Graduate Thesis Or Dissertation
 

Pyoluteorin as a signaling molecule regulating secondary metabolite production and transport genes in Pseudomonas fluorescens Pf-5

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  • A major factor in the ability of Pseudomonas fluorescens Pf-5 to act as a biological control agent is its production of antibiotics, including pyoluteorin (PLT), 2,4-diacetylphloroglucinol (2,4-DAPG) and pyrrolnitrin (PRN). The data provided in this thesis demonstrate that the presence of any of these antibiotics in the extracellular milieu affects production of that same antibiotic, as well as others, by Pf-5. Amending the growth medium with antibiotics had multiple effects on secondary metabolism in Pf-5. i) PLT positively regulated its own production, ii) 2,4-DAPG positively regulated its own production. iii) PLT suppressed 2,4-DAPG production. iv) 2,4- DAPG inhibited PLT production. v) PLT suppressed transcription of a heterologous ferric-pyoverdine uptake gene. vi) PRN exerted a slight inhibitory effect on PLT gene transcription and production. PLT autoinduction by Pf-5 was extensively characterized, and was shown to require concentrations of exogenous PLT in the nanomolar range. These low concentrations are comparable to those of many molecules proposed to function in signaling roles. PLT served as a signal between distinct populations of cells within the rhizosphere, where it prompted autoinduction by those cells. Aside from effects of Pf- 5 antibiotics on one another, I also described the positive effect of exogenous PLT on expression of a set of transport genes flanking the PLT biosynthetic gene cluster. Sequence data and experimental evidence suggests that these genes encode a transport apparatus for PLT. The deduced amino acid sequences for four adjacent open reading frames together resemble Type I secretion apparatuses, which typically function in transport of proteins rather than secondary metabolites. The intact transporter genes are necessary for optimal PLT production. Taken together, the data from the studies described herein demonstrate that i) the production of PLT by Pf-5 can affect the production of PLT by neighboring cells, and ii) PLT and other exogenous secondary metabolites have both autoregulatory and cross-regulatory effects in culture. Because Pf-5 derivatives engaged in PLT crossfeeding in the rhizosphere, it is likely that cross-feeding occurs for other secondary metabolites as well. Thus, production of an antibiotic by one cell can profoundly affect secondary metabolism in neighboring cells occupying natural habitats.
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