Over-expression studies in tissue culture almost certainly do not accurately reflect physiological signaling events. Similarly, what controls the balance between the levels of the various heterodimeric complexes in vivo is not known. Studies in Drosophila have elegantly shown that very small differences in nuclear concentrations of these factors, in their affinities for target DNA sites, and in cooperation or competition between Rel proteins and other transcription factors can have profound physiological consequences in organisms. Lastly, in many situations, it is not known how or which of the many genes induced by Rel/NF-kB factors in a given response contribute to that response. The recent development of methods to analyze genome-wide changes in gene expression (e.g., cDNA microarrays), which has already begun to uncover additional Rel/NF-kB-responsive genes, may clarify which Rel/NF-kB target genes are activated in a given response.
As described above, the structures of several Rel/NF-kB dimers on DNA or bound to IkB are known. In all cases, these structures have been derived from molecules that contain almost exclusively residues from the RH domain. As such, these studies provide rather static glimpses of these factors at work. Several molecular and biochemical studies indicate that Rel dimers assume distinct conformations when bound to DNA versus as free or IkB-bound dimers or when bound to different kB sites. Moreover, such studies have also indicated that C-terminal residues influence sequences within the RH domain. Furthermore, there is surprisingly little information about how any of the Rel/NF-kB complexes actually activate transcription when bound to DNA: that is, what are the co-activators or basal factors with which they interact to activate transcription? Therefore, we cannot accurately simulate the dynamic nature of the complex as it releases from IkB, enters the nucleus, binds to DNA, and enhances gene expression.
Although the discovery and characterization of the IkB kinase complex was a monumental step in our understanding of the regulation of this pathway, it raised almost as many questions as it has answered. For example, the following issues remain murky: 1) precisely which proteins are in the IKK complex in all cell types; 2) the exact size of the complex in all cell types; 3) whether the IKK complex has physiologically-relevant substrates other than IkB (almost certainly); 4) how the various NF-kB-activation pathways converge on IKK (for example, what and how many upstream kinases can activate IKK); 5) how is IKK activated by what appears to be induced clustering; 6) how is it that one subunit of this complex (IKKa) controls a specific developmental process, namely keratinocyte differentiation; 7) what other signaling pathways crosstalk via or to IKK; and 8) how do the two catalytic kinases within the IKK complex act on substrate proteins. X-ray crystal structural information on the IKK complex and its components may help answer some of these questions.
The study of v-Rel unequivocally demonstrates that Rel/NF-kB transcription factors can be oncogenic, and one would like to know how the activating mutations in v-Rel have altered its structure as compared to c-Rel. However, v-Rel has accumulated so many activating mutations that it may not be a precise model for the role of these transcription factors in human cancers, where a single mutation (or gene amplification event) has occurred. Thus, it is not known whether the rearrangements, mutations, and amplifications in Rel/NF-kB/IkB genes that have been repeatedly identified in several human cancers and the constitutive NF-kB signaling seen in certain human cancers or induced by oncogenic human viruses (e.g., EBV and HTLV-1) contribute to proliferation, abrogate growth suppression, influence the control of apoptosis, or affect all of these processes.
The involvement of Rel/NF-kB transcription factors in human inflammation and disease certainly establishes them as targets for therapeutics. Indeed, many common synthetic (e.g., aspirin), and traditional (e.g., green tea, curcumin) remedies target, at least in part, the Rel/NF-kB signaling pathway (see INHIBITORS at this site). It is likely that our knowledge of the molecular details of this pathway will enable us to develop more specific and potent inhibitors.
A suggested nomenclature for members of the Rel/NF-kB signal transduction pathway
Among the many publications on this topic, there are inconsistencies in the naming of genes and proteins in the Rel/NF-kB pathway. Although a system of nomenclature for the Rel/NF-kB transcription factors and IkB proteins was established previously (Nabel and Verma, 1993), we use and propose a slightly modified nomenclature. The revised nomenclature reflects the new members of this pathway, common usage over the past several years, and at times my own judgment. In most cases, the choice was quite simple, although the p65 vs. RelA decision continues to be a thorny one; (even though the p65 habit is hard to break, RelA is much more consistent and logical).