<I>Serratia</I> sp. strain ATCC 39006 ('<I>39006</I>') produces a bright red pigment prodigiosin (Pig), and a carbapenem antibiotic, 1-carbapen-2-em-3-carboxylic acid (Car). Recently it was shown that <I>39006</I> also produced N-acylhomoserine lactones (AHL), which act as quorum sensing (QS) signalling molecules. The notion that <I>39006</I> used a QS system to regulate phenotypes according to cell-density was investigated and characterised here. <I>N</I>-butanoyl-L-homoserine lactone (BHL) and <I>N</I>-hexanoyl-L-homoserine lactone (HHL) were identified as the QS signalling molecules by thin-layer chromatography. Ahl<SUP>-</SUP> mutants carrying mini-transposon insertions in a single ORF (<I>smaI</I>) were concomitantly deficient in the production of Pig, Car, 'white-opaque phenotype' and the synthesis of the extracellular enzymes pectate lyase and cellulase, all of which were restored by the addition of synthetic AHLs. All five phenotypes were also restored in the <I>smaI</I><SUP>-</SUP> mutant when grown with or near a BHL/HHL-producing strain. It was therefore concluded that <I>39006</I> possessed a previously uncharacterised QS system that played a central role in exoenzyme and secondary metabolite production. The <I>smaI</I> gene was sequenced and found to encode a protein (SmaI) similar to members of the LuxI family of AHL synthases. DNA sequencing also revealed another ORF, <I>smaR</I>, which encoded a protein (SmaR) similar to the LuxR family of QS response regulator proteins. The regulation of Pig and Car production with respect to quorum sensing was investigated in this study using <I>lacZ</I> transcriptional fusions to <I>pigA, pigH</I> (<I>pig</I> biosynthetic gene cluster) and <I>carA</I> (<I>car</I> biosynthetic gene cluster). Data supported a model in which the <I>smaIR</I> locus formed a complete regulatory switch where SmaR acted as a repressor of gene transcription at low cell density. At high cell-density, in the presence of AHLs, SmaR-mediated repression was lifted thus allowing transcription.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:598191 |
| Date | January 2002 |
| Creators | Crow, M. A. |
| Publisher | University of Cambridge |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
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