Genetic interaction analysis of <i>Candida glabrata</i> transcription factors <i>CST6</i> and <i>UPC2A</i> in the regulation of respiration and fluconazole susceptibility

<i>Candida glabrata</i> is the second most common cause of invasive candidiasis and is widely known to have reduced susceptibility to fluconazole relative to many other <i>Candida</i> spp. Upc2A is a transcription factor that regulates ergosterol biosynthesis gene expression under conditions of sterol stress such as azole drug treatment or hypoxia. Through an <i>in vitro</i> microevolution experiment, we found that loss-of-function mutants of the ATF/CREB transcription factor <i>CST6</i> suppresses the fluconazole hyper-susceptibility of the <i>upc2A</i>∆ mutant. Here, we confirm that the <i>cst6</i>∆ <i>upc2A</i>∆ mutants are resistant to fluconazole but not to hypoxia relative to the <i>upc2A</i>∆ mutant. Sterol analysis of these mutants indicates that this suppression phenotype is not due to restoration of ergosterol levels in the <i>cst6</i>∆ <i>upc2A</i>∆ mutant. Furthermore, increased expression of <i>CDR1</i>, the efflux pump implicated in the vast majority of azole-resistant <i>C. glabrata</i> strains, does not account for the suppression phenotype. Instead, our data suggest that this effect is due in part to increased expression of the adhesin <i>EPA3</i>, which has been shown by others to reduce fluconazole susceptibility in <i>C. glabrata</i>. In addition, we find that loss of both <i>UPC2A</i> and <i>CST6</i> reduces the expression of mitochondrial and respiratory genes and that this also contributes to the suppression phenotype as well as to the resistance of <i>cst6</i>∆ to fluconazole. These latter data further emphasize the connection between mitochondrial function and azole susceptibility.