A Novel cAMP-response Element, CRE1, Modulates Expression of nor-1 in Aspergillus parasiticus
Author:
Roze LV, Miller MJ, Rarick M, Mahanti N, Linz JE
Date: 11 November 2005
Abstract:
The level of aflatoxin accumulation in the filamentousfungus Aspergillus parasiticus is modulated by avariety of environmental cues. The presence of glucose(a preferred carbon source) in liquid and solid glucoseminimal salts (GMS) growth media strongly stimulatedaflatoxin accumulation. Peptone (a non-preferred carbonsource) in peptone minimal salts (PMS) mediastimulated only low levels of aflatoxin accumulation.Glucose stimulated transcription of the aflatoxinstructural genes ver-1 and nor-1 to similar intermediatelevels in liquid GMS, while on solid media, ver-1transcription was stimulated to 20-fold higher levelsthan nor-1. PMS liquid and solid media stimulated verylow or non-detectable levels of transcription of bothgenes. Electrophoretic mobility shift analysis using anor-1 promoter fragment (norR) and A. parasiticus cellprotein extracts revealed specific DNA-protein complexesof different mobility on GMS and PMS solid andliquid media. An imperfect cAMP-response element,CRE1, was identified in norR that mediated formationof the specific DNA-protein complexes. Mutation inCRE1 or AflR1 (AflR cis-acting site) caused up to a3-fold decrease in cAMP-mediated stimulation of nor-1promoter activity on GMS agar. South-Western blotanalysis identified a 32-kDa protein that specificallybound to norR. p32 could be co-immunoprecipitated byanti-AflR antibody and co-purified with an AflR-maltose-binding protein fusion demonstrating a physicalinteraction between AflR and p32 in vitro. We hypothesizethat p32 assists AflR in binding to the nor-1 promoter,thereby modulating nor-1 gene expression inresponse to environmental cues.
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