Summary The signaling module that specifies nuclear factor-κB (NF-κB) activation is a three-component system: NF-κB, inhibitor of NFκB (IκB), and IκB kinase complex (IKK). IKK receives upstream signals from the surface or inside the cell and converts itself into a catalytically active form leading to the destruction of IκB in the inhibited IκB: NF-κB complex, leaving active NF-κB free to regulate target genes. Hidden within this simple module are family members that all can undergo various modifications resulting in expansion of functional spectrum. Three-dimensional structures representing all three components are now available. These structures have allowed us to interpret cellular observations in molecular terms and at the same time helped us to bring forward new concepts focused towards understanding the specificity in the NF-κB activation pathway.

Conformational change in human IKK2 permits dimers to form higher-order oligomers that support interaction between kinase domains and promote activation through trans auto-phosphorylation.

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The X-ray structure of the nuclear factor-jB (NF-jB) p52:RelB:jB DNA complex reveals a new recognition feature not previously seen in other NF-jB:jB DNA complexes. Arg 125 of RelB is in contact with an additional DNA base pair. Surprisingly, the p52:RelB R125A mutant heterodimer shows defects both in DNA binding and in transcriptional activity only to a subclass of jB sites. We found that the Arg 125-sensitive jB sites contain more contiguous and centrally located A:T base pairs than do the insensitive sites. A protein-induced kink observed in this complex, which used an AT-rich jB site, might allow the DNA contact by Arg 125; such a kink might not be possible in complexes with non-AT-rich jB sites. Furthermore, we show that the p52:RelB heterodimer binds to a broader spectrum of jB sites when compared with the p50:RelA heterodimer. We suggest that the p52:RelB heterodimer is more adaptable to complement sequence and structural variations in jB sites when compared with other NF-jB dimers.

The NF-B family member RelB has many properties not shared by other family members such as restricted subunit association and lack of regulation by the classical IB proteins. We show that the protein level of RelB is significantly reduced in the absence of p100 and reduced even more when both p100 and p105 are absent. RelB stabilizes itself by directly interacting with p100, p105, and their processed products. However, RelB forms complexes with its partners using different interaction modes. Although the C-terminal ankyrin repeat domain of p105 is not involved in the RelB-p105 complex formation, all domains and flexible regions of each protein are engaged in the RelB-p100 complex. In several respects the RelB-p52 and RelB-p100 complexes are unique in the NF-B family. The N-terminal domain of p100/p52 interacts with RelB but not RelA. The transcriptional activation domain of RelB, but not RelA, directly interacts with the processing region of p100. These unique protein-protein contacts explain why RelB prefers p52 as its dimeric partner for transcriptional activity and is retained in the cytoplasm as an inhibited complex by p100. This association-mediated stabilization of RelB implies a possible role for RelB in the processing of p100 into p52.The dimeric NF-B transcription factors are formed from five family members, p50 (NF-B1), RelA (p65), p52 (NF-B2), c-Rel, and RelB. These proteins share an ϳ300-residue long homologous region located near the N terminus. This element, referred to as the Rel homology region (RHR), 5 is responsible for DNA binding, dimerization, inhibitor binding, and nuclear localization. p50 and p52 are the processed products of precursor proteins, p105 and p100, respectively (1, 2). RelA and c-Rel homo-and heterodimers are tightly regulated by a class of inhibitor proteins known as IB through the formation of stable IB-NF-B complexes that are unable to bind DNA. Activation of these dimers requires degradation of IB. A large number of stimuli activate IB degradation through phosphorylation of IB by IB kinase (IKK) leading to ubiquitination, 26 S proteasome recruitment, and degradation of IB by the proteasome (3, 4). Signaling pathways leading to NF-B activation through degradation of classical IB proteins (IB␣, IB␤, and IB⑀) are classified as the canonical pathways.RelB displays characteristics that are not shared by the other NF-B subunits: 1) Prototypical IB proteins do not regulate RelB-containing NF-B dimers (5, 6). 2) The RelB homodimer does not have DNA binding activity, suggesting that unlike other members, RelB may not form a stable detectable homodimer in vivo (7). 3) RelB has a N-terminal extension, known as the leucine zipper (LZ) domain because of the presence of a leucine-rich heptad repeat that is not present in other NF-B family members (8). 4) The x-ray crystal structure of the RelB dimerization domain (DD) revealed an intertwined domain swapped arrangement of the two monomers, suggesting that this RelB domain fold might be unstable and is stabilized by domain intertwining (9...