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Title: A functional gene cluster for toxoflavin biosynthesis in the genome of the soil bacterium Pseudomonas protegens Pf-5

item PHILMUS, BENJAMIN - Oregon State University
item Shaffer, Brenda
item YAN, QING - Oregon State University
item Loper, Joyce

Submitted to: Phytopathology
Publication Type: Abstract Only
Publication Acceptance Date: 6/3/2015
Publication Date: 11/1/2015
Citation: Philmus, B., Shaffer, B.T., Yan, Q., Loper, J.E. 2015. A functional gene cluster for toxoflavin biosynthesis in the genome of the soil bacterium Pseudomonas protegens Pf-5 [abstract]. Phytopathology. 105:54.

Interpretive Summary: This was a collaborative project in which Brenda Shaffer served as an integral member of the scientific team from the beginning to the end of the project. Specifically, Brenda generated all of the mutants included in the abstract, from designing primers to sequence analysis of the insertions and she did the experiments evaluating the mutants for antibiosis. She participated in planning the experimental design, and was a full participant in the discussions on how to trouble shoot the system, so that we were finally able to identify conditions (i.e., culture conditions as well as genetic backgrounds) conducive to toxoflavin production. She wrote up the methods and results for these experiments and approved the final version of this abstract.

Technical Abstract: Toxoflavin is a broad-spectrum toxin best known for its role in virulence of Burkholderia glumae, which causes panicle blight of rice. A gene cluster containing homologs of toxoflavin biosynthesis genes (toxA-E) of B. glumae is present in the genome of Pseudomonas protegens Pf-5, a biological control strain that produces many antimicrobial compounds. Repeated efforts to detect toxoflavin production by Pf-5 were unsuccessful so we attempted to associate antimicrobial activity with the gene cluster. Recognizing that this antimicrobial activity may be difficult to discern from the activities of other metabolites, we derived a mutant of Pf-5 (LK185) that lacked production of all of Pf-5’s known antibiotics. LK185 suppressed P. syringae DC3000 on an iron-amended medium (KMB-Fe), whereas toxC or toxD derivatives of LK185 did not suppress DC3000. We then observed that LK185 grown in KMB-Fe broth without shaking produced toxoflavin whereas toxC and toxD mutants did not produce toxoflavin. We propose that the biosynthetic pathways for toxoflavin and riboflavin share the first two steps because deletion of toxE diminished but did not eliminate toxoflavin production, suggesting that the paralog RibD compensates for the loss of ToxE. We also report that toxM, which is present in the toxoflavin cluster of Pf-5 but not B. glumae, encodes a monooxygenase that detoxifies toxoflavin; introduction of toxM into P. syringae DC3000 confered toxoflavin resistance.