bcl‐2 expression is often associated with poor prognosis in several types of tumors; however, the role of this molecule in breast cancer is still controversial. We found earlier that over‐expression of bcl‐2 in a human breast‐cancer cell line (MCF7^ADR) enhances its tumorigenicity and metastatic potential by inducing metastasis‐associated properties such as increased secretion of the matrix metalloproteinase‐9 (mmp‐9). In the present study, we investigated the effect of bcl‐2 over‐expression on the activity of the transcription factor NF‐κB, an important regulator of genes involved in tumor progression and invasion. Transient transfection experiments indicate that over‐expression of bcl‐2 in the MCF7^ADR cell line, enhances NF‐κB‐dependent transcriptional activity. Mobility‐shift analysis revealed an increase of NF‐κB DNA‐binding in bcl‐2‐over‐expressing clones that correlated with lower levels of the NF‐κB cytoplasmic inhibitor IκBα. Moreover, point mutations of 2 highly conserved residues within the BH1 and BH2 domains that abrogate the interaction of bcl‐2 with bax, or deletion of the N‐terminal BH4 domain, completely eliminate the ability of this molecule to up‐regulate NF‐κB‐dependent transactivation. Since mmp‐9 is a NF‐κB‐regulated gene, we also investigated whether bcl‐2 over‐expression up‐regulated mmp‐9 transcription. We found that induction of mmp‐9 mRNA correlates with the activation of an mmp‐9‐promoter‐reporter‐gene construct in transient transfection assay, and a mutation of the (−600)mmp‐9‐NF‐κB binding element abolishes this effect. The overall data indicate that bcl‐2‐mediated regulation of NF‐κB‐transcription‐factor activity may represent an important mechanism for the promotion of malignant behavior in MCF‐7^ADR cells. Int. J. Cancer 86:188–196, 2000. © 2000 Wiley‐Liss, Inc.