Inhibitors of protein kinase C (PKC) prevent activated transcription - role of events downstream of NF-κB DNA binding

In pulmonary A549 cells, the protein kinase C (PKC) inhibitor, Ro 31-8220, and the phosphotidylcholine-specific phospholipase C inhibitor, D609, prevent NF-{kappa}B-dependent transcription, yet NF-{kappa}B DNA binding is unaffected (Bergmann, M., Hart, L., Lindsay, M., Barnes, P. J., and Newton, R. (1998) J. Biol. Chem. 273, 6607–6610). We now show that this effect also occurs in BEAS-2B bronchial epithelial cells as well as with other PKC inhibitors (Goe 6976, GF109203X, and calphostin C) in A549 cells. Similarly, phorbol ester, a diacylglycerol mimetic, activates NF-{kappa}B-dependent transcription and potentiates tumor necrosis factor {alpha} (TNF{alpha})-induced NF-{kappa}B-dependent transcription, yet unlike TNF{alpha}, poorly activates I{kappa}B kinase (IKK) activity, I{kappa}B{alpha} degradation, or NF-{kappa}B DNA binding in both A549 and BEAS-2B cells. As phorbol ester-induced NF-{kappa}B-dependent transcription was relatively insensitive to the proteasome inhibitor, MG-132, PKC may affect NF-{kappa}B-dependent transcription via mechanisms other than the core IKK-I{kappa}B pathway. This is supported by Gal4 one hybrid analysis of p65/RelA transactivation, which was potentiated by TNF{alpha} and phorbol ester and was inhibited by Ro 31-8220 and D609. Additionally, a number of PKC isoforms, particularly the novel isoform PKC{epsilon}, induced p65/RelA transactivation. Phosphorylation of p65/RelA and cAMP-responsive element-binding protein (CREB)-binding protein (CBP) was increased by TNF{alpha} treatment and, in the case of CBP, was prevented by Ro 31-8220 or D609. However, p65/RelA-CBP interactions were unaffected by either compound. As this effect was not limited to NF-{kappa}B, but was a more general feature of inducible gene transcription, we suggest PKC isoforms may provide a point of intervention in diseases such as inflammation, or cancer, where activated gene expression is prominent