title: New Fks hot spot for acquired echinocandin resistance in Saccharomyces cerevisiae and its contribution to intrinsic resistance of Scedosporium species.
abstract: Echinocandins represent a new antifungal group with potent activity against Candida species. These lipopeptides inhibit the synthesis of β-1,3-glucan, the major cell wall polysaccharide. Acquired resistance or reduced echinocandin susceptibility (RES) is rare and associated with mutations in two "hot spot" regions of Fks1 or Fks2, the probable β-1,3-glucan synthases. In contrast, many fungi demonstrate intrinsic RES for reasons that remain unclear. We are using Saccharomyces cerevisiae to understand the basis for RES by modeling echinocandin-Fks interaction. Previously characterized mutations confer cross-RES; we screened for mutations conferring differential RES, implying direct interaction of that Fks residue with a variable echinocandin side chain. One mutant (in an fks1Δ background) exhibited ≥16-fold micafungin and anidulafungin versus caspofungin RES. Sequencing identified a novel Fks2 mutation, W714L/Y715N. Equivalent W695L/Y696N and related W695L/F/C mutations in Fks1 generated by site-directed mutagenesis and the isolation of a W695L-equivalent mutation in Candida glabrata confirmed the role of the new "hot spot 3" in RES. Further mutagenesis expanded hot spot 3 to Fks1 residues 690 to 700, yielding phenotypes ranging from cross-RES to differential hypersusceptibility. Fks1 sequences from intrinsically RES Scedosporium species revealed W695F-equivalent substitutions; Fks1 hybrids expressing Scedosporium prolificans hot spot 3 confirmed that this substitution imparts RES.
authors.affiliation: Department of Microbiology and Immunology, Drexel University College of Medicine, Philadelphia, PA 19129-1033, USA.
authors.forename: Michael E
authors.initials: ME
authors.lastname: Johnson
authors.forename: Santosh K
authors.initials: SK
authors.lastname: Katiyar
authors.forename: Thomas D
authors.initials: TD
authors.lastname: Edlind
mesh_terms: D000077612:Anidulafungin; D000935:Antifungal Agents / Q000494:pharmacology*; D001483:Base Sequence; D041221:Candida glabrata / Q000187:drug effects / Q000235:genetics; D000077336:Caspofungin; D002473:Cell Wall / Q000737:chemistry / Q000187:drug effects; D025141:Drug Resistance, Fungal / Q000235:genetics*; D054714:Echinocandins / Q000235:genetics / Q000494:pharmacology; D005964:Glucosyltransferases / Q000037:antagonists & inhibitors / Q000235:genetics; D055666:Lipopeptides / Q000494:pharmacology; D008565:Membrane Proteins / Q000235:genetics; D000077551:Micafungin; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D009154:Mutation; D011509:Proteoglycans; D012441:Saccharomyces cerevisiae / Q000187:drug effects* / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics; D021681:Scedosporium / Q000187:drug effects* / Q000235:genetics*; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D047071:beta-Glucans / Q000378:metabolism
chemical_list: D000935:Antifungal Agents; D054714:Echinocandins; D055666:Lipopeptides; D008565:Membrane Proteins; D011509:Proteoglycans; D029701:Saccharomyces cerevisiae Proteins; D047071:beta-Glucans; C010770:polysaccharide-K; D000077612:Anidulafungin; D005964:Glucosyltransferases; C040232:glucan synthase; C091618:FKS1 protein, S cerevisiae; C095724:GSC2 protein, S cerevisiae; D000077336:Caspofungin; D000077551:Micafungin