Regulation of production of the antifungal metabolite 2,4-diacetylphloroglucinol in Pseudomonas fluorescens F113: genetic analysis of phlF as a transcriptional repressor

The antifungal metabolite 2,4-diacetylphloroglucinol plays a major role in the biocontrol capabilities of Pseudomonas fluorescens. The phloroglucinol biosynthetic locus of P. fluorescens F113 has been isolated previously, Fro m nucleotide sequence data, a putative regulator gene (phlF) was identified upstream and divergently transcribed from the phlACBD phloroglucinol biosy nthetic genes. PhlF shows similarity to various transcriptional repressors in the EMBL database and exhibits a helix-turn-helix motif in its amino aci d sequence. phlF was cloned into an expression vector and the PhlF protein product was purified. Gel retardation experiments demonstrated PhlF to be a DNA-binding protein and showed that it binds to the phlA-phlF intergenic r egion. Introduction of phlF into P. fluorescens F113 in multiple copies res ulted in repression of phloroglucinol production in this strain, This effec t was mediated at the transcription level since the expression of a phlorog lucinol biosynthetic gene fusion in this background was equally repressed, Furthermore, the inactivation of phlF results in derepression of phlorogluc inol production in this strain.