|Corbell, N. - OREGON STATE UNIVERSITY|
|Reifenrath, C. - OREGON STATE UNIVERSITY|
Submitted to: International Congress on Pseudomonas Molecular Biology and Biotechnology
Publication Type: Proceedings
Publication Acceptance Date: June 1, 1997
Publication Date: N/A
Technical Abstract: Production of the extracellular metabolites pyoluteorin, 2,4- diacetylphloroglucinol, and hydrogen cyanide (HCN); an extracellular protease; and tryptophan-side-chain oxidase (TSO) are controlled by the apdA (=lemA) and gacA genes, which encode a sensor kinase and response regulator, respectively, in Pseudomonas fluorescens Pf-5. Colonies with a ApdA-/GacA- morphology (large, orange, and highly fluorescent) arise with remarkable frequency in cultures of P. fluorescens: they comprised up to 40% of the total colonies harvested from six-day cultures of Pf-5 grown in nutrient yeast broth. Certain colonies with the altered morphology exhibited only a subset of the phenotypes controlled by these global regulators. For example, a Pf- 5 derivative with a single point mutation in the carboxy-terminal region of GacA did not produce pyoluteorin, pyrrolnitrin, 2,4- diacetylphloroglucinol or protease, but did produce HCN and TSO at less than wildtype levels. The point mutation was located thus, the carboxy-terminal region GacA may confer specificity for certain target genes. In Pf-5, wildtype apdA and gacA genes cloned in low copy number plasmids such as pRK415 (1-4 copies/cell) restore the wildtype phenotype to GacA- and ApdA- mutants, respectively. When present on a high copy plasmid (i.e. pME6000, 16-20 copies/cell), however, apdA does not complement either ApdA- or GacA- mutants and gacA complements both ApdA- and GacA- mutants. We speculate that proper function of the ApdA/GacA system requires a stoichiometric balance between the sensor kinase and the response regulator that is disrupted by high copy numbers of either apdA or gacA.