Mutations in drive increased and expression and azole resistance in .

has emerged as a fungal pathogen of particular concern owing in part to its propensity to exhibit antifungal resistance, especially to the commonly prescribed antifungal fluconazole. A mutation in , which encodes a zinc cluster transcription factor, has been shown to confer increased resistance to fluconazole. In this work, we aimed to determine how mutations in exert this effect. mutations leading to A640V, A657V, and F862_N866del, found in fluconazole-resistant clinical isolates, were introduced into two susceptible Clade I backgrounds using CRISPR-Cas9 gene editing. These mutations conferred increased fluconazole resistance, as well as increased resistance to other triazoles as measured by broth microdilution. RNA-seq revealed that the ATP-binding cassette (ABC) transporter gene as well as the major facilitator superfamily (MFS) transporter gene were both upregulated in the presence of these mutations. Disruption of increased susceptibility in strains with mutations, whereas disruption of had little to no effect. However, disruption of both and resulted in an additional increase in susceptibility as compared with disruption of alone. mutations, leading to A640V, A657V, and F862_N866del all result in increased resistance to fluconazole and other triazole antifungals and increased expression of both and in . Together, these data suggest is the primary driver of resistance conferred by these mutations.