Transcriptional regulation of the Antifungal Protein in Aspergillus giganteus

Abstract

Regulation of the expression of the afp gene that codes for the Antifungal Protein of Aspergillus giganteus was investigated using a reporter system. For this purpose, the E. coli reporter gene uidA encoding beta-glucuronidase (GUS) was placed under the control of the afp promoter. No homologous integration of the reporter construct into the afp site was observed among 156 transformants tested. In one of the transformants carrying a single, ectopically integrated, copy of the construct, GUS and AFP both displayed exactly the same temporal expression patterns under various cultivation conditions, as assayed by Northern and protein analyses. Thus, this transformant was used to identify factors that are involved in the transcriptional regulation of afp expression. Expression is only detectable in the vegetative mycelium, whereas no expression occurs in aerial hyphae or conidia, indicating that afp expression is developmentally regulated. Transcription of afp is regulated by ambient pH, being suppressed under acidic conditions and strongly induced under alkaline conditions. This observation suggests that PacC regulates the afp gene, which is consistent with the presence of two putative PacC binding sites within the 5' upstream region. Transcription is not subject to carbon catabolite repression or nitrogen metabolite repression. The expression of afp is up-regulated by heat shock, upon growth in the presence of excess NaCl and ethanol, and under conditions of carbon starvation. In contrast, expression decreases slightly in the presence of hydrogen peroxide and under nitrogen starvation. These data are compatible with the presence of a putative heat shock element (NTTCNNGANTTCN) and five putative C(4)T stress-responsive elements within the afp promoter.

Updating genome annotation for the microbial cell factoryAspergillus nigerusing gene co-expression networks

“…Ribosomally synthesized antifungal peptides of the AFP family are promising molecules for use in medical or agricultural applications to combat humanand plant-pathogenic fungi (40). We and others could show that expression of their cognate genes are under tight temporal and spatial regulation in their native hosts and precedes asexual sporulation (24,(41)(42)(43).…”

Antifungal proteins: More than antimicrobials?

“…The P. chrysogenum paf and the A. giganteus afp exhibit a temporal and spatial regulation during asexual differentiation (Hegedus et al. , 2011a; Meyer et al. , 2002).…”

“…No gene expression occurs in conidiophores or conidia (Hegedus et al. , 2011a; Meyer, 2008; Meyer et al. , 2002).…”

Genetic manipulation of fungal strains for the improvement of heterologous genes expression (a mini-review)

“…Upstream regulatory regions of gluc78 (which encodes GLUC78, a 78-kDa exo-1,3-glucosidase) (Donzelli et al, 2001), ech42 and nag1 (which encodes CHIT73, a 73-kDa N-acetylhexosaminidase) (Peterbauer et al, 1996) from T. atroviride contain clustered putative binding sites, such as GATA, stress response elements, BrlA response elements and AbaA response elements, suggesting that they are regulated by a number of stimuli. A study showed that complex response elements existing in the promoter region of the Antarctosaurus giganteus AFP gene (encoded a antifungal protein) are responsible for its efficient transcription in its expression vectors (Meyer et al, 2002).…”