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Thermo-regulation in Pseudomonas syringae: Phenotypes and Genetics

  • Author(s): Hockett, Kevin Loren
  • Advisor(s): Lindow, Steven E
  • et al.
Abstract

Pseudomonas syringae is an important member of the phyllosphere microbial community and has been studied for many decades both as a model saprophyte, residing epiphytically, and a pathogen, living in the plant apoplast. While much is known about the traits that contribute to P. syringae's success as a phyllosphere microbe, one area that is not well understood, is the contribution of temperature dependent gene regulation, or thermo-regulation, to P. syringae's epiphytic colonization and survival strategies.

Flagellar-mediated motility is a trait conserved among plant-associated Pseudomonads. In P. syringae, motility contributes both to epiphytic colonization and survival, as well as pathogenicity by allowing cells to invade into the leaf interior. We have found that multiple forms of flagellar-mediated motility, including swarming and swimming motility, are thermo-repressed at around 30 °C. Repression of swarming results from reduced expression of both flagellar genes, including flagellin, encoded by fliC, as well genes involved in regulation and biosynthesis of syringafactin, the major surfactant produced by P. syringae B728a. Thermo-regulation of the flagellum is context dependent, being influenced by the nutrient status of the agar, which together with temperature contribute to a heterogeneous swimming phenotype at elevated temperatures. The heterogeneous swimming phenotype may represent a so-called "bet-hedging" strategy, which may be an important strategy for colonization of the leaf surface under varying and unpredictable weather conditions.

Investigation into the molecular determinants of thermo-regulation revealed several regulators, which are all conserved across the Pseudomonas genus, potentially indicating these are conserved thermo-regulators with in this important group of organisms. flgM as well as fleN were both involved in thermo-regulation of the flagellum. Additionally, fleN appeared to be important for the heterogeneous swimming phenotype, as mutations in Spontaneous Hot Swimming (SHS) mutants, which swim at elevated temperatures with a cool, diffuse swimming phenotype, commonly mapped to this gene, as determined by genome resequencing. Two separate loci, an acyl-CoA dehydrogenase and a nudix hydrolase, together contributed to thermo-regulation of genes involved in the regulation and synthesis of syringafactin. Interestingly, mutations that contributed to thermo-regulation of the flagellum did not affect thermo-regulation of syringafactin, while mutations affecting thermo-regulation of syringafactin did not affect thermo-regulation of the flagellum, indicating that thermo-regulation of these two traits are independently controlled.

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