Intersubunit Complementation of Sugar Signal Transduction in VirA Heterodimers and Posttranslational Regulation of VirA Activity inAgrobacterium tumefaciens

Wise, Arlene A.; Voinov, Luba; Binns, Andrew N.

doi:10.1128/jb.187.1.213-223.2005

Cited by 22 publications

(19 citation statements)

References 49 publications

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“…In contrast, complete removal of medium components from the inoculation medium, i.e., only D. W., gave significantly lower transformation efficiency (6.7%), suggesting the importance of maltose during infection period for the successful transformation with Agrobacterium. Previous studies demonstrated that sugars greatly increased the expression of vir genes in combination with AS (Shimoda et al 1990;Wise et al 2005). In the present study, difference in the presence of maltose in medium was made only at short period of inoculation but not at subsequent cocultivation period.…”

Section: Effect Of Different Stage Of Plbs After Subculture and Inocusupporting

confidence: 48%

Agrobacterium-mediated transformation of protocorm-like bodies in Dendrobium Formidible ‘Ugusu’

Phlaetita

Chin

Tokuhara

et al. 2015

Plant Biotechnology

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Agrobacterium-mediated genetic transformation system was established in Dendrobium Formidible 'Ugusu' by inoculating PLBs with A. tumefaciens strain EHA101 (pIG121Hm) harboring hygromycin phosphotranferase (hpt) and neomycin phosphotranferase II (nptII) genes as selectable marker gene and β-glucuronidase (gus) gene as a reporter gene. For obtaining the optimum conditions for the transformation, several factors such as the stage of PLBs after subculture, bacterial concentrations, kind of inoculation medium, inoculation time and inoculation condition (with or without rotary shaking), were examined. After inoculation, PLBs were cocultivated for 3 days and then transferred for selection onto 2.5 g l −1 gellan gum-solidified ND medium containing 10 g l −1 maltose, 20 mg l −1 hygromycin and 20 mg l −1 meropenem. Hygromycin-resistant plantlets were regenerated from secondary PLBs after 4 months of selection. Transformation of these plants was confirmed by GUS histochemical assay, PCR and Southern blot analyses. The highest transformation efficiency of 18.5% was obtained when PLBs 3 weeks after subculture were inoculated with 1 : 10 diluted bacteria (OD 600 ≈0.1) with liquid medium containing only 10 g l −1 maltose and 100 µM acetosyringone with shaking for 30 min.

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Section: Effect Of Different Stage Of Plbs After Subculture and Inocusupporting

confidence: 48%

Agrobacterium-mediated transformation of protocorm-like bodies in Dendrobium Formidible ‘Ugusu’

Phlaetita

Chin

Tokuhara

et al. 2015

Plant Biotechnology

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“…The presence of biologically relevant dimers has been demonstrated both biophysically and genetically for the Tar chemoreceptor and histidine kinases such as PhoQ, DcuS, NarX, and TorS (20,32,77,79). The evidence for VirA dimers comes by way of genetic analysis using intersubunit complementation (9,66,73) and by chemical cross-linking analysis indicating that VirA exists as a dimer even in the absence of the inducing ligands (57). While these earlier studies affirm that VirA exists as a dimer and that dimerization is critical to VirA function, our investigations demonstrate periplasmic domain dimerization in the absence of tethering via transmembrane and cytoplasmic domains.…”

Section: Discussionmentioning

confidence: 99%

The Integrity of the Periplasmic Domain of the VirA Sensor Kinase Is Critical for Optimal Coordination of the Virulence Signal Response inAgrobacterium tumefaciens

et al. 2011

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The plant pathogen Agrobacterium tumefaciens responds to three main signals at the plant-bacterium interface: phenolics, such as acetosyringone (AS), monosaccharides, and acidic pH (ϳ5.5). These signals are transduced via the chromosomally encoded sugar binding protein ChvE and the Ti plasmid-encoded VirA/VirG two-component regulatory system, resulting in the transcriptional activation of the Ti plasmid virulence genes. Here, we present genetic and physical evidence that the periplasmic domain of VirA dimerizes independently of other parts of the protein, and we examine the effects of several engineered mutations in the periplasmic and transmembrane regions of VirA on vir-inducing capacity as indicated by AS sensitivity and maximal level of vir-inducing activity at saturating AS levels. The data indicate that helix-breaking mutations throughout the periplasmic domain of VirA or mutations that reposition the second transmembrane domain (TM2) of VirA relieve the periplasmic domain's repressive effects on the maximal activity of this kinase in response to phenolics, effects normally relieved only when ChvE, sugars, and low pH are also present. Such relief, however, does not sensitize VirA to low concentrations of phenolics, the other major effect of the ChvE-sugar and low pH signals. We further demonstrate that amino acid residues in a small Trg-like motif in the periplasmic domain of VirA are crucial for transmission of the ChvE-sugar signal to the cytoplasmic domain. These experiments provide evidence that small perturbations in the periplasmic domain of VirA can uncouple sugar-mediated changes in AS sensitivity from the sugar-mediated effects on maximal activity.

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“…pAW10 and pYW47 are similar in that they are both derivatives of pYW10 (31,37). Derivatives of these plasmids are expected to be present in Agrobacterium at levels between 15 and 20 copies per pTi (37). Plasmid pAW168 was constructed by first moving the 7-kb HpaI insert from pVRA6 to pAW24.…”

Section: Methodsmentioning

confidence: 99%

The Receiver Domain of Hybrid Histidine Kinase VirA: an Enhancing Factor for vir Gene Expression in Agrobacterium tumefaciens

et al. 2010

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The plant pathogen Agrobacterium tumefaciens expresses virulence (vir) genes in response to chemical signals found at the site of a plant wound. VirA, a hybrid histidine kinase, and its cognate response regulator, VirG, regulate vir gene expression. The receiver domain at the carboxyl end of VirA has been described as an inhibitory element because its removal increased vir gene expression relative to that of full-length VirA. However, experiments that characterized the receiver region as an inhibitory element were performed in the presence of constitutively expressed virG. We show here that VirA's receiver domain is an activating factor if virG is expressed from its native promoter on the Ti plasmid. When virA⌬R was expressed from a multicopy plasmid, both sugar and the phenolic inducer were essential for vir gene expression. Replacement of wild-type virA on pTi with virA⌬R precluded vir gene induction, and the cells did not accumulate VirG or induce transcription of a virG-lacZ fusion in response to acetosyringone. These phenotypes were corrected if the virG copy number was increased. In addition, we show that the VirA receiver domain can interact with the VirG DNA-binding domain.

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Intersubunit Complementation of Sugar Signal Transduction in VirA Heterodimers and Posttranslational Regulation of VirA Activity inAgrobacterium tumefaciens

Cited by 22 publications

References 49 publications

<i>Agrobacterium</i>-mediated transformation of protocorm-like bodies in <i>Dendrobium</i> Formidible ‘Ugusu’

<i>Agrobacterium</i>-mediated transformation of protocorm-like bodies in <i>Dendrobium</i> Formidible ‘Ugusu’

The Integrity of the Periplasmic Domain of the VirA Sensor Kinase Is Critical for Optimal Coordination of the Virulence Signal Response inAgrobacterium tumefaciens

The Receiver Domain of Hybrid Histidine Kinase VirA: an Enhancing Factor for vir Gene Expression in Agrobacterium tumefaciens

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