title: Click beetle luciferases as dual reporters of gene expression in Candida albicans.
abstract: Synthetic genes encoding functional luciferases of the click beetle (CB) Pyrophorus plagiophthalamus have been expressed in the human fungal pathogen Candida albicans. Both green- and red-emitting CB luciferases (CaCBGluc and CaCBRluc) were produced with high efficiency in transformants under transcriptional control of the growth-dependent ACT1 promoter, as well as by the HWP1 and UME6 promoters, which are upregulated during hyphal morphogenesis, as well as by the YWP1 and EFG1 promoters, which are downregulated. For all hyphally regulated genes, relative bioluminescence values derived from promoter fusions approximated relative transcript levels of native genes, although downregulation of YWP1 promoter activity required correction for the stability of CB luciferases (approximate half-lives 30 min for CaCBRluc and 80 min for CaCBGluc, as determined by immunoblotting). Importantly, the activity of both luciferases could be separately monitored in a single strain, in intact cells, in lysed cells or in cell extracts using luciferin as single substrate and inhibition of hypha formation by farnesol could be easily detected by the HWP1p-CaCBRluc fusion. The results suggest that CB luciferases are convenient tools to measure gene expression in C. albicans and may facilitate screenings for antifungal compounds.
authors.affiliation: Department Biologie, Molekulare Mykologie, Heinrich-Heine-Universität Düsseldorf, 1/26 12, Duesseldorf, D-40225, Germany.
authors.forename: Mario
authors.initials: M
authors.lastname: Kapitan
authors.affiliation: Department Biologie, Molekulare Mykologie, Heinrich-Heine-Universität Düsseldorf, 1/26 12, Duesseldorf, D-40225, Germany.
authors.forename: Isabel
authors.initials: I
authors.lastname: Eichhof
authors.affiliation: Department Biologie, Molekulare Mykologie, Heinrich-Heine-Universität Düsseldorf, 1/26 12, Duesseldorf, D-40225, Germany.
authors.forename: Quentin
authors.initials: Q
authors.lastname: Lagadec
authors.affiliation: Department Biologie, Molekulare Mykologie, Heinrich-Heine-Universität Düsseldorf, 1/26 12, Duesseldorf, D-40225, Germany.
authors.forename: Joachim F
authors.initials: JF
authors.lastname: Ernst
mesh_terms: D000199:Actins / Q000235:genetics; D000818:Animals; D002176:Candida albicans / Q000235:genetics / Q000378:metabolism*; D001517:Coleoptera / Q000201:enzymology*; D004268:DNA-Binding Proteins / Q000235:genetics; D005656:Fungal Proteins / Q000235:genetics; D015870:Gene Expression / Q000235:genetics*; D015966:Gene Expression Regulation, Fungal / Q000235:genetics*; D008156:Luciferases / Q000235:genetics*; D008163:Luminescent Measurements; D008562:Membrane Glycoproteins / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics; D014157:Transcription Factors / Q000235:genetics
chemical_list: C404682:ACT1 protein, Candida albicans; D000199:Actins; D004268:DNA-Binding Proteins; C121438:EFG1 protein, Candida albicans; D005656:Fungal Proteins; C118102:HWP1 protein, Candida albicans; D008562:Membrane Glycoproteins; D014157:Transcription Factors; D008156:Luciferases