scholarly journals ChvD, a Chromosomally Encoded ATP-Binding Cassette Transporter-Homologous Protein Involved in Regulation of Virulence Gene Expression in Agrobacterium tumefaciens

2001 ◽  
Vol 183 (11) ◽  
pp. 3310-3317 ◽  
Author(s):  
Zhenying Liu ◽  
Mark Jacobs ◽  
Dennis A. Schaff ◽  
Colleen A. McCullen ◽  
Andrew N. Binns
Keyword(s):  
Gene Expression ◽  
Agrobacterium Tumefaciens ◽  
Mutant Strain ◽  
Molecular Genetic ◽  
Constitutive Expression ◽  
Virulence Gene ◽  
Molecular Genetic Analysis ◽  
Atp Binding ◽  
Virulence Gene Expression ◽  
Atp Binding Cassette

ABSTRACT A yeast two-hybrid screen searching for chromosomally encoded proteins that interact with the Agrobacterium tumefaciensVirB8 protein was carried out. This screen identified an interaction candidate homologous to the partial sequence of a gene that had previously been identified in a transposon screen as a potential regulator of virG expression, chvD. In this report, the cloning of the entire chvD gene is described and the gene is sequenced and characterized. Insertion of a promoterless lacZ gene into the chvD locus greatly attenuated virulence and vir gene expression. Compared to that of the wild-type strain, growth of thechvD mutant was reduced in rich, but not minimal, medium. Expression of chvD, as monitored by expression of β-galactosidase activity from the chvD-lacZ fusion, occurred in both rich and minimal media as well as under conditions that induce virulence gene expression. The ChvD protein is highly homologous to a family of ATP-binding cassette transporters involved in antibiotic export from bacteria and has two complete Walker box motifs. Molecular genetic analysis demonstrated that disruption of either Walker A box, singly, does not inactivate this protein's effect on virulence but that mutations in both Walker A boxes renders it incapable of complementing a chvD mutant strain. Constitutive expression of virG in the chvDmutant strain restored virulence, supporting the hypothesis that ChvD controls virulence through effects on virG expression.

scholarly journals Involvement of Agrobacterium tumefaciens Galacturonate Tripartite ATP-Independent Periplasmic (TRAP) Transporter GaaPQM in Virulence Gene Expression

2015 ◽  
Vol 82 (4) ◽  
pp. 1136-1146 ◽  
Author(s):  
Jinlei Zhao ◽  
Andrew N. Binns
Keyword(s):  
Gene Expression ◽  
Agrobacterium Tumefaciens ◽  
Galacturonic Acid ◽  
Virulence Gene ◽  
Promoter Regions ◽  
Content Type ◽  
Virulence Gene Expression ◽  
Sugar Binding ◽  
Neutral Sugars ◽  
High Ligand

ABSTRACTMonosaccharides capable of serving as nutrients for the soil bacteriumAgrobacterium tumefaciensare also inducers of thevirregulon present in the tumor-inducing (Ti) plasmid of this plant pathogen. One such monosaccharide is galacturonate, the predominant monomer of pectin found in plant cell walls. This ligand is recognized by the periplasmic sugar binding protein ChvE, which interacts with the VirA histidine kinase that controlsvirgene expression. Although ChvE is also a member of the ChvE-MmsAB ABC transporter involved in the utilization of many neutral sugars, it is not involved in galacturonate utilization. In this study, a putative tripartite ATP-independent periplasmic (TRAP) transporter, GaaPQM, is shown to be essential for the utilization of galacturonic acid; we show that residue R169 in the predicted sugar binding site of the GaaP is required for activity. The gene upstream ofgaaPQM(gaaR) encodes a member of the GntR family of regulators. GaaR is shown to repress the expression ofgaaPQM, and the repression is relieved in the presence of the substrate for GaaPQM. Moreover, GaaR is shown to bind putative promoter regions in the sequences required for galacturonic acid utilization. Finally,A. tumefaciensstrains carrying a deletion ofgaaPQMare more sensitive to galacturonate as an inducer ofvirgene expression, while the overexpression ofgaaPQMresults in strains being less sensitive to thisvirinducer. This supports a model in which transporter activity is crucial in ensuring thatvirgene expression occurs only at sites of high ligand concentration, such as those at a plant wound site.

scholarly journals Reconstitution of Acetosyringone-Mediated Agrobacterium tumefaciens Virulence Gene Expression in the Heterologous Host Escherichia coli

2001 ◽  
Vol 183 (12) ◽  
pp. 3704-3711 ◽  
Author(s):  
Scott M. Lohrke ◽  
Hongjiang Yang ◽  
Shouguang Jin
Keyword(s):  
Gene Expression ◽  
Escherichia Coli ◽  
Agrobacterium Tumefaciens ◽  
Virulence Genes ◽  
Virulence Gene ◽  
Dependent Manner ◽  
Gene Encoding ◽  
Virulence Gene Expression ◽  
E Coli ◽  
Glucose Binding

ABSTRACT The ability to utilize Escherichia coli as a heterologous system in which to study the regulation ofAgrobacterium tumefaciens virulence genes and the mechanism of transfer DNA (T-DNA) transfer would provide an important tool to our understanding and manipulation of these processes. We have previously reported that the rpoA gene encoding the alpha subunit of RNA polymerase is required for the expression of lacZ gene under the control of virB promoter (virBp::lacZ) in E. colicontaining a constitutively active virG gene [virG(Con)]. Here we show that an RpoA hybrid containing the N-terminal 247 residues from E. coli and the C-terminal 89 residues from A. tumefaciens was able to significantly express virBp::lacZ in E. coli in a VirG(Con)-dependent manner. Utilization oflac promoter-driven virA and virGin combination with the A. tumefaciens rpoA construct resulted in significant inducer-mediated expression of thevirBp::lacZ fusion, and the level ofvirBp::lacZ expression was positively correlated to the copy number of the rpoA construct. This expression was dependent on VirA, VirG, temperature, and, to a lesser extent, pH, which is similar to what is observed in A. tumefaciens. Furthermore, the effect of sugars on virgene expression was observed only in the presence of thechvE gene, suggesting that the glucose-binding protein ofE. coli, a homologue of ChvE, does not interact with the VirA molecule. We also evaluated other phenolic compounds in induction assays and observed significant expression with syringealdehyde, a low level of expression with acetovanillone, and no expression with hydroxyacetophenone, similar to what occurs in A. tumefaciens strain A348 from which the virA clone was derived. These data support the notion that VirA directly senses the phenolic inducer. However, the overall level of expression of thevir genes in E. coli is less than what is observed in A. tumefaciens, suggesting that additional gene(s) from A. tumefaciens may be required for the full expression of virulence genes in E. coli.

scholarly journals Identification of Plant Factors Inducing Virulence Gene Expression in Agrobacterium tumefaciens.

1991 ◽  
Vol 39 (9) ◽  
pp. 2347-2350 ◽  
Author(s):  
Yan-Nong SONG ◽  
Masaaki SHIBUYA ◽  
Yutaka EBIZUKA ◽  
Ushio SANKAWA
Keyword(s):  
Gene Expression ◽  
Agrobacterium Tumefaciens ◽  
Virulence Gene ◽  
Virulence Gene Expression

scholarly journals Identification of an Orphan Response Regulator Required for the Virulence of Francisella spp. and Transcription of Pathogenicity Island Genes

2007 ◽  
Vol 75 (7) ◽  
pp. 3305-3314 ◽  
Author(s):  
Nrusingh P. Mohapatra ◽  
Shilpa Soni ◽  
Brian L. Bell ◽  
Richard Warren ◽  
Robert K. Ernst ◽  
...  
Keyword(s):  
Gene Expression ◽  
Mutant Strain ◽  
Lipid A ◽  
Response Regulator ◽  
Model Organism ◽  
Virulence Gene ◽  
Pathogenicity Island ◽  
Effective Vaccine ◽  
Gene Encoding ◽  
Virulence Gene Expression

ABSTRACT Francisella tularensis is a category A agent of biowarfare/biodefense. Little is known about the regulation of virulence gene expression in Francisella spp. Comparatively few regulatory factors exist in Francisella, including those belonging to two-component systems (TCS). However, orphan members of typical TCS can be identified. To determine if orphan TCS members affect Francisella gene expression, a gene encoding a product with high similarity to the Salmonella PmrA response regulator (FTT1557c/FNU0663.2) was deleted in Francisella novicida (a model organism for F. tularensis). The F. novicida pmrA mutant was defective in survival/growth within human and murine macrophage cell lines and was 100% defective in virulence in mice at a dose of up to 108 CFU. In addition, the mutant strain demonstrated increased susceptibility to antimicrobial peptide killing, but no differences were observed between the lipid A of the mutant and the parental strain, as has been observed with pmrA mutants of other microbes. The F. novicida pmrA mutant was 100% protective as a single-dose vaccine when challenge was with 106 CFU of F. novicida but did not protect against type A Schu S4 wild-type challenge. DNA microarray analysis identified 65 genes regulated by PmrA. The majority of these genes were located in the region surrounding pmrA or within the Francisella pathogenicity island (FPI). These FPI genes are also regulated by MglA, but MglA does not regulate pmrA, nor does PmrA regulate MglA. Thus, the orphan response regulator PmrA is an important factor in controlling virulence in F. novicida, and a pmrA mutant strain is an effective vaccine against homologous challenge.

scholarly journals The Global Regulator CodY Regulates Toxin Gene Expression in Bacillus anthracis and Is Required for Full Virulence

2009 ◽  
Vol 77 (10) ◽  
pp. 4437-4445 ◽  
Author(s):  
Willem van Schaik ◽  
Alice Château ◽  
Marie-Agnès Dillies ◽  
Jean-Yves Coppée ◽  
Abraham L. Sonenshein ◽  
...  
Keyword(s):  
Gene Expression ◽  
Bacillus Anthracis ◽  
Mutant Strain ◽  
Virulence Gene ◽  
Etiologic Agent ◽  
Toxin Gene ◽  
Promoter Regions ◽  
Virulence Gene Expression ◽  
Toxin Gene Expression ◽  
Virulence Regulator

ABSTRACT In gram-positive bacteria, CodY is an important regulator of genes whose expression changes upon nutrient limitation and acts as a repressor of virulence gene expression in some pathogenic species. Here, we report the role of CodY in Bacillus anthracis, the etiologic agent of anthrax. Disruption of codY completely abolished virulence in a toxinogenic, noncapsulated strain, indicating that the activity of CodY is required for full virulence of B. anthracis. Global transcriptome analysis of a codY mutant and the parental strain revealed extensive differences. These differences could reflect direct control for some genes, as suggested by the presence of CodY binding sequences in their promoter regions, or indirect effects via the CodY-dependent control of other regulatory proteins or metabolic rearrangements in the codY mutant strain. The differences included reduced expression of the anthrax toxin genes in the mutant strain, which was confirmed by lacZ reporter fusions and immunoblotting. The accumulation of the global virulence regulator AtxA protein was strongly reduced in the mutant strain. However, in agreement with the microarray data, expression of atxA, as measured using an atxA-lacZ transcriptional fusion and by assaying atxA mRNA, was not significantly affected in the codY mutant. An atxA-lacZ translational fusion was also unaffected. Overexpression of atxA restored toxin component synthesis in the codY mutant strain. These results suggest that CodY controls toxin gene expression by regulating AtxA accumulation posttranslationally.

scholarly journals The MexE/MexF/AmeC Efflux Pump of Agrobacterium tumefaciens and Its Role in Ti Plasmid Virulence Gene Expression

2020 ◽  
Vol 202 (8) ◽  
Author(s):  
Andrew N. Binns ◽  
Jinlei Zhao
Keyword(s):  
Gene Expression ◽  
Agrobacterium Tumefaciens ◽  
Efflux Pump ◽  
Bacterial Pathogen ◽  
Virulence Gene ◽  
Ti Plasmid ◽  
Content Type ◽  
Virulence Gene Expression ◽  
Extracellular Milieu ◽  
Phenol Derivatives

ABSTRACT Expression of the tumor-inducing (Ti) plasmid virulence genes of Agrobacterium tumefaciens is required for the transfer of DNA from the bacterium into plant cells, ultimately resulting in the initiation of plant tumors. The vir genes are induced as a result of exposure to certain phenol derivatives, monosaccharides, and low pH in the extracellular milieu. The soil, as well as wound sites on a plant—the usual site of the virulence activity of this bacterium—can contain these signals, but vir gene expression in the soil would be a wasteful utilization of energy. This suggests that mechanisms may exist to ensure that vir gene expression occurs only at the higher concentrations of inducers typically found at a plant wound site. In a search for transposon-mediated mutations that affect sensitivity for the virulence gene-inducing activity of the phenol, 3,5-dimethoxy-4-hydroxyacetophenone (acetosyringone [AS]), an RND-type efflux pump homologous to the MexE/MexF/OprN pump of Pseudomonas aeruginosa was identified. Phenotypes of mutants carrying an insertion or deletion of pump components included hypersensitivity to the vir-inducing effects of AS, hypervirulence in the tobacco leaf explant virulence assay, and hypersensitivity to the toxic effects of chloramphenicol. Furthermore, the methoxy substituents on the phenol ring of AS appear to be critical for recognition as a pump substrate. These results support the hypothesis that the regulation of virulence gene expression is integrated with cellular activities that elevate the level of plant-derived inducers required for induction so that this occurs preferentially, if not exclusively, in a plant environment. IMPORTANCE Expression of genes controlling the virulence activities of a bacterial pathogen is expected to occur preferentially at host sites vulnerable to that pathogen. Host-derived molecules that induce such activities in the plant pathogen Agrobacterium tumefaciens are found in the soil, as well as in the plant. Here, we tested the hypothesis that mechanisms exist to suppress the sensitivity of Agrobacterium species to a virulence gene-inducing molecule by selecting for mutant bacteria that are hypersensitive to its inducing activity. The mutant genes identified encode an efflux pump whose proposed activity increases the concentration of the inducer necessary for vir gene expression; this pump is also involved in antibiotic resistance, demonstrating a relationship between cellular defense activities and the control of virulence in Agrobacterium.

scholarly journals Agrobacteria reprogram virulence gene expression by controlled release of host-conjugated signals

2019 ◽  
Vol 116 (44) ◽  
pp. 22331-22340 ◽  
Author(s):  
Chao Wang ◽  
Fuzhou Ye ◽  
Changqing Chang ◽  
Xiaoling Liu ◽  
Jianhe Wang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Salicylic Acid ◽  
Agrobacterium Tumefaciens ◽  
Virulence Gene ◽  
Tumor Formation ◽  
Bacterial Virulence ◽  
In Planta ◽  
Virulence Gene Expression ◽  
Successful Infection ◽  
Plant Metabolite

It is highly intriguing how bacterial pathogens can quickly shut down energy-costly infection machinery once successful infection is established. This study depicts that mutation of repressor SghR increases the expression of hydrolase SghA in Agrobacterium tumefaciens, which releases plant defense signal salicylic acid (SA) from its storage form SA β-glucoside (SAG). Addition of SA substantially reduces gene expression of bacterial virulence. Bacterial vir genes and sghA are differentially transcribed at early and later infection stages, respectively. Plant metabolite sucrose is a signal ligand that inactivates SghR and consequently induces sghA expression. Disruption of sghA leads to increased vir expression in planta and enhances tumor formation whereas mutation of sghR decreases vir expression and tumor formation. These results depict a remarkable mechanism by which A. tumefaciens taps on the reserved pool of plant signal SA to reprogram its virulence upon establishment of infection.

scholarly journals The Tzs Protein and Exogenous Cytokinin Affect Virulence Gene Expression and Bacterial Growth of Agrobacterium tumefaciens

2013 ◽  
Vol 103 (9) ◽  
pp. 888-899 ◽  
Author(s):  
Hau-Hsuan Hwang ◽  
Fong-Jhih Yang ◽  
Tun-Fang Cheng ◽  
Yi-Chun Chen ◽  
Ying-Ling Lee ◽  
...  
Keyword(s):  
Gene Expression ◽  
Agrobacterium Tumefaciens ◽  
Bacterial Growth ◽  
Deletion Mutant ◽  
Virulence Gene ◽  
Protein Accumulation ◽  
Virulence Gene Expression ◽  
Crown Gall Disease ◽  
Wide Range ◽  
Exogenous Cytokinin

The soil phytopathogen Agrobacterium tumefaciens causes crown gall disease in a wide range of plant species. The neoplastic growth at the infection sites is caused by transferring, integrating, and expressing transfer DNA (T-DNA) from A. tumefaciens into plant cells. A trans-zeatin synthesizing (tzs) gene is located in the nopaline-type tumor-inducing plasmid and causes trans-zeatin production in A. tumefaciens. Similar to known virulence (Vir) proteins that are induced by the vir gene inducer acetosyringone (AS) at acidic pH 5.5, Tzs protein is highly induced by AS under this growth condition but also constitutively expressed and moderately upregulated by AS at neutral pH 7.0. We found that the promoter activities and protein levels of several AS-induced vir genes increased in the tzs deletion mutant, a mutant with decreased tumorigenesis and transient transformation efficiencies, in Arabidopsis roots. During AS induction and infection of Arabidopsis roots, the tzs deletion mutant conferred impaired growth, which could be rescued by genetic complementation and supplementing exogenous cytokinin. Exogenous cytokinin also repressed vir promoter activities and Vir protein accumulation in both the wild-type and tzs mutant bacteria with AS induction. Thus, the tzs gene or its product, cytokinin, may be involved in regulating AS-induced vir gene expression and, therefore, affect bacterial growth and virulence during A. tumefaciens infection.

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