Modeling and functional analysis of AEBP1, a transcriptional repressor

Abstract

Adipocyte enhancer binding protein 1 (AEBP1) is a transcriptional repressor of the aP2 gene, which encodes the adipocyte lipid binding protein and is involved in the differentiation of preadipocytes into mature adipocytes. It is an isoform of aortic carboxypeptidase-like protein (ACLP), which is a part of the extracellular matrix. AEBP1 and ACLP contain a conserved carboxypeptidase domain which is critical for the function of AEBP1 as a transcriptional repressor. Homology modeling and multiple alignment of AEBP1 homologues were performed to identify putative domains and critical residues that were then deleted or mutated in mouse AEBP1. Expression of wild-type and mutant AEBP1 proteins in CHO cells was performed, and their function in transcriptional repression was assayed by luciferase assay. All deletion forms of AEBP1 were able to repress transcription driven by the aP2 promoter. The DNA binding domain of AEBP1 was mapped by electrophoretic mobility shift assays to a region of the C-terminus rich in basic residues. However, wild-type AEBP1 was not able to interact strongly with DNA, suggesting that AEBP1 might function predominantly as a corepressor, independent of DNA binding. AEBP1 was also found to interact with Ca2+/calmodulin through this basic region, suggesting another mechanism of functional regulation.