Restoring virulence to mutants lacking subunits of multiprotein machines: functional complementation of a <i>Brucella virB5</i> mutant

Sprynski, Nicolas; Félix, Christine; O’Callaghan, David; Vergunst, Annette C.

doi:10.1016/j.fob.2012.03.003

Cited by 7 publications

(3 citation statements)

References 32 publications

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“…Complementation for growth of the non-polar rex mutant in B. cereus ATCC 14579 was not possible using a pHT304-based plasmid. This is probably due to the presence of multiple copies of the rex promoter region and/or overexpression of rex [38] . Therefore, to validate the role of Rex in aerobic respiration and anaerobic fermentation in B. cereus , we deleted the rex gene of B cereus F4430/73 [39] and cultured the mutant strain under both aerobiosis and anaerobiosis.…”

Section: Resultsmentioning

confidence: 99%

Proteomic Evidences for Rex Regulation of Metabolism in Toxin-Producing Bacillus cereus ATCC 14579

et al. 2014

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The facultative anaerobe, Bacillus cereus, causes diarrheal diseases in humans. Its ability to deal with oxygen availability is recognized to be critical for pathogenesis. The B. cereus genome comprises a gene encoding a protein with high similarities to the redox regulator, Rex, which is a central regulator of anaerobic metabolism in Bacillus subtilis and other Gram-positive bacteria. Here, we showed that B. cereus rex is monocistronic and down-regulated in the absence of oxygen. The protein encoded by rex is an authentic Rex transcriptional factor since its DNA binding activity depends on the NADH/NAD+ ratio. Rex deletion compromised the ability of B. cereus to cope with external oxidative stress under anaerobiosis while increasing B. cereus resistance against such stress under aerobiosis. The deletion of rex affects anaerobic fermentative and aerobic respiratory metabolism of B. cereus by decreasing and increasing, respectively, the carbon flux through the NADH-recycling lactate pathway. We compared both the cellular proteome and exoproteome of the wild-type and Δrex cells using a high throughput shotgun label-free quantitation approach and identified proteins that are under control of Rex-mediated regulation. Proteomics data have been deposited to the ProteomeXchange with identifier PXD000886. The data suggest that Rex regulates both the cross-talk between metabolic pathways that produce NADH and NADPH and toxinogenesis, especially in oxic conditions.

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Section: Resultsmentioning

confidence: 99%

Proteomic Evidences for Rex Regulation of Metabolism in Toxin-Producing Bacillus cereus ATCC 14579

et al. 2014

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“…S1A and B. Others have shown that improper expression of virB components including the overexpression of VirB5 can lead to B. suis 1330 attenuation (Sprynski , et al , 2012). Also, as discussed above, the expression of the VirB operon is significantly altered in the ΔBM-LOV-HK strain compared to wild type.…”

Section: Discussionmentioning

confidence: 99%

Decreasedin vivovirulence and altered gene expression by aBrucella melitensislight-sensing histidine kinase mutant

et al. 2014

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Brucella species utilize diverse virulence factors. Previously, Brucella abortus light sensing histidine kinase was identified as important for cellular infection. Here, we demonstrate that a Brucella melitensis LOV-HK (BM-LOV-HK) mutant strain has strikingly different gene expression than wild type. General stress response genes including the alternative sigma factor rpoE1 and its anti-anti sigma factor phyR were downregulated, while flagellar, quorum sensing, and type IV secretion system genes were upregulated in the ΔBM-LOV-HK strain versus wild type. Contextually, expression results agree with other studies of transcriptional regulators involving ΔrpoE1, ΔphyR, ΔvjbR, and ΔblxR (ΔbabR) Brucella strains. Additionally, deletion of BM-LOV-HK decreases virulence in mice. During C57BL/6 mouse infection, the ΔBM-LOV-HK strain had 2 logs less CFUs in the spleen 3 days post infection but similar levels 6 days post infection compared to wild type. Infection of IRF-1−/− mice more specifically define ΔBM-LOV-HK strain attenuation with fewer bacteria in spleens and significantly increased survival of mutant versus wild type infected IRF-1−/− mice. Upregulation of flagella, quorum sensing, and VirB genes, along with downregulation of rpoE1 and related sigma factor, rpoH2 (BMEI0280) suggest that BM-LOV-HK modulates both quorum sensing and general stress response regulatory components to control Brucella gene expression on a global level.

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“…The results showed that entD was overexpressed in the complemented strain (Δ entD -pHT304 entD ) compared with the wild-type strain (WT/pHT304). This probably explains why the wild-type phenotype was not restored by complementation (Sprynski et al, 2012 ).…”

Section: Resultsmentioning

confidence: 99%

Proteomics identifies Bacillus cereus EntD as a pivotal protein for the production of numerous virulence factors

Omer

Alpha-Bazin

Brunet³

et al. 2015

Front. Microbiol.

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Bacillus cereus is a Gram-positive pathogen that causes a wide variety of diseases in humans. It secretes into the extracellular milieu proteins that may contribute directly or indirectly to its virulence. EntD is a novel exoprotein identified by proteogenomics of B. cereus ATCC 14579. We constructed a ΔentD mutant and analyzed the impact of entD disruption on the cellular proteome and exoproteome isolated from early, late, and stationary-phase cultures. We identified 308 and 79 proteins regulated by EntD in the cellular proteome and the exoproteome, respectively. The contribution of these proteins to important virulence-associated functions, including central metabolism, cell structure, antioxidative ability, cell motility, and toxin production, are presented. The proteomic data were correlated with the growth defect, cell morphology change, reduced motility, and reduced cytotoxicity of the ΔentD mutant strain. We conclude that EntD is an important player in B. cereus virulence. The function of EntD and the putative EntD-dependent regulatory network are discussed. To our knowledge, this study is the first characterization of an Ent family protein in a species of the B. cereus group.

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Restoring virulence to mutants lacking subunits of multiprotein machines: functional complementation of a Brucella virB5 mutant

Cited by 7 publications

References 32 publications

Proteomic Evidences for Rex Regulation of Metabolism in Toxin-Producing Bacillus cereus ATCC 14579

Proteomic Evidences for Rex Regulation of Metabolism in Toxin-Producing Bacillus cereus ATCC 14579

Decreasedin vivovirulence and altered gene expression by aBrucella melitensislight-sensing histidine kinase mutant

Proteomics identifies Bacillus cereus EntD as a pivotal protein for the production of numerous virulence factors

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