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CovS Inactivation Reduces CovR Promoter Binding at Diverse Virulence Factor Encoding Genes in Group A Streptococcus

ABSTRACT: CovS Inactivation Reduces CovR Promoter Binding at Diverse Virulence Factor Encoding Genes in Group A Streptococcus

PROVIDER: PRJNA793225 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR17382743.fastq.gz	Fastqsanger.gz
	SRR17382744.fastq.gz	Fastqsanger.gz
	SRR17382745.fastq.gz	Fastqsanger.gz
	SRR17382746.fastq.gz	Fastqsanger.gz
	SRR17382747.fastq.gz	Fastqsanger.gz

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CovS Inactivation Reduces CovR Promoter Binding at Diverse Virulence Factor Encoding Genes in Group A Streptococcus

Project description:We performed CovR ChIP-seq analysis in the emm1 strain MGAS2221 and its CovS kinase deficient derivative strain 2221-CovS-E281A. We identified that CovR bound in the promoter regions of nearly all virulence factor encoding genes in the CovR regulon. Additionally, direct CovR binding was observed for numerous genes encoding proteins involved in amino acid metabolism, but we found limited direct CovR binding to genes encoding other transcriptional regulators. The consensus sequence AATRANAAAARVABTAAA was present in the promoters of genes directly regulated by CovR, and mutations of highly conserved positions within this motif relieved CovR repression of the hasA and M2221_0187 promoters. Analysis of strain 2221-CovS-E281A revealed that binding of CovR at repressed, but not activated, promoters is highly dependent on CovR~P state. CovR repressed virulence factor encoding genes could be grouped dependent on how variation in CovR~P differentially impacted DNA binding and gene transcript levels.

2022-01-03 | GSE192795 | GEO

Hyperphosphorylation of the Group A Streptococcal Control of Virulence Regulator Increases Promoter Occupancy Specifically at Virulence Factor Encoding Genes

Project description:The control of virulence two-component gene regulatory system (CovRS) is critical to the pathogenesis of many medically important streptococci. In emm1 group A streptococci (GAS), CovR directly binds the promoters of numerous GAS virulence factor encoding genes. Elimination of CovS phosphatase activity increases CovR phosphorylation (CovR~P) levels and abrogates GAS virulence. Given the emm type-specific diversity of CovRS function, herein we used ChIP-seq to define global CovR DNA occupancy in the wild-type emm3 strain MGAS10870 (medium CovR~P) and its CovS phosphatase-negative derivative 10870-CovS-T284A (high CovR~P). In the wild-type emm3 strain, 89% of the previously identified emm1 CovR binding sites present in the emm3 genome were also enriched; additionally, we ascertained unique CovR binding, primarily to genes in mobile genetic elements and other sites of inter-strain chromosomal differences. Elimination of phosphatase activity specifically increased CovR occupancy at the promoters of a broad array of CovR repressed virulence factor encoding genes, including those encoding the key GAS regulator Mga and M protein. However, a limited number of promoters had augmented enrichment at low CovR~P levels. Differential motif searches using sequences enriched at high vs. low CovR~P levels revealed two distinct binding patterns. At high CovR~P, a pseudo-palindromic AT-rich consensus sequence consistent with CovR binding as a dimer was determined. Conversely, sequences specifically enriched at low CovR~P contained isolated “ATTARA” motifs suggesting an interaction with a monomer. These data extend understanding of global CovR DNA occupancy beyond emm1 GAS and provide a mechanism for previous observations regarding hypovirulence induced by CovS phosphatase abrogation.

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Transcriptomic analysis of the major regulator of virulence CovR in Group B Streptococcus strains BM110 and NEM16.

Project description:To define the transcriptional response associated to a CovR inactivation we performed RNA-Seq in GBS strains BM110 and NEM316. We used mutants in which CovR is inactivated following a two base-pairs chromosomal substitution (AT->CC) resulting in the translation of a CovRD53A variant unable to be phosphorylated by the histidine kinase CovS. We also used a ∆covR mutant in strain BM110 in which the covR sequence is deleted from the chromosome.

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Project description:Two covR mutant derivatives of parental strain MGAS2221 were recovered from mice experimentally infected with MGAS2221 and shown to differ in terms of the number and concentration of secreted proteins. One of the covR mutant strains had a secretion phenotype identical to a covS mutant strain, while the other had a secretion phenotype identical to a constructed covR mutant strain. To further investigate the potential differences between the two covR mutant strains we performed expression microarray analysis. Single cultures of each of the four GAS strains tested were grown in THY broth to early exponential phase (O.D. 0.2). Two volumes of RNA protect were added, the samples incubated at room temperature for 5 minutes, and the bacteria collected through centrifugation. Total RNA was isolated via a mechaniscal disruption method, converted to cDNA, fragmented, labeled, and hybridized to our Affymetrix microarray. Estimates of gene expression were calculated using GCOS software v1.4. Data represent probes for serotype M1.

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2013-02-26 | E-MEXP-3703 | biostudies-arrayexpress

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Project description:We sought to determine how single amino acid changes in the active site of CovS autophosphorylation and subsequent phosphorylation of the response regulator CovR affect the gene expression profile of group A streptococcus There were 6 strains analyzed, each in quadruplicate replicates: 1) wild-type GAS; 2) covS deletion strain; 3) covS with alanine at AA 280; 4) covS with valine at AA 281; 5) covS with valine at AA 284; 6) covS with alanine at AA 457

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Effect of CovR polymorphisms on GAS gene expression

Project description:In this study we sought to determine the effects of single amino acid replacements in the control of virulence regulatory (CovR) regulatory protein on group A streptococcal global gene expression. We compared the transcriptomes of the serotype M3 strain MGAS10870 with four derivative strains: 1) a strain in which CovR had been completely inactivated (CovR knockout); 2) a strain in which the Arg 144 amino acid was changed to a cysteine (CovR-R144C); 3) a strain in which the Arg 158 amino acid was changed to a cysteine (CovR-R158C); and 4) a strain in which the the Asn 193 amino acid was changed to an isoleucine (CovR-N193I). Duplicate samples of each strain were grown to mid-exponential phase and the relative transcriptomes were compared using an Affymetrix GeneChipl.

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RNA sequencing analysis of covR-deficient Streptococcus agalactiae

Project description:Total RNA was isolated from mid-log phase Streptococcus agalactiae cells deficient in CovR (∆covR strain GU2400), grown in Todd-Hewitt broth (THB) medium and sequenced using Illumina NextSeq500

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Project description:The goal of this study were to compare the transcriptoms of CcpA and CovR in group A Streptococcus. Isogenic mutant strains were created in which CcpA and CovR were inactivated alone and in combination. The transcriptomes of the four strains (wild-type, CcpA mutant, CovR mutant, and ccpA-covR double mutant) were then compared.

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