Bcl-2 activates the transcription factor NFκB through the degradation of the cytoplasmic inhibitor IκBα

Nuclear factor kappa B (NF kappa B) is a ubiquitously expressed transcription factor that is regulated by the cytoplasmic inhibitor protein I kappa B alpha. Biological agents such as tumor necrosis factor alpha (TNF alpha), which activate NF kappa B, result in the rapid degradation of I kappa B alpha Adenoviral-mediated gene transfer of Bcl-2 prevents apoptosis of neonatal ventricular myocytes induced by TMF alpha, In view of the growing evidence that NF kappa B may play an important role in regulating apoptosis, we determined whether TNF alpha! and Bcl-2 could modulate the activity of NF kappa B in ventricular myocytes. Stimulation of myocytes with TNFa: resulted in a 2.1-fold increase (p < 0.001) in NF kappa B-dependent gene transcription and nuclear DNA binding Similarly, a 1.9-fold increase (p < 0.0002) ire NF kappa B-dependent gene transcription was observed in myocytes expressing Bcl-2. Nuclear DNA binding activity of NF kappa B was significantly increased in myocytes expressing Bcl-2, with a concomitant reduction in I kappa B alpha protein level. The BcI-2-mediated loss of I kappa B alpha could be prevented by the proteasome inhibitor lactacystin, consistent with the notion that the targeted degradation of I kappa B alpha consequent to overexpression of BcI-2 utilizes the ubiquitin-proteasome pathway. This was further tested in human 293 cells in which the N-terminal region of I kappa B alpha was identified to be an important regulatory site for Bcl-2. Deletion of this region or a serine to alanine substitution mutant at amino acids 32 and 36, which are defective for both phosphorylation and degradation were more resistant than wild type I kappa B alpha to the inhibitory effects of Bcl-2. To our knowledge, this provides the first evidence for the regulation of I kappa B alpha by Eel-2 and suggests a link between Eel-2 and the NF kappa B signaling pathway in the suppression of apoptosis.