Genome-wide expression profiling of the response to azole, polyene, echinocandin, and pyrimidine antifungal agents in Candida albicans

Abstract

Antifungal agents exert their activity through a variety of mechanisms, some of which are poorly understood. We examined changes in the gene expression profile of Candida albicans following exposure to representatives of the four currently available classes of antifungal agents used in the treatment of systemic fungal infections. Ketoconazole exposure increased expression of genes involved in lipid, fatty acid, and sterol metabolism, including NCP1, MCR1, CYB5, ERG2, ERG3, ERG10, ERG25, ERG251, and that encoding the azole target, ERG11. Ketoconazole also increased expression of several genes associated with azole resistance, including CDR1, CDR2, IFD4, DDR48, and RTA3. Amphotericin B produced changes in the expression of genes involved in small-molecule transport (ENA21), and in cell stress (YHB1, CTA1, AOX1, and SOD2). Also observed was decreased expression of genes involved in ergosterol biosynthesis, including ERG3 and ERG11. Caspofungin produced changes in expression of genes encoding cell wall maintenance proteins, including the beta-1,3-glucan synthase subunit GSL22, as well as PHR1, ECM21, ECM33, and FEN12. Flucytosine increased the expression of proteins involved in purine and pyrimidine biosynthesis, including YNK1, FUR1, and that encoding its target, CDC21. Real-time reverse transcription-PCR was used to confirm microarray results. Genes responding similarly to two or more drugs were also identified. These data shed new light on the effects of these classes of antifungal agents on C. albicans.

FIG. 1.

Distribution of KTZ-responsive genes. Genes were annotated and assigned to functional categories. The average expression ratio from two independent experiments is shown. Genes shaded in gray are indicative of major responses associated with the drug's mechanism of action. Genes in boldface have been previously associated with azole resistance.

FIG. 2.

Distribution of AMB-responsive genes. Gene annotations and expression values are as in Fig. 1. Genes shaded in gray are indicative of major responses associated with the drug's mechanism of action.

FIG. 3.

Distribution of CPF-responsive genes. Gene annotations and expression values are as in Fig. 1. Genes shaded in gray are indicative of major responses associated with the drug's mechanism of action.

FIG. 4.

Distribution of 5-FC-responsive genes. Gene annotations and expression values are as in Fig. 1. Genes shaded in gray are indicative of major responses associated with the drug's mechanism of action.

FIG. 5.

Quantitative real-time RT-PCR analysis of genes identified as differentially expressed by microarray experiments. Eight genes identified as differentially expressed by microarray analysis were examined by quantitative real-time RT-PCR with gene-specific primers. Data are shown as mean ± S.D. (A) Gene expression changes in 8 genes (2 per drug) detected by real-time RT-PCR. (B) Validation of drug-specific gene expression responses.