The identification of a gene family in the Saccharomyces cerevisiae ergosterol biosynthesis pathway

Margaret H.Lai, Martin Bard, Charles A.Pierson, John F.Alexander, Mark Goebl, Guy T. Carter, Donald R. Kirsch

Research output: Contribution to journalArticle

57 Scopus citations

Abstract

The Saccharomyces cerevisiae ERG24 gene, encoding sterol Δ14 reductase (Erg24p), was cloned by selecting strains carrying sequences on a 2μ-based vector for resistance to the morpholine fungicide, fenpropimorph (Fp). Four distinct plasmid inserts which conferred Fp resistance (FpR) were recovered (plasmids pML99, pML100, pML101 and pML103). Although Fp is reported to inhibit activity of Erg24p and sterol Δ8-Δ7 isomerase (Erg2p; encoded by ERG2), none of the inserts had restriction maps resembling ERG2. In addition, a 2μ plasmid overexpression of the ERG2 sequence did not produce FpR. Characterization studies were focused on plasmid pML100, because it was the only plasmid to confer FpR consistently when tested in a number of different genetic backgrounds. Tests with a panel of fungicides indicated that pML100 conferred significant resistance only to compounds (Fp, tridemorph, fenpropidin and azasterol) which have a shared site of action, Erg24p. An insertional disruption of pML100 resulted in an obligate anaerobic phenotype, indicating a lesion in sterol biosynthesis. Sterol analysis of the disrupted mutant demonstrated the accumulation of ignosterol, indicating a loss of Erg24p activity. A SphI-XbaI fragment of pML100 was sequenced, revealing the presence of an ORF encoding a 438-amino-acid protein, which is highly similar to those encoded by two previously reported yeast drug sensitivity genes, sts1+ (Schizosaccharomyces pombe)and YGL022 (S. cerevisiae). Analyses of these genes demonstrated that strains carrying disruptions of sts1+ or YGL022 have ergosterol biosynthesis defects in the enzyme, sterol C-24(28) reductase (Erg4p; encoded by ERG4). In addition, YGL022 complemented an erg4 mutation in S. cerevisiae and a YGL022 gene disruption showed allelism with an erg4 mutation, providing further support that YGL022 encodes Erg4p.

Original languageEnglish (US)
Pages (from-to)41-49
Number of pages9
JournalGene
Volume140
Issue number1
DOIs
StatePublished - 1994

Keywords

  • ERG24
  • ERG4
  • fenpropimorph
  • fungicide
  • ignosterol
  • morpholine
  • sterol A14 reductase
  • Sterol biosynthesis
  • sterol C-24(28) reductase
  • tetrad analysis
  • tridemorph

ASJC Scopus subject areas

  • Genetics

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  • Cite this

    H.Lai, M., Bard, M., A.Pierson, C., F.Alexander, J., Goebl, M., Carter, G. T., & Kirsch, D. R. (1994). The identification of a gene family in the Saccharomyces cerevisiae ergosterol biosynthesis pathway. Gene, 140(1), 41-49. https://doi.org/10.1016/0378-1119(94)90728-5