Regulation of clusterin gene expression by transforming growth factor beta

Transforming growth factor beta (TGF beta) induces the expression of a wide variety of genes in many cell types. Our previous studies have shown that TGF beta stimulates both clusterin mRNA and protein levels, and induces its accumulation in the nucleus of CCL64 cells. To further investigate the molecular mechanism of clusterin mRNA induction by TGF beta, we created a 1.3-kilobase rat clusterin promoter/luciferase reporter construct. We demonstrate that TGF beta enhances luciferase activity 2.5-6fold in transient transfection assays of epithelial, endothelial, and fibroblast cell lines. Deletional analysis reveals that an AP-1-binding site (5'-TGAGTCA) in the minimal promoter region is necessary for initiating transactivation by TGF beta. A single T to G base mutation in the AP-1 site (5'-TGAGGCA) abolishes TGF beta-induced clusterin promoter transactivation. In transcription factor decoy experiments, 23-mer oligonucleotides of wild type AP-1 reduce TGF beta induction of clusterin mRNA levels and promoter transactivation, while all oligonucleotide containing the mutated AP-1 site has no effect. Two specific protein kinase C inhibitors, GF109203X and calphostin C, block TGF beta-induced clusterin mRNA levels and promoter transactivation. Together these results indicate that TGF beta regulates clusterin gene expression through an AP-1 site and its cognate transcription factor AP-1, and requires the involvement of protein kinase C.