title: Expression of a reporter gene interrupted by the Candida albicans group I intron is inhibited by base analogs.
abstract: We previously reported the identification of an intron (CaLSU) in the 25S ribosomal RNA of some Candida albicans yeast strains. CaLSU was shown to self-splice and has the potential to adopt a secondary structure typical of group I introns. The presence of CaLSU inC. albicans strains correlates with a high degree of susceptibility to base analog antifungal agents, 5-fluorocytosine (5-FC) or 5-fluorouracil (5-FU). Cell death, resulting from addition of base analogs to growing cultures, precluded demonstration of a causal relationship between CaLSU presence and susceptibility to base analogs. In the present study, CaLSU was inserted in a non-essential lacZ reporter gene and expression was examined in Saccharomyces cerevisiae. Different mutations affecting in vitro self-splicing also had similar effects on reporter gene expression in vivo. This indicates that in vivo removal of CaLSU from the reporter gene occurs through the typical self-splicing mechanism of group I introns. Base analogs inhibited expression of the reporter gene product in a concentration-dependent manner upon their addition to the cultures. This supports a model in which disruption of intron secondary structure, consecutive to the incorporation of nucleotide analogs, is a major factor determining the susceptibility of C.albicans cells to base analogs.
authors.affiliation: Département de Microbiologie et Immunologie, Université de Montréal, PO Box 6128, Station Centre-Ville, Montréal, Québec H3C 3J7, Canada.
authors.forename: S
authors.initials: S
authors.lastname: Mercure
authors.forename: L
authors.initials: L
authors.lastname: Cousineau
authors.forename: S
authors.initials: S
authors.lastname: Montplaisir
authors.forename: P
authors.initials: P
authors.lastname: Belhumeur
authors.forename: G
authors.initials: G
authors.lastname: Lemay
mesh_terms: D000935:Antifungal Agents / Q000494:pharmacology; D015152:Blotting, Northern; D015153:Blotting, Western; D002176:Candida albicans / Q000737:chemistry*; D002455:Cell Division / Q000187:drug effects; D005437:Flucytosine / Q000494:pharmacology; D005472:Fluorouracil / Q000494:pharmacology; D015966:Gene Expression Regulation, Fungal / Q000235:genetics*; D017930:Genes, Reporter / Q000235:genetics; D007438:Introns / Q000235:genetics*; D007763:Lac Operon / Q000235:genetics; D016297:Mutagenesis, Site-Directed / Q000235:genetics; D009690:Nucleic Acid Conformation; D012326:RNA Splicing / Q000235:genetics; D012335:RNA, Ribosomal / Q000235:genetics; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
chemical_list: D000935:Antifungal Agents; D012335:RNA, Ribosomal; C065039:RNA, ribosomal, 25S; D005437:Flucytosine; D001616:beta-Galactosidase; D005472:Fluorouracil