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Streptomyces-derived quorum-sensing systems engineered for adjustable transgene expression in mammalian cells and mice.

ABSTRACT: Prokaryotic transcriptional regulatory elements have been adopted for controlled expression of cloned genes in mammalian cells and animals, the cornerstone for gene-function correlations, drug discovery, biopharmaceutical manufacturing as well as advanced gene therapy and tissue engineering. Many prokaryotes have evolved specific molecular communication systems known as quorum-sensing to coordinate population-wide responses to physiological and/or physicochemical signals. A generic bacterial quorum-sensing system is based on a diffusible signal molecule that prevents binding of a repressor to corresponding operator sites thus resulting in derepression of a target regulon. In Streptomyces, a family of butyrolactones and their corresponding receptor proteins, serve as quorum-sensing systems that control morphological development and antibiotic biosynthesis. Fusion of the Streptomyces coelicolor quorum-sensing receptor (ScbR) to a eukaryotic transactivation domain (VP16) created a mammalian transactivator (SCA) which binds and adjusts transcription from chimeric promoters containing an SCA-specific operator module (P(SPA)). Expression of erythropoietin or the human secreted alkaline phosphatase (SEAP) by this quorum-sensor-regulated gene expression system (QuoRex) could be fine-tuned by non-toxic butyrolactones in a variety of mammalian cells including human primary and mouse embryonic stem cells. Following intraperitoneal implantation of microencapsulated Chinese hamster ovary cells transgenic for QuoRex-controlled SEAP expression into mice, the serum levels of this model glycoprotein could be adjusted to desired concentrations using different butyrolactone dosing regimes.

SUBMITTER: Weber W

PROVIDER: S-EPMC167645 | biostudies-literature | 2003 Jul

REPOSITORIES: biostudies-literature

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Streptomyces-derived quorum-sensing systems engineered for adjustable transgene expression in mammalian cells and mice.

Weber Wilfried W Schoenmakers Ronald R Spielmann Manuela M El-Baba Marie Daoud MD Folcher Marc M Keller Bettina B Weber Cornelia C CC Link Nils N van de Wetering Petra P Heinzen Christoph C Jolivet Benoît B Séquin Urs U Aubel Dominique D Thompson Charles J CJ Fussenegger Martin M

Nucleic acids research 20030701 14

Prokaryotic transcriptional regulatory elements have been adopted for controlled expression of cloned genes in mammalian cells and animals, the cornerstone for gene-function correlations, drug discovery, biopharmaceutical manufacturing as well as advanced gene therapy and tissue engineering. Many prokaryotes have evolved specific molecular communication systems known as quorum-sensing to coordinate population-wide responses to physiological and/or physicochemical signals. A generic bacterial quo ...[more]

PMID: 12853648

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Project description:Quorum sensing (QS) systems communicate bacterial population and stimulate microbial pathogenesis through signaling molecules. Inhibition of QS signals potentially suppresses microbial infections. Antimicrobial properties of Streptomyces have been extensively studied, however, less is known about quorum sensing inhibitory (QSI) activities of Streptomyces. This study explored the QSI potential of Streptomyces isolated from soil. Sixty-five bacterial isolates were purified from soil samples with morphological characteristics of Streptomyces. The three isolates: S6, S12, and S17, exhibited QSI effect by screening with the reporter, Chromobacterium violaceum. Isolate S17 was identified as Streptomyces coelicoflavus by sequencing of the hypervariable regions (V1-V6) of 16S rRNA and was assigned gene bank number KJ855087. The QSI effect of the cell-free supernatant of isolate S17 was not abolished by proteinase K indicating the non-enzymatic activity of QSI components of S17. Three major compounds were isolated and identified, using spectroscopic techniques (1D, 2D NMR, and Mass spectrometry), as behenic acid (docosanoic acid), borrelidin, and 1H-pyrrole-2-carboxylic acid. 1H-pyrrole-2-carboxylic acid inhibited QS and related virulence factors of Pseudomonas aeruginosa PAO1 including; elastase, protease, and pyocyanin without affecting Pseudomonas viability. At the molecular level, 1H-pyrrole-2-carboxylic acid suppressed the expression of QS genes (lasI, lasR, lasA, lasB, rhlI, rhlR, pqsA, and pqsR). Moreover, QSI activity of S17 was assessed under different growth conditions and ISP2 medium supplemented with glucose 0.4% w/v and adjusted at pH 7, showed the highest QSI action. In conclusion, 1H-pyrrole-2-carboxylic acid, one of the major metabolites of Streptomyces isolate S17, inhibited QS and virulence determinants of P. aeruginosa PAO1. The findings of the study open the scope to exploit the in vivo efficacy of this active molecule as anti-pathogenic and anti-virulence of P. aeruginosa.

Dataset Information

Streptomyces-derived quorum-sensing systems engineered for adjustable transgene expression in mammalian cells and mice.

Publications

Streptomyces-derived quorum-sensing systems engineered for adjustable transgene expression in mammalian cells and mice.

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