ABSTRACTAntifungal agents exert their activity through a variety of mechanisms, some of which are poorly understood. We examined changes in the gene expression profile ofCandida albicansfollowing 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, includingNCP1,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, includingCDR1,CDR2,IFD4,DDR48, andRTA3. Amphotericin B produced changes in the expression of genes involved in small-molecule transport (ENA21), and in cell stress (YHB1,CTA1,AOX1, andSOD2). Also observed was decreased expression of genes involved in ergosterol biosynthesis, includingERG3andERG11. Caspofungin produced changes in expression of genes encoding cell wall maintenance proteins, including the β-1,3-glucan synthase subunitGSL22, as well asPHR1,ECM21,ECM33, andFEN12. Flucytosine increased the expression of proteins involved in purine and pyrimidine biosynthesis, includingYNK1,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 onC. albicans.