The role of Rel/NF-κB transcription factors in B lymphocyte survival
Introduction
The Rel/NF-κB family of transcription factors comprises a group of related proteins that function as homo- or hetero-dimers. The five subunits, c-Rel RelA(p65), RelB, NF-κB1 and NF-κB2, each encoded by a distinct locus, share a conserved amino terminal domain (Rel Homology Domain (RHD)) that encompasses sequences essential for DNA binding, dimerization, and nuclear import [1]. The type I sub-group comprises c-Rel, RelA, and RelB, each of which possesses a carboxyl-terminal transcriptional trans-activation domain in addition to the RHD [1]. The type II sub-group members NF-κB1(p50) and NF-κB2(p52) have only the RHD and both are derived by proteolysis from the amino terminus of larger precursors, p105 (NF-κB1) and p100 (NF-κB2), respectively [1]. Since p50 and p52 lack intrinsic transactivating properties, they act either as transcriptional repressors in the form of homo-dimers or they modulate as hetero-dimers the trans-activating function of type I family members [1].
In most cells, the major proportion of Rel/NF-κB proteins resides in the cytoplasm in a latent state, sequestered by inhibitory proteins, called IκB proteins [2]. All IκB proteins contain ankyrin-like repeats, motifs that mediate protein–protein interaction and in IκB proteins they are essential for association with Rel/NF-κB proteins [2]. IκBα, IκBβ and IκBε act in trans to retain c-Rel and RelA containing dimers in the cytoplasm. Moreover, the Rel/NF-κB precursor proteins, NF-κB1 p105 and NF-κB2 p100 contain ankyrin repeats, which act in a cis-inhibitory fashion to prevent the nuclear localization of these proteins [2].
A broad range of signals, including mitogens, growth factors, cytokines and stress stimuli activate an IκB-specific kinase complex (IKK) that contains two related subunits with kinase activity (IKKα and IKKβ) plus a third, non-catalytic subunit (IKKγ) [3]. IKK-mediated phosphorylation targets IκB proteins for ubiquitination and proteasome-dependent degradation, thereby liberating Rel/NF-κB transcription factors to enter the nucleus and activate gene transcription [3].
The nuclear import of Rel/NF-κB factors can be activated by two distinct signaling pathways (outlined in Fig. 1), termed the ‘classical’ and the ‘alternate’ activation pathways. The classical pathway involves the activation of hetero-dimers, such as NF-κB1/RelA or RelA/c-Rel, that are normally retained in the cytoplasm by IκBα-like inhibitory proteins [4]. Signals from certain receptors, including antigen receptors (TCR or BCR) IL-1-R or TNF-R1, lead to the phosphorylation-dependent activation of the IKK complex, which in turn phosphorylates IκB on two N-terminal serine residues, thereby targeting IκB for degradation [4]. This exposes the previously masked nuclear localization signals in the Rel/NF-κB protein, resulting in its nuclear import. In the nucleus, these dimeric transcription factors recognize and bind to decameric sequences, termed κB motifs, found in the transcriptional regulatory regions of many genes, particularly those encoding proteins involved in immune, inflammatory and acute phase responses [1]. Studies using cells deficient in various IKK proteins indicated that IKKβ and IKKγ, but not IKKα are the essential initiators of the ‘classical activation pathway’ [5]. In the ‘alternate activation pathway,’ certain receptors, such as lymphotoxin-β (LTβ) receptor [6] and BAFF receptor 3 (BAFF-R3) [7], [8] activate IKKα which in turn initiates the proteolytic processing of NF-κB2 p100 by phosphorylating this precursor protein within its carboxyl-terminal half [9]. This promotes nuclear import and transcriptional activation of RelB in the form of p52/RelB dimers. The nature of the protease that processes NF-κB2 p100 is presently not clear.
Amongst the many types of target genes that Rel/NF-κB transcription factors regulate are some that directly control cell survival. Mammals have two distinct apoptosis signaling pathways [10], [11], one that is initiated by ligation of ‘death receptors’ (members of the TNF-R family with an intra-cellular death domain, e.g. Fas, TRAIL receptors) and the other which is regulated by pro- and anti-apoptotic proteins of the Bcl-2 family. It appears that Rel/NF-κB transcription factors can regulate expression of genes that function in either of these two pathways [12]. ‘Bona fide’ Rel/NF-κB target genes include those encoding Bcl-2, its pro-survival homologs, Bcl-x and A1, or FLIP, which is an inhibitor of ‘death receptor’ signaling. The mechanisms by which other putative Rel/NF-κB target genes implicated in cell survival, such as cIAP1, cIAP2, A20, TRAF1, TRAF2, and GADD45β, regulate the apoptosis machinery are unclear. Genetic and biochemical studies have indicated that the ‘classical’ and the ‘alternate’ NF-κB signaling pathways, which are activated by distinct stimuli may control the expression of distinct sets of pro-survival genes [12].
Section snippets
The role of Rel/NF-κB transcription factors in B lymphopoiesis
In the bone marrow, productive IgH gene rearrangement in B cell progenitors allows expression of the so-called pre-B cell receptor, composed of the IgH chain plus the invariant λ5 and Vpre-B chains, which is essential for survival, proliferation and differentiation [10]. Following productive IgL chain gene rearrangement, immature B cells exit the bone marrow, and undergo differentiation and selection steps within the spleen. Ultimately, only ∼5% of B cells produced in the bone marrow are
B cell maturation defects caused by loss of Rel/NF-κB signaling are associated with reduced cell survival
A common theme to emerge from the analysis of the IKK-deficient mice and the mutant mice lacking combinations of two Rel/NF-κB subunits is that the absence of mature B cells was associated with impaired cell survival. The nfkb1−/−nfkb2−/− T1 B cells had an abnormally high rate of spontaneous apoptosis in culture [7]. Similarly, a higher than normal turnover of both sIgMhisIgDlo and sIgMhisIgDhi rela−/−c-rel−/− B cells in vivo was associated with accelerated spontaneous death of these B cell
BAFF receptor signaling via the Rel/NF-κB pathway is essential for B cell survival and maturation
Although a clear link has been established between Rel/NF-κB activation, Bcl-2-like pro-survival gene expression and B cell maturation, only recently has the identity of a key extracellular signal that activates this pathway been identified. Genetic and biochemical experiments have demonstrated that the tumor necrosis factor (TNF) family member, termed B cell activating factor or BAFF, is essential for the development of long-lived, mature B cells [34]. BAFF is expressed as a homo-trimeric type
Can BAFF/BR3 activation of NF-κB2 explain everything about B cell survival?
Although the BAFF/BR3/NF-κB2/Bcl-2 connection clearly defines a key link between surface receptor-mediated activation of transcription factors inducing B cell survival and differentiation, it appears unlikely that this fully accounts for the overall role of Rel/NF-κB signaling in B cell development and selection. For example, whereas the combined absence of NF-κB2 plus NF-κB1 blocks B cell development at the T1 stage [7], [23], the developmental checkpoint at which BAFF initiates p100
Survival of peripheral lymphocytes during antigenic activation
Upon activation by antigens or mitogens, mature B cells undergo growth, division and ultimately differentiation to Ig-secreting plasma cells or memory B cells. The proliferative expansion of activated B cells is limited by apoptosis and apoptosis of plasma cells also plays a critical role in terminating humoral immune responses. In mature, resting B cells, NF-κB1/c-Rel hetero-dimers are the most abundant Rel/NF-κB factors, consistent with the progressive increase in expression of these subunits
Conclusions and perspectives
It has been clearly established that sustained survival of developing B cells and mature, recirculating B cells requires signals from two receptors, the BCR and BR3, that activate two distinct Rel/NF-κB signaling pathways that cooperate to induce expression of several pro-survival members of the Bcl-2 family. Deregulation of this signaling mechanism, for example by over-expression of BAFF [34] or Bcl-2 [43] or loss of the Bcl-2 antagonist Bim [44], [45], can lead to autoimmune disease or
Acknowledgements
We wish to thank all of our past and present colleagues, particularly J. Adams, S. Cory, R. Grumont, R. Gugasyan, M. Grossmann, G. Grigoriadis, D. Nesic, D. Vaux, D. Huang, A. Harris and S. Bath, for their help with our research and for discussions. Work in our laboratories is supported by grants and fellowships from the NHMRC (Canberra), the NIH, the Dr. Josef Steiner Cancer Research Foundation (Bern, Switzerland), the Leukaemia and Lymphoma Society of America, the Cancer Research Institute
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