Cloning and characterization of a novel zinc finger transcriptional repressor - A direct role of the zinc finger motif in repression

We have identified a novel transcriptional repressor, AEBP2, that binds to a regulatory sequence (termed AE-1) located in the proximal promoter region of the aP2 gene that encodes the adipose fatty acid-binding protein. Seque nce analysis of AEBP2 cDNA revealed that it encodes a protein containing th ree Gli-Kruppel (Cys(2)-His(2))-type zinc fingers. Northern blot analysis r evealed two transcripts (4.5 and 3.5 kilobases) which were ubiquitously exp ressed in every mouse tissue examined. In co-transfection assays, AEBP2 rep ressed transcription from the homologous aP2 promoter containing multiple c opies of the AE-1 sequence. Moreover, a chimeric construct encoding a fusio n AEBP2 protein with the Gal4 DNA-binding domain was able to repress the tr anscriptional activity of a heterologous promoter containing the Gal4-bindi ng sequence. The transcriptional repression function of AEBP2 was completel y abolished when one of the conserved histidine residues and a flanking ser ine residue in the middle zinc finger were replaced with an arginine residu e. The defective transcriptional repression function of the mutant derivati ve was due neither to lack of expression nor to a failure to localize to th e nucleus. Moreover, both the wild-type and mutant derivative of either the histidine-tagged recombinant AEBP2 proteins or the in vitro translated Gal 4-AEBP2 fusion proteins were equally able to bind to the target DNA. These results suggest that a portion of the zinc finger structure may play a dire ct role in transcriptional repression function, but not in DNA binding.