Streptomyces-derived quorum-sensing systems engineered for adjustable transgene expression in mammalian cells and mice.
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.
Weber
Wilfried
W
Institute of Biotechnology, Swiss Federal Institute of Technology, ETH Hoenggerberg, CH-8093 Zurich, Switzerland.
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
D015107:4-Butyrolactone / Q000096:biosynthesis* / Q000494:pharmacology; D000818:Animals; D001426:Bacterial Proteins; D001667:Binding, Competitive; D016466:CHO Cells; D002460:Cell Line; D006224:Cricetinae; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D005260:Female; D005786:Gene Expression Regulation / Q000187:drug effects; D005818:Genetic Engineering / Q000379:methods*; D006801:Humans; D051379:Mice; D008822:Mice, Transgenic; D010957:Plasmids / Q000235:genetics; D013302:Streptomyces / Q000235:genetics* / Q000378:metabolism; D014162:Transfection; D014407:Tumor Cells, Cultured
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; C446304:ScbR protein, Streptomyces coelicolor; D015107:4-Butyrolactone