Organization and environmental regulation of the Pseudomonas syringae pv. syringae 61 hrp cluster

Xiao, Yingxian; Lu, Yingjian; Heu, Sunggi; Hutcheson, Steven W.

doi:10.1128/jb.174.6.1734-1741.1992

Cited by 152 publications

(128 citation statements)

References 44 publications

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“…b-Glucuronidase activity (GUS) was quantified by a fluorescence assay, as described previously (Xiao et al, 1992). The pellet was lysed in 500 ml GUS extraction buffer (50 mM Na 2 HPO 4 , adjusted to pH 7?0, 10 mM EDTA disodium salt, 0?1 % N-laurolylsarcosyl sodium salt, 0?1 % Triton X-100, 0?07 % b-mercaptoethanol), and incubated on ice for 30 min.…”

Section: Methodsmentioning

confidence: 99%

Topological and deletion analysis of CorS, a Pseudomonas syringae sensor kinase

Smirnova

Ullrich

2004

Microbiology

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A modified two-component regulatory system consisting of two response regulators, CorR and CorP, and the histidine protein kinase CorS, regulates the thermoresponsive production of the phytotoxin coronatine (COR) in Pseudomonas syringae PG4180. COR is produced at the virulence-promoting temperature of 18 6C, but not at 28 6C, the optimal growth temperature of PG4180. Assuming that the highly hydrophobic N-terminus of CorS might be involved in temperature-signal perception, the membrane topology of CorS was determined using translational phoA and lacZ fusions, leading to a topological model for CorS with six transmembrane domains (TMDs). Interestingly, three PhoA fusions located downstream of the sixth TMD showed a thermoresponsive phenotype. Enzymic activity, immunoblot, and protease-sensitivity assays were performed to localize the CorS derivatives, to analyse the expression level of hybrid proteins and to examine the model. In-frame deletions of the last four, or all six TMDs gave rise to non-functional CorS. The results indicated that the transmembrane region is important for CorS to function as a temperature sensor, and that the membrane topology of CorS might be involved in signal perception.

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Section: Methodsmentioning

confidence: 99%

Topological and deletion analysis of CorS, a Pseudomonas syringae sensor kinase

Smirnova

Ullrich

2004

Microbiology

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“…(Sambrook et al, 1989) Wilson et al (1992). Fluorometric analysis of GUS activity (Xiao et al, 1992) was carried out using a Fluorolite-1000 micro-plate reader (Dynatech) and 96-well microtitre plates.…”

Section: Methodsmentioning

confidence: 99%

Temperature-responsive genetic loci in the plant pathogen Pseudomonas syringae pv. glycinea The GenBank accession numbers for the nucleotide sequences of mutants 560, 561, 562, 563, 564, 568, 570, 574, 590, 591, 593, 596, 599, 601, 605, 608, 613, 617, 618, 626 and 632 determined in this work are AF274322–AF274342, respectively.

et al. 2000

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Plant-pathogenic bacteria may sense variations in environmental factors, such as temperature, to adapt to plant-associated habitats during pathogenesis or epiphytic growth. The bacterial blight pathogen of soybean, Pseudomonas syringae pv. glycinea PG4180, preferentially produces the phytotoxin coronatine at 18 SC and infects the host plant under conditions of low temperature and high humidity. A miniTn5-based promoterless glucuronidase (uidA) reporter gene was used to identify genetic loci of PG4180 preferentially expressed at 18 or 28 SC. Out of 7500 transposon mutants, 61 showed thermoregulated uidA expression as determined by a three-step screening procedure. Two-thirds of these mutants showed an increased reporter gene expression at 18 SC whilst the remainder exhibited higher uidA expression at 28 SC. MiniTn5-uidA insertion loci from these mutants were subcloned and their nucleotide sequences were determined. Several of the mutants induced at 18 SC contained the miniTn5-uidA insertion within the 328 kb coronatine biosynthetic gene cluster. Among the other mutants with increased uidA expression at 18 SC, insertions were found in genes encoding formaldehyde dehydrogenase, short-chain dehydrogenase and mannuronan C-5-epimerase, in a plasmid-borne replication protein, and in the hrpT locus, involved in pathogenicity of P. syringae. Among the mutants induced at 28 SC, insertions disrupted loci with similarities to a repressor of conjugal plasmid transfer, UV resistance determinants, an isoflavanoid-degrading enzyme, a HU-like DNAbinding protein, two additional regulatory proteins, a homologue of bacterial adhesins, transport proteins, LPS synthesis enzymes and two proteases. Genetic loci from 13 mutants did not show significant similarities to any database entries. Results of plant inoculations showed that three of the mutants tested were inhibited in symptom development and in planta multiplication rates. Temperature-shift experiments suggested that all of the identified loci showed a rather slow induction of expression upon change of temperature.

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“…HrpL recognizes a consensus sequence motif (''harp box'') that has been identified in the upstream regions of many hrp and avr genes (4). The expression of hrp genes of many P. syringae pathovars can also be induced in vitro when bacteria are grown in defined minimal medium with low pH and containing certain sugars or sugar alcohols as carbon sources (5)(6)(7).…”

mentioning

confidence: 99%

Hrp pilus: An hrp -dependent bacterial surface appendage produced by Pseudomonas syringae pv. tomato DC3000

Roine

Wei

Yuan

et al. 1997

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

323

279

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Hypersensitive response and pathogenicity (hrp) genes control the ability of major groups of plant pathogenic bacteria to elicit the hypersensitive response (HR) in resistant plants and to cause disease in susceptible plants. A number of Hrp proteins share significant similarities with components of the type III secretion apparatus and f lagellar assembly apparatus in animal pathogenic bacteria. Here we report that Pseudomonas syringae pv. tomato strain DC3000 (race 0) produces a filamentous surface appendage (Hrp pilus) of 6-8 nm in diameter in a solid minimal medium that induces hrp genes. Formation of the Hrp pilus is dependent on at least two hrp genes, hrpS and hrpH (recently renamed hrcC), which are involved in gene regulation and protein secretion, respectively. Our finding of the Hrp pilus, together with recent reports of Salmonella typhimurium surface appendages that are involved in bacterial invasion into the animal cell and of the Agrobacterium tumefaciens virB-dependent pilus that is involved in the transfer of T-DNA into plant cells, suggests that surface appendage formation is a common feature of animal and plant pathogenic bacteria in the infection of eukaryotic cells. Furthermore, we have identified HrpA as a major structural protein of the Hrp pilus. Finally, we show that a nonpolar hrpA mutant of P. syringae pv. tomato DC3000 is unable to form the Hrp pilus or to cause either an HR or disease in plants.Major groups of Gram-negative plant pathogenic bacteria belonging to genera Erwinia, Pseudomonas, Ralstonia, and Xanthomonas contain hypersensitive reaction and pathogenicity (hrp) genes. These genes control the ability of these bacteria to initiate interactions with plants, including elicitation of the hypersensitive reaction (HR), characterized by rapid localized death of plant cells at the pathogen infection site in resistant plants and causation of disease in susceptible plants (1, 2).hrp genes of Pseudomonas syringae are expressed in planta as a result of a regulatory cascade involving the gene products of hrpS and hrpR, positive transcriptional regulators, and of hrpL, an alternative sigma factor (3, 4). HrpL recognizes a consensus sequence motif (''harp box'') that has been identified in the upstream regions of many hrp and avr genes (4). The expression of hrp genes of many P. syringae pathovars can also be induced in vitro when bacteria are grown in defined minimal medium with low pH and containing certain sugars or sugar alcohols as carbon sources (5-7).The 25-kb hrp͞hrmA gene cluster of Pseudomonas syringae pv. syringae strain 61 is sufficient to enable nonpathogenic strains of Pseudomonas fluorescens and Escherichia coli to elicit the HR in nonhost plants (8). Sixteen of the 25 genes in this completely sequenced hrp͞hrmA gene cluster are either predicted or shown to be required for secretion of harpin Pss , a proteinaceous elicitor of the HR encoded by hrpZ (9, 10). Nine of these hrp genes, recently renamed hrc genes (11), are broadly conserved among P. syringae pathovars, Erwi...

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Organization and environmental regulation of the Pseudomonas syringae pv. syringae 61 hrp cluster

Cited by 152 publications

References 44 publications

Topological and deletion analysis of CorS, a Pseudomonas syringae sensor kinase

Topological and deletion analysis of CorS, a Pseudomonas syringae sensor kinase

Hrp pilus: An hrp -dependent bacterial surface appendage produced by Pseudomonas syringae pv. tomato DC3000

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