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Analyzing the Transcriptomes of Two Quorum-Sensing Controlled Transcription Factors, RcsA and LrhA, Important for Pantoea stewartii Virulence.

ABSTRACT: The Gram-negative proteobacterium Pantoea stewartii subsp. stewartii causes wilt disease in corn plants. Wilting is primarily due to bacterial exopolysaccharide (EPS) production that blocks water transport in the xylem during the late stages of infection. EsaR, the master quorum-sensing (QS) regulator in P. stewartii, modulates EPS levels. At low cell densities EsaR represses or activates expression of a number of genes in the absence of its acyl homoserine lactone (AHL) ligand. At high cell densities, binding of AHL inactivates EsaR leading to derepression or deactivation of its direct targets. Two of these direct targets are the key transcription regulators RcsA and LrhA, which in turn control EPS production and surface motility/adhesion, respectively. In this study, RNA-Seq was used to further examine the physiological impact of deleting the genes encoding these two second-tier regulators. Quantitative reverse transcription PCR (qRT-PCR) was used to validate the regulation observed in the RNA-Seq data. A GFP transcriptional fusion reporter confirmed the existence of a regulatory feedback loop in the system between LrhA and RcsA. Plant virulence assays carried out with rcsA and lrhA deletion and complementation strains demonstrated that both transcription factors play roles during establishment of wilt disease in corn. These efforts further define the hierarchy of the QS-regulated network controlling plant virulence in P. stewartii.

SUBMITTER: Kernell Burke A

PROVIDER: S-EPMC4689408 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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Analyzing the Transcriptomes of Two Quorum-Sensing Controlled Transcription Factors, RcsA and LrhA, Important for Pantoea stewartii Virulence.

Kernell Burke Alison A Duong Duy An DA Jensen Roderick V RV Stevens Ann M AM

PloS one 20151223 12

The Gram-negative proteobacterium Pantoea stewartii subsp. stewartii causes wilt disease in corn plants. Wilting is primarily due to bacterial exopolysaccharide (EPS) production that blocks water transport in the xylem during the late stages of infection. EsaR, the master quorum-sensing (QS) regulator in P. stewartii, modulates EPS levels. At low cell densities EsaR represses or activates expression of a number of genes in the absence of its acyl homoserine lactone (AHL) ligand. At high cell den ...[more]

PMID: 26699719

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Project description:Introduction: Pantoea stewartii subsp. stewartii is a proteobacterium that causes Stewart’s wilt disease in corn plants. The bacteria form a biofilm in the xylem of infected plants and produce capsule that blocks water transport, eventually causing wilt. At low cell densities, the quorum-sensing (QS) regulatory protein EsaR is known to directly repress expression of esaR itself as well as the capsular synthesis operon transcription regulator, rcsA, and a 2,5-diketogluconate reductase, dkgA. It simultaneously directly activates expression of genes for a putative small RNA, esaS, the glycerol utilization operon, glpFKX, and another transcriptional regulator, lrhA. At high bacterial cell densities, all of this regulation is relieved when EsaR binds an acylated homoserine lactone signal, which is synthesized constitutively over growth. QS-dependent gene expression is critical for the establishment of disease in the plant. Objective: However, the identity of the full set of genes controlled by EsaR/QS is unknown. A proteomic approach previously identified around 30 proteins in the QS regulon. In this study, a whole-transcriptome, next-generation sequencing analysis of rRNA-depleted RNA from QS-proficient and -deficient P. stewartii strains was performed to identify additional targets of EsaR. Result: EsaR-dependent transcriptional regulation of a subset of differentially expressed genes was confirmed by quantitative reverse transcription-PCR (qRT-PCR). Electrophoretic mobility shift assays demonstrated that EsaR directly bound 10 newly identified target promoters. Overall, the QS regulon of P. stewartii orchestrates three major physiological responses: capsule and cell envelope biosynthesis, surface motility and adhesion, and stress response. Comparison of gene expression in two strains of Pantoea stewartii, QS-proficient DC283 ES∆IR (pSVB60) and QS-deficient DC283 ES∆IR (pBBR1MCS-3) to identify those genes that are directly regulated by the master Quorum-sensing regulator, EsaR.

Dataset Information

Analyzing the Transcriptomes of Two Quorum-Sensing Controlled Transcription Factors, RcsA and LrhA, Important for Pantoea stewartii Virulence.

Publications

Analyzing the Transcriptomes of Two Quorum-Sensing Controlled Transcription Factors, RcsA and LrhA, Important for Pantoea stewartii Virulence.

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