Two different, adjacent and divergent zinc finger binding sites are necessary for CREA-mediated carbon catabolite repression in the proline gene cluster of Aspergillus nidulans

Abstract

CREA is the negative regulator mediating carbon catabolism repression in Aspergillus nidulans. We have determined all the sites in the DNA region between the prnD and prnB genes of the proline degradation cluster of this organism which are able to bind a fusion protein containing the zinc finger domain of CREA. The consensus sequence derived for CREA binding is 5'-SYGGRG-3', but not all possible sites derived from this consensus do in fact bind. The binding of at least some sequences of the form 5'-SYGGAG-3' is context dependent. Two different and divergent sites, separated by one base pair (5'-GCGGAGACCCCAG-3'), contain the previously sequenced derepressed mutations and are essential for carbon catabolite repression in vivo. We have studied the binding of CREA to the region by DNase I and methylation protection, and by methylation and depurination interference. We propose that this pair of sites is the physiological, cis-acting element responsible for the carbon catabolite repression of prnB transcription.