Functional expression of CXCR4 in Saccharomyces cerevisiae in the development of powerful tools for the pharmacological characterization of CXCR4.

Zi xuan Wang, James Broach, Stephen C. Peiper

Research output: Contribution to journalArticle

16 Citations (Scopus)

Abstract

CXCR4, the receptor for stromal cell-derived factor (SDF)-1, was expressed in Saccharomyces cerevisiae, coupled to the pheromone response pathway via a chimeric Galpha subunit. Engagement of CXCR4 by SDF-1 resulted in expression of reporter genes, HIS3 or lacZ, under the transcriptional control of a FUS1 promoter, which is pheromone-responsive. CXCR4 mutants with constitutive signaling activity were generated by random mutagenesis of receptor coding sequences and selection for complementation of histidine auxotrophy in the yeast strain by autonomous expression of the FUS1-HIS3 reporter gene. Linkage of CXCR4 to the pheromone response pathway in yeast provides a system that lends itself to screening of receptor antagonists. The use of constitutively active mutants to screen for inhibitors of the weak partial agonist and inverse agonist pharmacologic types offers a sensitive, efficient approach that is independent of ligand.

Original languageEnglish (US)
Pages (from-to)115-127
Number of pages13
JournalMethods in molecular biology (Clifton, N.J.)
Volume332
StatePublished - Jan 1 2006

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Pheromones
Saccharomyces cerevisiae
Chemokine CXCL12
Pharmacology
Reporter Genes
Yeasts
CXCR4 Receptors
Histidine
Mutagenesis
Ligands

All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Genetics

Cite this

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abstract = "CXCR4, the receptor for stromal cell-derived factor (SDF)-1, was expressed in Saccharomyces cerevisiae, coupled to the pheromone response pathway via a chimeric Galpha subunit. Engagement of CXCR4 by SDF-1 resulted in expression of reporter genes, HIS3 or lacZ, under the transcriptional control of a FUS1 promoter, which is pheromone-responsive. CXCR4 mutants with constitutive signaling activity were generated by random mutagenesis of receptor coding sequences and selection for complementation of histidine auxotrophy in the yeast strain by autonomous expression of the FUS1-HIS3 reporter gene. Linkage of CXCR4 to the pheromone response pathway in yeast provides a system that lends itself to screening of receptor antagonists. The use of constitutively active mutants to screen for inhibitors of the weak partial agonist and inverse agonist pharmacologic types offers a sensitive, efficient approach that is independent of ligand.",
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