Genomic profiling of the response of Candida albicans to itraconazole treatment using a DNA microarray

Abstract

The application of genome-wide expression profiling to determine how drugs achieve their therapeutic effect has provided the pharmaceutical industry with an exciting new tool for drug mode-of-action studies. We used DNA chip technology to study cellular responses to perturbations of ergosterol biosynthesis caused by the broad-spectrum antifungal agent itraconazole. Simultaneous examination of over 6,600 Candida albicans gene transcript levels, representing the entire genome, upon treatment of cells with 10 microM itraconazole revealed that 296 genes were responsive. For 116 genes transcript levels were decreased at least 2.5-fold, while for 180 transcript levels were similarly increased. A global upregulation of ERG genes in response to azole treatment was observed. ERG11 and ERG5 were found to be upregulated approximately 12-fold. In addition, a significant upregulation was observed for ERG6, ERG1, ERG3, ERG4, ERG10, ERG9, ERG26, ERG25, ERG2, IDII, HMGS, NCP1, and FEN2, all of which are genes known to be involved in ergosterol biosynthesis. The effects of itraconazole on a wide variety of known metabolic processes are discussed. As over 140 proteins with unknown function were responsive to itraconazole, our analysis might provide-in combination with phenotypic data-first hints of their potential function. The present report is the first to describe the application of DNA chip technology to study the response of a major human fungal pathogen to drug treatment.

FIG. 1

Correlation graph. Normalized hybridization signals from one representative experiment from Cy5 (untreated wild-type C. albicans cells) and Cy3 (itraconazole-treated C. albicans cells) channels were plotted against each other on a logarithmic scale. The data series have a correlation coefficient of 0.93. Datum points with differential expression less than 2.5-fold fall between the two dotted lines. The dashed line is the perfect diagonal.

FIG. 2

Pie chart grouping responsive (more than 2.5-fold change) genes in functional classes (n = 296); 5% of the responsive genes belong to the ergosterol biosynthetic pathway, and 4% are associated with lipid and fatty acid metabolism. Forty-one percent of the responsive genes are of unknown function.

FIG. 3

Effect of itraconazole (24-h treatment; final concentration, 10 μM) on expression of C. albicans genes involved in the ergosterol biosynthetic pathway; responsive genes are shown in bold; fold changes are shown in parentheses. ∗, not present on DNA chip. CoA, coenzyme A; HMG, 3-hydroxy-3-methylglutaryl.

FIG. 3

Effect of itraconazole (24-h treatment; final concentration, 10 μM) on expression of C. albicans genes involved in the ergosterol biosynthetic pathway; responsive genes are shown in bold; fold changes are shown in parentheses. ∗, not present on DNA chip. CoA, coenzyme A; HMG, 3-hydroxy-3-methylglutaryl.