Sequence‐specific binding by Aspergillus nidulans AflR, a C6 zinc cluster protein regulating mycotoxin biosynthesis

• Published in: Molecular microbiology Vol. 28; no. 6; pp. 1355 - 1365
• Main Authors: Fernandes, Mary, Keller, Nancy P., Adams, Thomas H.
• Format: Journal Article
• Language: English
• Published: Oxford BSL Blackwell Science Ltd 01.06.1998; Blackwell Publishing Ltd
• Subjects: Aflatoxins - genetics; Artificial Gene Fusion; Aspergillus nidulans - genetics; Aspergillus nidulans - metabolism; Base Sequence; Binding Sites; DNA Footprinting; DNA, Fungal - metabolism; DNA-Binding Proteins - genetics; DNA-Binding Proteins - isolation & purification; DNA-Binding Proteins - metabolism; Fungal Proteins; Gene Expression Regulation, Fungal; Genes, Reporter; Glucuronidase - genetics; Glucuronidase - metabolism; Molecular Sequence Data; Multigene Family; Promoter Regions, Genetic; Sterigmatocystin - biosynthesis; Transcription Factors; Transcriptional Activation; Zinc Fingers
• ISSN: 0950-382X; 1365-2958
• DOI: 10.1046/j.1365-2958.1998.00907.x

The Aspergillus nidulans aflR gene is found within a 60 kb gene cluster that includes ≈24 other genes that putatively function in the production of the aflatoxin‐related mycotoxin sterigmatocystin. Previous work showed that AflR is a C6 zinc binuclear cluster protein that is conserved across Aspergillus spp. and functions as a pathway‐specific transcription factor in activating expression of other cluster genes. In this report, we demonstrate that A. nidulans AflR (AnAflR) is a 45 kDa protein that binds to the palindromic sequence 5′‐TCG(N5)CGA‐3′ found in the promoter regions of several aflatoxin and sterigmatocystin cluster genes (stc genes). The in vivo relevance of this AnAflR binding site was assessed by examining the contribution of the three TCG(N5)CGA elements in the 1.1 kb promoter region of stcU using gene fusions with the bacterial uidA gene encoding β‐glucuronidase (GUS). By mutating one, two or all three of the AnAflR‐binding elements and examining GUS activity in wild‐type aflR or ΔaflR A. nidulans strains, we found that stc gene activation required both AnAflR and at least one TCG(N5)CGA AflR binding site.