Regulation of nuclear factor κB transactivation -: Implication of phosphatidylinositol 3-kinase and protein kinase C ξ IN c-Rel activation by tumor necrosis factor α

Transactivation by c-Rel (nuclear factor kappaB) was dependent on phosphorylation of several serines in the transactivation domain, indicating that it is a phosphorylation-dependent Ser-rich domain. By Ser --> Ala mutational and deletion analysis, we have identified two regions in this domain: 1) a C-terminal region (amino acids 540-588), which is required for basal activity; and 2) the 422-540 region, which responds to external stimuli as tumor necrosis factor (TNF) alpha or phorbol myristate acetate plus ionomycin. Ser from 454 to 473 were shown to be required for TNF alpha -induced activation, whereas Ser between 492 and 519 were required for phorbol myristate acetate plus ionomycin activation. Phosphatidylinositol S-kinase (PI3K) and protein kinase C (PKC) zeta were identified as downstream signaling molecules of TNF alpha -activation of c-Rel transactivating activity. Interestingly, dominant negative forms of PI3K inhibited PKC zeta activation and dominant negative PKC zeta inhibited PI3K-mediated activation of c-Rel transactivating activity, indicating a cross-talk between both enzymes. We have identified the critical role of different Ser for PKC zeta- and PI3K-mediated responses. Interestingly, those c-Rel mutants not only did not respond to TNF alpha but also acted as dominant negative forms of nuclear factor kappaB activation.