Transcriptional regulation of fibronectin gene by phorbol myristate acetate in hepatoma cells: A negative role for NF-kappa B

The transcriptional regulation of the fibronectin (FN) gene in hepatoma cel ls by phorbol myristate acetate (PMA) was investigated. PMA increased the s ynthesis and mRNA levels of FN and its promoter activity in Hep3B hepatoma cells. The PMA-induced activation of FN expression was blocked by a protein kinase C (PKC) inhibitor and did not require a new protein synthesis. Dele tion analysis revealed that the sequence between positions -69 and +136 of the FN gene was responsible for the PMA induction. Two PMA-inducible nuclea r protein complexes were found to bind to a putative NF-kappa B site at -41 and were identified as a p65/p50 heterodimer and a p50/50 homodimer of NF- kappa B family. Mutations in the -41 NF-kappa B site, however, did not bloc k the PMA induction of the FN promoter but rather enhanced it. Overexpressi on of p65 increased the FN promoter activity. While overexpression of p50 a lone did not affect the promoter activity, it decreased the p65-induced act ivation of the FN promoter. Mutations in the -41 NF-kappa B site attenuated the p50-mediated suppression of the p65 transactivation of the FN promoter . Deletion of the sequence between +1 and +136 decreased the basal and PMA- induced activities of the FN promoter. This study shows that PMA induces th e transcription of the FN gene in hepatoma cells via the PKC pathway. The D NA sequence between +1 and +136 is responsible, at least in part, for the P MA-induced activation of the FN gene, while the -41 NF-kappa B binding site plays as a negative regulatory element for it. In addition, this study is the first to show a role for NF-kappa E p65 in the transcriptional activati on of the FN gene. (C) 2000 Wiley-iiss, Inc.