Conserved Fungal Genes as Potential Targets for Broad-Spectrum Antifungal Drug Discovery

Abstract

ABSTRACT The discovery of novel classes of antifungal drugs depends to a certain extent on the identification of new, unexplored targets that are essential for growth of fungal pathogens. Likewise, the broad-spectrum capacity of future antifungals requires the target gene(s) to be conserved among key fungal pathogens. Using a genome comparison (or concordance) tool, we identified 240 conserved genes as candidates for potential antifungal targets in 10 fungal genomes. To facilitate the identification of essential genes in Candida albicans , we developed a repressible C. albicans MET3 (Ca MET3 ) promoter system capable of evaluating gene essentiality on a genome-wide scale. The Ca MET3 promoter was found to be highly amenable to controlled gene expression, a prerequisite for use in target-based whole-cell screening. When the expression of the known antifungal target C. albicans ERG1 was reduced via down-regulation of the Ca MET3 promoter, the Ca ERG1 conditional mutant strain became hypersensitive, specifically to its inhibitor, terbinafine. Furthermore, parallel screening against a small compound library using the Ca ERG1 conditional mutant under normal and repressed conditions uncovered several hypersensitive compound hits. This work therefore demonstrates a streamlined process for proceeding from selection and validation of candidate antifungal targets to screening for specific inhibitors.