The <i>att</i> locus of <i>Rhodococcus fascians</i> strain D188 is essential for full virulence on tobacco through the production of an autoregulatory compound

Maes, Tania; Vereecke, Danny; Ritsema, Tita; Cornelis, Karen; Thu, Hang Ngo Thi; Montagu, Marc Van; Holsters, Marcelle; Goethals, Koen

doi:10.1046/j.1365-2958.2001.02615.x

Cited by 54 publications

(62 citation statements)

References 52 publications

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“…In this context, it is interesting that the augmentation of Trp was one of the highest measured. Production of the autoregulatory compound by the att locus involves Arg biosynthesis, and expression of the att genes is inhibited by Orn and Arg (Maes et al, 2001). Again, it is remarkable that the concentrations of these amino acids were lower than those of the controls at the onset of the interaction.…”

Section: Discussionmentioning

confidence: 99%

“…Similarly, transcript and metabolite data indicated that the flux through the TCA cycle was also reduced when the disease became fully established, although the concentration of succinate remained high throughout the interaction and pyruvate levels were elevated at the early stages of infection. Interestingly, virulence gene expression in R. fascians is induced by an autoregulatory compound and the induction levels are significantly increased in the presence of succinate and pyruvate (Temmerman et al, 2000;Maes et al, 2001). The decreasing amounts of Ser and Gly, together with the increase in glycerate levels, may point to photorespiratory Gly decarboxylation that would add to the reduced photosynthetic and TCA cycle activities (Atkin et al, 2000;Raghavendra and Padmasree, 2003;NunesNesi et al, 2007).…”

Section: Discussionmentioning

confidence: 99%

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An Integrated Genomics Approach to Define Niche Establishment byRhodococcus fascians

et al. 2008

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Rhodococcus fascians is a Gram-positive phytopathogen that induces shooty hyperplasia on its hosts through the secretion of cytokinins. Global transcriptomics using microarrays combined with profiling of primary metabolites on infected Arabidopsis (Arabidopsis thaliana) plants revealed that this actinomycete modulated pathways to convert its host into a niche. The transcript data demonstrated that R. fascians leaves a very characteristic mark on Arabidopsis with a pronounced cytokinin response illustrated by the activation of cytokinin perception, signal transduction, and homeostasis. The microarray data further suggested active suppression of an oxidative burst during the R. fascians pathology, and comparison with publicly available transcript data sets implied a central role for auxin in the prevention of plant defense activation. Gene Ontology categorization of the differentially expressed genes hinted at a significant impact of infection on the primary metabolism of the host, which was confirmed by subsequent metabolite profiling. The much higher levels of sugars and amino acids in infected plants are presumably accessed by the bacteria as carbon and nitrogen sources to support epiphytic and endophytic colonization. Hexoses, accumulating from a significantly increased invertase activity, possibly inhibited the expression of photosynthesis genes and photosynthetic activity in infected leaves. Altogether, these changes are indicative of sink development in symptomatic tissues. The metabolomics data furthermore point to the possible occurrence of secondary signaling during the interaction, which might contribute to symptom development. These data are placed in the context of regulation of bacterial virulence gene expression, suppression of defense, infection phenotype, and niche establishment.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

An Integrated Genomics Approach to Define Niche Establishment byRhodococcus fascians

et al. 2008

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“…Bacterial isolates had always been grown in relatively rich media, but from recent insights, we know that essential virulence genes are not expressed under these conditions (22,27,32). Therefore, we determined the CK profiles of strain D188 and its plasmid-free derivative D188-5 as reference, grown in defined minimal medium optimized for virulence gene expression (22).…”

Section: Discussionmentioning

confidence: 99%

Identification ofRhodococcus fascianscytokinins and their modus operandi to reshape the plant

Pertry

Václavíková

Depuydt

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

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Decades ago, the importance of cytokinins (CKs) during Rhodococcus fascians pathology had been acknowledged, and an isopentenyltransferase gene had been characterized in the fas operon of the linear virulence plasmid, but hitherto, no specific CK(s) could be associated with virulence. We show that the CK receptors AHK3 and AHK4 of Arabidopsis thaliana are essential for symptom development, and that the CK perception machinery is induced upon infection, underlining its central role in the symptomatology. Three classical CKs [isopentenyladenine, trans-zeatin, and cis-zeatin (cZ)] and their 2-methylthio (2MeS)-derivatives were identified by CK profiling of both the pathogenic R. fascians strain D188 and its nonpathogenic derivative D188 -5. However, the much higher CK levels in strain D188 suggest that the linear plasmid is responsible for the virulenceassociated production. All R. fascians CKs were recognized by AHK3 and AHK4, and, although they individually provoked typical CK responses in several bioassays, the mixture of bacterial CKs exhibited clear synergistic effects. The cis-and 2MeS-derivatives were poor substrates of the apoplastic CK oxidase/dehydrogenase enzymes and the latter were not cytotoxic at high concentrations. Consequently, the accumulating 2MeScZ (and cZ) in infected Arabidopsis tissue contribute to the continuous stimulation of tissue proliferation. Based on these results, we postulate that the R. fascians pathology is based on the local and persistent secretion of an array of CKs.phytopathogen ͉ actinomycete ͉ phytohormone T he fine-tuned balance of plant regulators has a key role in the growth and development of plants. Many plant-associated bacteria can influence their hosts by either modulating the phytohormone production or producing the phytohormones themselves. The main advantages for the bacteria are increased nutrient release, suppression of defense, and/or disease establishment (1, 2). Hyperplasia-inducing bacteria, such as Pantoea agglomerans and Pseudomonas savastanoi, secrete high amounts of cytokinins (CKs) and auxins to facilitate or initiate gall development (3, 4), and Agrobacterium tumefaciens genetically transforms plant cells to convert them into CK and auxin (and opine) factories (5).In contrast to the undifferentiated galls induced by the bacteria mentioned above, the Actinomycete Rhodococcus fascians that shares persistence strategies with the closely related human pathogen Mycobacterium tuberculosis (6) provokes the formation of differentiated leafy galls, consisting of numerous shoot primordia whose further outgrowth is inhibited (7). The shooty symptoms can be partially mimicked by exogenous addition of CKs (8, 9), and analyses of culture supernatants of different nonisogenic virulent and avirulent R. fascians strains grown under rich culture conditions identified 11 different CKs: methylaminopurine, 2-methylthioisopentenyladenine (2MeSiP), iP, cis-zeatin (cZ), trans-zeatin (tZ), dihydrozeatin (DZ), 2MeScZ, and their respective ribosides (10-14). Except for iP, the p...

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“…The fasA-F and the fasA-D regions were amplified via PCR from genomic DNA of strain D188 with the primer pairs fasAstart-fasFend and fasAstart-fasDend (Supplementary Table 1), respectively, and cloned as XbaI/HindIII fragments into pRF37 (a bifunctional cloning vector for R. fascians) (Desomer et al 1990), yielding the cloning intermediates pRF37fasA/F and pRF37fasA/D. The promoter of the att operon of R. fascians (Maes et al 2001) was amplified via PCR from genomic DNA of strain D188 with the primers attpromstart and attpromend and cloned as an XbaI fragment into pRF37fasA/F and pRF37fasA/D, yielding the final constructs pRF37fasOE-A/F and pRF37fasOE-A/D. These plasmids were introduced into D188-fas5 and D188-fasD ) via electroporation and selection on phleomycin (Desomer et al 1990).…”

Section: Construction Of Overexpression Plasmids and Generation Of Famentioning

confidence: 99%

Rhodococcus fascians Impacts Plant Development Through the Dynamic Fas-Mediated Production of a Cytokinin Mix

Pertry¹,

Václavíková²,

Gemrotová³

et al. 2010

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The phytopathogenic actinomycete Rhodococcus fascians D188 relies mainly on the linear plasmid-encoded fas operon for its virulence. The bacteria secrete six cytokinin bases that synergistically redirect the developmental program of the plant to stimulate proliferation of young shoot tissue, thus establishing a leafy gall as a niche. A yeast-based cytokinin bioassay combined with cytokinin profiling of bacterial mutants revealed that the fas operon is essential for the enhanced production of isopentenyladenine, trans-zeatin, cis-zeatin, and the 2-methylthio derivatives of the zeatins. Cytokinin metabolite data and the demonstration of the enzymatic activities of FasD (isopentenyltransferase), FasE (cytokinin oxidase/dehydrogenase), and FasF (phosphoribohydrolase) led us to propose a pathway for the production of the cytokinin spectrum. Further evaluation of the pathogenicity of different fas mutants and of fas gene expression and cytokinin signal transduction upon infection implied that the secretion of the cytokinin mix is a highly dynamic process, with the consecutive production of a tom initiation wave followed by a maintenance flow.Rhodococcus fascians is a phytopathogenic actinomycete with a very broad host range that causes important commercial losses in the ornamentals industry because it triggers severe malformations of shoots, referred to as leafy galls (Depuydt et al. 2008b). In strain D188, the virulence determinants are encoded by a large conjugative linear plasmid, pFiD188 , and the pathology is induced by the secretion of a mix of six synergistically acting cytokinins: isopentenyladenine (iP), trans-zeatin (tZ), cis-zeatin (cZ), and their 2-methylthio (2MeS) derivatives (Pertry et al. 2009). In Arabidopsis thaliana, these cytokinins are perceived by the receptors AHK3 and CRE1/AHK4 (Pertry et al. 2009), activating a signaling cascade that stimulates cell proliferation and meristematic gene expression and, ultimately, results in the establishment of a specific niche (Depuydt et al. 2008a.Comparison of the cytokinin profiles of two near-isogenic strains, D188 and its plasmid-free derivative, D188-5, has shown that a basal level of the six cytokinins is produced by a chromosomally encoded pathway. However, the much higher levels of iP, cZ, tZ, 2MeScZ, and 2MeStZ secreted by strain D188 strongly suggest an additional linear plasmid-encoded de novo biosynthetic pathway (Pertry et al. 2009). By sequence analysis of pFiD188, the fas operon was identified, consisting of six genes putatively involved in cytokinin biosynthesis and essential for virulence (Fig. 1). FasA is similar to P450-type cytochrome monooxygenases. The N-terminal region of FasB corresponds to 4Fe-3S-type ferredoxins of Actinomycetes, whereas its carboxy-terminus is homologous to the α subunit of pyruvate dehydrogenase. FasC is similar to the β subunit of the latter enzyme. Both FasB and FasC have a binding site for the cofactor thiamine pyrophosphate (Crespi et al. 1994). FasD is an isopentenyltransferase (Ipt) protein that mediates th...

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The att locus of Rhodococcus fascians strain D188 is essential for full virulence on tobacco through the production of an autoregulatory compound

Cited by 54 publications

References 52 publications

An Integrated Genomics Approach to Define Niche Establishment byRhodococcus fascians

An Integrated Genomics Approach to Define Niche Establishment byRhodococcus fascians

Identification ofRhodococcus fascianscytokinins and their modus operandi to reshape the plant

Rhodococcus fascians Impacts Plant Development Through the Dynamic Fas-Mediated Production of a Cytokinin Mix

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