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Microbiology 147 (2001), 2127-2139
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Microbiology (2001), 147, 2127-2139.
© 2001 Society for General Microbiology

Genetics and Molecular Biology

Quorum-sensing-dependent regulation of biosynthesis of the polyketide antibiotic mupirocin in Pseudomonas fluorescens NCIMB 10586

A. Kassem El-Sayed1, Joanne Hothersall1 and Christopher M. Thomas1

School of Biosciences, University of Birmingham, Edgbaston, Birmingham B15 2TT, UK1

Author for correspondence: Christopher M. Thomas. Tel: +44 121 414 5903. Fax: +44 121 414 5925. e-mail: c.m.thomas{at}bham.ac.uk

Mupirocin (pseudomonic acid) is a polyketide antibiotic, targeting isoleucyl-tRNA synthase, and produced by Pseudomonas fluorescens NCIMB 10586. It is used clinically as a topical treatment for staphylococcal infections, particularly in contexts where there is a problem with methicillin-resistant Staphylococcus aureus (MRSA). In studying the mupirocin biosynthetic cluster the authors identified two putative regulatory genes, mupR and mupI, whose predicted amino acid sequences showed significant identity to proteins involved in quorum-sensing-dependent regulatory systems such as LasR/LuxR (transcriptional activators) and LasI/LuxI (synthases for N-acylhomoserine lactones – AHLs – that activate LasR/LuxR). Inactivation by deletion mutations using a suicide vector strategy confirmed the requirement for both genes in mupirocin biosynthesis. Cross-feeding experiments between bacterial strains as well as solvent extraction showed that, as predicted, wild-type P. fluorescens NCIMB 10586 produces a diffusible substance that overcomes the defect of a mupI mutant. Use of biosensor strains showed that the MupI product can activate the Pseudomonas aeruginosa lasRlasI system and that P. aeruginosa produces one or more compounds that can replace the MupI product. Insertion of a xylE reporter gene into mupA, the first ORF of the mupirocin biosynthetic operon, showed that together mupR/mupI control expression of the operon in such a way that the cluster is switched on late in exponential phase and in stationary phase.

Keywords: N-acylhomoserine lactone, Staphylococcus aureus, reporter genes, genetic manipulation, diffusible activator

Abbreviations: AHL, N-acylhomoserine lactone; C4-HSL, N-butyryl-L-homoserine lactone; C6-HSL, N-hexanoyl-L-homoserine lactone; 3-oxo-C6-HSL, N-(3-oxohexanoyl)-L-homoserine lactone; C10-HSL, N-decanoyl-L-homoserine lactone; 3-OH-C6-HSL, N-(3-hydroxyhexanoyl)-L-homoserine lactone; 3-OH-C8-HSL, N-(3 hydroxyoctanoyl)-L-homoserine lactone; 3-OH-C10-HSL, N-(3 hydroxydecanoyl)-L-homoserine lactone; 3-oxo-C12-HSL, N-(3-oxododecanoyl)-L-homoserine lactone; 3-OH-C14:1-HSL, N-(3-hydroxytetradecenoyl)-L-homoserine lactone

The GenBank accession numbers for the sequences determined in this work are AF318063 (mupA), AF318064 (mupR) and AF318065 (mupI).




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