Impact Factor
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    • "This finding suggests a possible two-component signal transduction system, where the periplasmic sensor histidine kinase is responsible for sensing stimuli and a second component regulates the virulence effector, namely the TAA gene (Figure 5). The bacterial prototype for this system is the Bordetella pertussis BvgAS two-component regulatory system which is involved in the expression of many adhesins and toxins (Jones et al., 2005). It is now important to obtain experimental data in order to validate the hypothesis raised by this in silico analysis. "
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    ABSTRACT: Trimeric autotransporter adhesins (TAAs) are multimeric surface proteins exclusively found in bacteria. They are involved in various biological traits of pathogenic Gram-negative bacteria including adherence, biofilm formation, invasion, survival within eukaryotic cells, serum resistance, and cytotoxicity. TAAs have a modular architecture composed by a conserved membrane-anchored C-terminal domain and a variable number of stalk and head domains. In this study, a bioinformatic approach has been used to analyze the distribution and architecture of TAAs among Burkholderia cepacia complex (Bcc) genomes. Fifteen genomes were probed revealing a total of 74 encoding sequences. Compared with other bacterial species, the Bcc genomes contain a large number of TAAs (two genes to up to eight genes, such as in B. cenocepacia). Phylogenetic analysis showed that the TAAs grouped into at least eight distinct clusters. TAAs with serine-rich repeats are clearly well separated from others, thereby representing a different evolutionary lineage. Comparative gene mapping across Bcc genomes reveals that TAA genes are inserted within conserved synteny blocks. We further focused our analysis on the epidemic strain B. cenocepacia J2315 in which seven TAAs were annotated. Among these, three TAA-encoding genes (BCAM019, BCAM0223, and BCAM0224) are organized into a cluster and are candidates for multifunctional virulence factors. Here we review the current insights into the functional role of BCAM0224 as a model locus.
    Frontiers in Cellular and Infection Microbiology 12/2011; 1:13. DOI:10.3389/fcimb.2011.00013 · 3.72 Impact Factor
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    • "A large proportion of Bordetella genes undergo a change in their expression during the infectious cycle [23-25] some of which correspond to phenotypic modulation under the control of a two-component system named BvgA/BvgS (reviewed in [26]). In this system BvgS acts as a sensor anchored in the inner membrane and BvgA as the transcriptional activator [27,28]. "
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    ABSTRACT: Small bacterial RNAs (sRNAs) have been shown to participate in the regulation of gene expression and have been identified in numerous prokaryotic species. Some of them are involved in the regulation of virulence in pathogenic bacteria. So far, little is known about sRNAs in Bordetella, and only very few sRNAs have been identified in the genome of Bordetella pertussis, the causative agent of whooping cough. An in silico approach was used to predict sRNAs genes in intergenic regions of the B. pertussis genome. The genome sequences of B. pertussis, Bordetella parapertussis, Bordetella bronchiseptica and Bordetella avium were compared using a Blast, and significant hits were analyzed using RNAz. Twenty-three candidate regions were obtained, including regions encoding the already documented 6S RNA, and the GCVT and FMN riboswitches. The existence of sRNAs was verified by Northern blot analyses, and transcripts were detected for 13 out of the 20 additional candidates. These new sRNAs were named Bordetella pertussis RNAs, bpr. The expression of 4 of them differed between the early, exponential and late growth phases, and one of them, bprJ2, was found to be under the control of BvgA/BvgS two-component regulatory system of Bordetella virulence. A phylogenetic study of the bprJ sequence revealed a novel, so far undocumented repeat of ~90 bp, found in numerous copies in the Bordetella genomes and in that of other Betaproteobacteria. This repeat exhibits certain features of mobile elements. We shown here that B. pertussis, like other pathogens, expresses sRNAs, and that the expression of one of them is controlled by the BvgA/BvgS system, similarly to most virulence genes, suggesting that it is involved in virulence of B. pertussis.
    BMC Genomics 04/2011; 12:207. DOI:10.1186/1471-2164-12-207 · 3.99 Impact Factor
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