title: Contributions of both ATP-Binding Cassette Transporter and Cyp51A Proteins Are Essential for Azole Resistance in Aspergillus fumigatus.
abstract: While azole drugs targeting the biosynthesis of ergosterol are effective antifungal agents, their extensive use has led to the development of resistant organisms. Infections involving azole-resistant forms of the filamentous fungusAspergillus fumigatusare often associated with genetic changes in thecyp51Agene encoding the lanosterol α14 demethylase target enzyme. Both a sequence duplication in thecyp51Apromoter (TR34) and a substitution mutation in the coding sequence (L98H) are required for the full expression of azole resistance. A mechanism commonly observed in pathogenic yeast such asCandida albicansinvolves gain-of-function mutations in transcriptional regulatory proteins that induce expression of genes encoding ATP-binding cassette (ABC) transporters. We and others have found that an ABC transporter protein called Cdr1B (here referred to as AbcG1) is required for wild-type azole resistance inA. fumigatusHere, we test the genetic relationship between the TR34 L98H allele ofcyp51Aand anabcG1null mutation. Loss of AbcG1 from a TR34 L98Hcyp51A-containing strain caused a large decrease in the azole resistance of the resulting double-mutant strain. We also generated antibodies that enabled the detection of both the wild-type and L98H forms of the Cyp51A protein. The introduction of the L98H lesion into thecyp51Agene led to a decreased production of immunoreactive enzyme, suggesting that this mutant protein is unstable. Our data confirm the importance of AbcG1 function during azole resistance even in a strongly drug-resistant background.
authors.affiliation: Department of Molecular Physiology and Biophysics, Carver College of Medicine, University of Iowa, Iowa City, Iowa, USA.
authors.forename: Sanjoy
authors.initials: S
authors.lastname: Paul
authors.affiliation: Department of Internal Medicine, Carver College of Medicine, University of Iowa, Iowa City, Iowa, USA.
authors.forename: Daniel
authors.initials: D
authors.lastname: Diekema
authors.affiliation: Department of Molecular Physiology and Biophysics, Carver College of Medicine, University of Iowa, Iowa City, Iowa, USA scott-moye-rowley@uiowa.edu.
authors.forename: W Scott
authors.initials: WS
authors.lastname: Moye-Rowley
mesh_terms: D000070998:ATP Binding Cassette Transporter, Subfamily G, Member 1 / Q000235:genetics*; D000935:Antifungal Agents / Q000494:pharmacology*; D001228:Aspergillosis / Q000188:drug therapy / Q000382:microbiology; D001232:Aspergillus fumigatus / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D001393:Azoles / Q000494:pharmacology*; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics*; D025141:Drug Resistance, Fungal / Q000235:genetics*; D005656:Fungal Proteins / Q000235:genetics*; D017353:Gene Deletion; D008826:Microbial Sensitivity Tests; D011401:Promoter Regions, Genetic / Q000235:genetics; D065819:Voriconazole / Q000494:pharmacology
chemical_list: D000070998:ATP Binding Cassette Transporter, Subfamily G, Member 1; D000935:Antifungal Agents; D001393:Azoles; D005656:Fungal Proteins; D003577:Cytochrome P-450 Enzyme System; C465526:cytochrome P-450 CYP51A, Aspergillus; D065819:Voriconazole
keywords: ATP-binding cassette transporter; Aspergillus fumigatus; Western blotting; azole resistance; cyp51A; genetic analysis