De-repression of CSP-1 activates adaptive responses to antifungal azoles.

Xi Chen,Wenzhao Wang,Wei Xue,Shaojie Li,Shiping Wei,Xingyu Liu,Zhenying Zhang,Xianyun Sun,Jun Zhou

doi:10.1038/srep19447

Abstract

Antifungal azoles are the major drugs that are used to treat fungal infections. This study found that in response to antifungal azole stress, Neurospora crassa could activate the transcriptional responses of many genes and increase azole resistance by reducing the level of conidial separation 1 (CSP-1), a global transcription repressor, at azole-responsive genes. The expression of csp-1 was directly activated by the transcription factors WC-1 and WC-2. Upon ketoconazole (KTC) stress, the transcript levels of wc-1 and wc-2 were not changed, but csp-1 transcription rapidly declined. A chromatin immunoprecipitation-quantitative polymerase chain reaction analysis revealed a rapid reduction in the WC-2 enrichment at the csp-1 promoter upon KTC treatment, which might be responsible for the KTC-induced csp-1 downregulation. Deletion of csp-1 increased resistance to KTC and voriconazole, while csp-1 overexpression increased KTC susceptibility. CSP-1 transcriptionally repressed a number of azole-responsive genes, including genes encoding the azole target ERG11, the azole efflux pump CDR4, and the sterol C-22 desaturase ERG5. Deletion of csp-1 also reduced the KTC-induced accumulation of ergosterol intermediates, eburicol, and 14α-methyl-3,6-diol. CSP-1 orthologs are widely present in filamentous fungi, and an Aspergillus fumigatus mutant in which the csp-1 was deleted was resistant to itraconazole.

Highlights

Antifungal azoles, including imidazoles and triazoles, are the major drugs used to treat fungal infections
Our previous digital gene expression (DGE) data showed that the azole target-encoding gene erg[11] (NCU02624), the sterol C-22 desaturase-encoding gene erg[5] (NCU05278), and the azole pump-encoding gene cdr[4] (NCU05591) showed dramatic transcriptional increases upon KTC treatment in a wild-type (WT) N. crassa strain[15]
The transcriptional downregulation of csp-1 under KTC stress was confirmed by quantitative real-time polymerase chain reaction

Summary

Introduction

Antifungal azoles, including imidazoles and triazoles, are the major drugs used to treat fungal infections. Fungi are able to adapt to azole stress by altering the expression of many genes. In previous studies of fungal adaptation and resistance to antifungal azoles, the majority of the efforts were focused on genes that were upregulated during azole stress. For most of the genes that were downregulated under azole stress, the effects of their downregulation were not studied in depth. In S. cerevisiae, Nrg1p regulates glucose metabolism and deletion of either NRG1 or NRG2 enhances the resistance of cells to salt and oxidative stress, and decreases tolerance to freezing[13]. Our previously published digital gene expression (DGE) data showed that csp-1 was downregulated by ketoconazole (KTC) stress[15]. We demonstrated that downregulation of csp-1 promoted transcriptional responses by several genes to KTC and conferred resistance to the drug. We showed that deleting either wc-1 or wc-2 increased KTC resistance

Methods

Results

Conclusion