Project description:Genome sequence data results are reported from experimental and bioinfomatic work using the technique 'Bulk Segregant Analysis' to determine the genetic basis of observed resistance to the azole antifungal compound itraconazole in the opportunistic fungal pathogen Aspergillus fumigatus.
Project description:The Negative cofactor 2 (NCT) complex is an evolutionally conserved heterodimeric transcription factor. In Aspergillus fumigatus, the NCT complex consists of two subunits NctA and NctB. Through a genome-wide screening of a transcription factor null mutant strains, we found that loss of the NCT complex leads to a multi-drug resistance phenotype including the azoles (itraconazole, voriconazole and posaconazole) as well as the salvage therapeutic amphotericin B, and terbinafine. To obtain further insight into the molecular mechanisms driving the azole-resistance in the NCT complex null mutants, we analyzed genome-wide binding profiles of NctA using chromatin-immunoprecipitation sequencing (ChIP-seq). Our ChIP-seq analysis revealed that NCT complex binds the promoters of several ergosterol biosynthetic genes, their transcriptional regulators, and the azole efflux pump cdr1B. Taken together, these results suggest that the NCT complex plays a role as a master regulator of drug resistance in A. fumigatus.
Project description:Genomic DNA from five strains, Aspergillus fumigatus Af71, Aspergillus fumigatus Af294, Aspergillus clavatus, Neosartorya fenneliae, and Neosartorya fischeri, were co-hybridized with that of Aspergillus fumigatus Af293 and compared.
Project description:Aspergillus fumigatus transcription factor AtrR is a critical determinant of the azole resistance phenotype of this organism. AtrR positively regulates expression of a range of genes involved in azole resistance including the ATP-binding cassette transporter-encoding locus abcG1 (cdr1B/abcC) as well as the gene that encodes the enzymatic target of azole drugs cyp51A. Homology searches of A. fumigatus AtrR against a range of fungal species identified highly conserved 25 amino acid region in the carboxy-terminus. To determine the contribution of this region to AtrR function, we prepared a mutant from of the atrR that lacked this region (Δ855-879). We compared the response of an isogenic wild-type strain to a strain expressing the Δ855-879 AtrR derivative using RNA-seq, both in the absence and the presence of voriconazole challenge. The resulting data indicate that this 25 amino acid segment of AtrR is required for expression of some but not all AtrR target genes.
Project description:The Negative cofactor 2 (NCT) complex is an evolutionally conserved heterodimeric transcription factor. In Aspergillus fumigatus, the NCT complex consists of two subunits NctA and NctB. Through a genome-wide screening of a transcription factor null mutant strains, we found that loss of the NCT complex leads to a multi-drug resistance phenotype including the azoles (itraconazole, voriconazole and posaconazole) as well as the salvage therapeutic amphotericin B, and terbinafine. To obtain further insight into the molecular mechanisms driving the azole-resistance in the NCT complex null mutants, we analyzed genome-wide gene expression profiles of the nctA and the nctB null mutants using RNA-seq. Our expression profiling revealed that disruption of the genes lead to upregulation of several ergosterol biosynthetic genes, their transcriptional activators, and the azole efflux pump cdr1B. Taken together, these results suggest that the NCT complex plays a role as a master regulator of drug resistance in A. fumigatus.
Project description:In lung diseases caused by the major mould pathogen Aspergillus fumigatus the pulmonary epithelium is destroyed by invasive growth of fungal hyphae, a process thought to require fungal proteases. Here we show that the A. fumigatus pH-responsive transcription factor PacC governs expression of secreted proteases during invasive lung infections and is required for epithelial invasion and pathogenicity. In addition, A. fumigatus M-NM-^TpacC mutants aberrantly remodel the fungal cell wall during infection. This study defines distinct PacC-mediated mechanisms of host damage during pulmonary aspergillosis. ch1: treatment protocol Temporal transcriptional profiling of ATCC46645 strain and isogenic M-NM-^TpacC Aspergillus fumigatus mutant during murine infection
