NF-kB transcription factors regulate the expression of genes that promote immunity, inflammation and cell survival. The p105 precursor protein of NF-kB1 p50 acts as an NF-kB inhibitory protein (IkB), retaining associated NF-kB/Rel subunits inactive in the cytoplasm of cells. The pro-inflammatory cytokine tumor necrosis factor- [alpha] (TNF[alpha]) and bacterial lipopolysacharide (LPS) stimulate p105 proteolysis via IkB kinase (IKK) mediated phoshorylation of p105. Subsequent ubiquitination of p105 precedes its degradation by the proteasome, releasing associated NF-kB/Rel subunits to translocate to the nucleus. In this study, it is demonstrated that p105 binds to IKK via the death domain (DD) motif in its C-terminus. Furthermore, the p105 DD is required for the efficient phosphorylation and proteolysis of p105 induced by IKK or in response to TNF stimulation. These data suggest that the p105 DD is a docking domain that couples p105 to signal-induced proteolysis. The MAP 3-kinase tumor progression locus-2 (TPL-2), which is required for the activation of MEK1/2 MAP 2-kinases in response to LPS in macrophages, associates stoichiometrically with p105. Here, it is demonstrated that TPL-2 stability relies on its high affinity association with p105 through two interaction sites. While the TPL-2 C- terminus binds to residues 497-538 of p105, the p105 DD interacts with the TPL-2 kinase domain. Binding to the p105 DD inhibits TPL-2 MEK kinase activity, but concomitant interaction of the TPL-2 C-terminus with residues 497-538 of p105 is required for efficient TPL-2 inhibition by p105. Consequently, the C-terminally truncated form of TPL-2 is insensitive to p105 regulation in vivo, which may explain why such a mutation is oncogenic. These data indicate that in addition to its role as a precursor for p50 and a cytoplasmic inhibitor of NF-kB, p105 is a negative regulator of TPL-2
