IκBα degradation and nuclear factor-κB DNA binding are insufficient for interleukin-1β and tumor necrosis factor-α-induced κb-dependent transcription. Requirement for an additional activation pathway

Two closely related IκBα kinases as well as the upstream kinase, NIK, which integrates interleukin-1β (IL-1β)- and tumor necrosis factor (TNF)- α-dependent activation of the transcription factor NF-κB have recently been described. However, in this emerging pathway the role of previously identified components of cytokine-induced NF-κB activation, namely phosphatidylcholine-specific phospholipase C and protein kinase C, remains unclear. We now show that, in A549 human alveolar epithelial cells, the activation of a stably transfected NF-κB-dependent reporter gene by TNF-α and IL-1β is completely blocked by the phosphatidylcholine-specific phospholipase C inhibitor D609 and the protein kinase C inhibitor R031-8220. However, IL-1β-induced IκBα degradation as well as NF-κB nuclear translocation and DNA binding, as determined by Western blot and electro- mobility shift assay, respectively, are not affected by these inhibitors. A similar effect, although less pronounced, is observed with the p38 mitogen- activated protein kinase inhibitor SB 203580. On the basis of these data we propose the existence of a second signaling pathway induced by IL-1β and TNF-!a that is activated in parallel to the cascade leading to IκBα degradation and is specifically required for NF-κB-dependent transcriptional competency.