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.

Abstract

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.

Detection of and Response to Signals Involved in Host-Microbe Interactions by Plant-Associated Bacteria

“…These studies allow monitoring of the spatial distribution and fluctuations of physicochemical factors that are relevant for the microbes inhabiting the plant-associated environments. The factors studied so far have included UV irradiation, temperature, water potential, and iron availability on surfaces of leaves (17,251,268,499) and carbon, phosphate, nitrogen, iron, and oxygen availability in the soil (217,241,271,275). Bioreporter strains have also been used to detect products of plant metabolism that are released into the surrounding environment and are used by the associated microbes as nutrients (53, 239,285,329).…”

The alternative sigma factor AlgT, but not alginate synthesis, promotes in planta multiplication of Pseudomonas syringae pv. glycinea

“…glycinea is most virulent on its host plant (Ullrich et al, 2000). Optical density of cultures was monitored continuously until the cultures entered the late stationary phase (Fig.…”

“…The infection process involves epiphytic colonization of leaf surfaces, establishment of infection sites via entry into the plant apoplast, multiplication within host tissue and development of disease symptoms (Alfano & Collmer, 1996;Hirano & Upper, 2000). P. syringae and other plant-pathogenic bacteria may cause 'cold-weather' diseases and many genes studied with respect to temperature exhibit increased transcription at temperatures well below the respective growth optima (Ullrich et al, 2000). This includes virulence determinants such as those directing bacteriato-plant gene transfer, plant cell-wall-degrading enzymes, phytotoxins, ice nucleation activity, the type III protein secretion machinery and EPS production (Smirnova et al, 2001).…”

The Pseudomonas syringae Genome Encodes a Combined Mannuronan C-5-epimerase and O-Acetylhydrolase, Which Strongly Enhances the Predicted Gel-forming Properties of Alginates

“…Ullrich et al (32) used a promoter-trapping strategy to identify P. syringae PG4180 promoters with induced expression at 18°C when compared with 28°C. Sequencing of several hundred nucleotides of the transcriptional fusion contained in plasmid p561 revealed the presence of an open reading frame with 53% amino acid identity to the extracellular epimerase AlgE2 from A. vinelandii (32). In this report, we describe the molecular cloning and characterization of this gene (designated psmE) and show that it encodes a bifunctional enzyme possessing both G block-forming ME activity and mannuronan-O-acetylhydrolase activity.…”