Fluconazole resistance and expression in mediated by the hyperactive Tac1-5 transcriptional activator requires Tlo proteins.

is an opportunistic fungal pathogen associated with superficial and systemic infections in humans. Azole antifungal resistance in is of clinical concern, and both oral and systemic infections can be difficult to treat due to the lack of alternative antifungal drugs. Expression of a hyperactive form of the transcription factor Tac1 is a major contributor to azole resistance in isolates resulting in the increased expression of the azole efflux pump Cdr1. In this study, we investigated whether the Mediator tail component Med2, encoded by the expanded (n=14) gene family of , was required for Tac1 activity. A homozygous gain-of-function point mutation was introduced into WT, Δ and Δ strains of which enables them to express hyperactive Tac1. qRT-PCR analysis revealed that Δ- had reduced basal and fluphenazine-induced expression relative to WT- strains and exhibited reduced levels of resistance to fluconazole and terbinafine. Individual copies of representatives from each of the alpha, beta and gamma clades were reintroduced into Δ- to investigate their ability to restore Tac1-activated resistance. These studies show that alpha and beta genes could restore fluconazole resistance in the Δ- background, whereas gamma clade genes did not result in any detectable phenotypic complementation. Transcript profiling showed that reintroduction of α led to increased expression of -activated genes such as . Further analysis using ChIP-qPCR revealed that Tloα1 localizes to the drug response element which is the site where Tac1 binds to the promoter. These data have identified that the gene family is required for the expression of Tac1-mediated fluconazole resistance. However, this effect is confined to members of the alpha and beta, but not the gamma, clades.