2 DIFFERENT, ADJACENT AND DIVERGENT ZINC-FINGER BINDING-SITES ARE NECESSARY FOR CREA-MEDIATED CARBON CATABOLITE REPRESSION IN THE PROLINE GENE-CLUSTER OF ASPERGILLUS-NIDULANS

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