Abstract
Constitutive overexpression of the MDR1 (multidrug resistance) gene, which encodes a multidrug efflux pump of the major facilitator superfamily, is a frequent cause of resistance to fluconazole and other toxic compounds in clinical Candida albicans strains, but the mechanism of MDR1 upregulation has not been resolved. By genome-wide gene expression analysis we have identified a zinc cluster transcription factor, designated as MRR1 (multidrug resistance regulator), that was coordinately upregulated with MDR1 in drug-resistant, clinical C. albicans isolates. Inactivation of MRR1 in two such drug-resistant isolates abolished both MDR1 expression and multidrug resistance. Sequence analysis of the MRR1 alleles of two matched drug-sensitive and drug-resistant C. albicans isolate pairs showed that the resistant isolates had become homozygous for MRR1 alleles that contained single nucleotide substitutions, resulting in a P683S exchange in one isolate and a G997V substitution in the other isolate. Introduction of these mutated alleles into a drug-susceptible C. albicans strain resulted in constitutive MDR1 overexpression and multidrug resistance. By comparing the transcriptional profiles of drug-resistant C. albicans isolates and mrr1Δ mutants derived from them and of C. albicans strains carrying wild-type and mutated MRR1 alleles, we defined the target genes that are controlled by Mrr1p. Many of the Mrr1p target genes encode oxidoreductases, whose upregulation in fluconazole-resistant isolates may help to prevent cell damage resulting from the generation of toxic molecules in the presence of fluconazole and thereby contribute to drug resistance. The identification of MRR1 as the central regulator of the MDR1 efflux pump and the elucidation of the mutations that have occurred in fluconazole-resistant, clinical C. albicans isolates and result in constitutive activity of this trancription factor provide detailed insights into the molecular basis of multidrug resistance in this important human fungal pathogen.
Highlights
The yeast Candida albicans is usually a harmless commensal in many healthy people where it resides on mucosal surfaces of the gastrointestinal and urogenital tract, but it can cause superficial as well as life-threatening systemic infections, especially in immunocompromised patients [1]
By comparing the transcription profiles of MDR1 overexpressing, clinical C. albicans isolates and matched, drugsusceptible isolates from the same patients, we identified a transcription factor, termed multidrug resistance regulator 1 (MRR1), which was upregulated in all resistant isolates and turned out to be a central regulator of MDR1 expression
Inactivation of MRR1 in clinical isolates abolished MDR1 expression and affected fluconazole resistance even more strongly than deletion of the MDR1 efflux pump itself, indicating that additional Mrr1p target genes, which were identified by genome-wide gene expression analysis, contribute to fluconazole resistance
Summary
The yeast Candida albicans is usually a harmless commensal in many healthy people where it resides on mucosal surfaces of the gastrointestinal and urogenital tract, but it can cause superficial as well as life-threatening systemic infections, especially in immunocompromised patients [1]. C. albicans can develop resistance to fluconazole, especially during long-term treatment of oropharyngeal candidiasis, which frequently affects HIV-infected persons and AIDS patients [2]. Molecular fingerprinting of serial C. albicans isolates from recurrent episodes of oropharyngeal candidiasis has shown that fluconazole resistance usually develops in previously susceptible strains, and such serial isolates from the same patient, so-called matched isolates, have proved an excellent tool to study the molecular basis of drug resistance [3,4,5,6,7,8,9,10].
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