In Candida albicans, Upc2 is a zinc-cluster transcription factor that targets genes including those of the ergosterol biosynthesis pathway. To date there have been three documented UPC2 gain-of-function (GOF) mutations recovered from fluconazole-resistant clinical isolates that contribute to an increase in ERG11 expression and decreased fluconazole susceptibility. In a group of 62 fluconazole-resistant isolates, we found that 47 of these overexpressed ERG11 by at least two-fold over that of an average expression of 3 unrelated fluconazole susceptible strains. Of those 47 isolates, 29 contained a mutation in UPC2, whereas the remaining 18 isolates did not. Of the isolates containing mutations in UPC2, we recovered eight distinct mutations resulting in single putative amino acid substitutions: G648D, G648S, A643T, A643V, Y642F, G304R, A646V and W478C. Seven of these resulted in increased ERG11 expression, increased cellular ergosterol, and decreased susceptibility to fluconazole as compared to the wild-type strain. Genome-wide transcriptional analysis was performed for the four strongest Upc2 amino acid substitutions (A643V, G648D, G648S and Y642F). Genes commonly upregulated in all four mutations included those involved in ergosterol biosynthesis, in oxidoreductase activity, the major facilitator efflux pump encoded by the MDR1 gene, and the uncharacterized ATP binding cassette transporter CDR11. These findings demonstrate that gain-of-function mutations in UPC2 are more prevalent than previously thought among clinical isolates, make a significant contribution to azole antifungal resistance, but do not account for ERG11 overexpression in all such isolates of C. albicans.
Overall design: We examined the expression of genes in response to the presence of 4 gain-of-function alleles of the zinc-cluster transcription factor Upc2. The global gene expression of each mutant Upc2 strain was compared to that of the wildtype strain SC5314.
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