We have demonstrated that p52–deficient animals (a) have reduced numbers of B cells, consistent with a loss of B cell follicles within their spleens and lymph nodes, (b) are unable to form germinal centers and are impaired in antibody responses to TD antigens, (c) lack follicular dendritic cell networks, (d) lack MMs in the splenic MZ and are impaired in exclusion of cells from the white pulp. We have also demonstrated that the inability to generate germinal centers does not track with lymphocytes, as adoptively transferred p52-deficient lymphocytes can form germinal centers in RAG-1–deficient mice. Together, these data reveal a defect in p52 null mutant mice in select aspects of antigen-dependent activation of lymphocytes, and they implicate accessory cells as primary effectors of unique and critical functions for p52 in wild-type animals (especially in follicular dendritic cells; see below).
Many defects observed in p52 null mice are very similar to those observed in Bcl-3–deficient animals (32
). This supports the notion that a ternary complex of DNA-binding p52 homodimers and Bcl-3 (14
) may regulate expression of some critical genes. Nevertheless, these two proteins are likely to have separate functions as well. Mice deficient in p52 not only lack p52 homodimers, they also lack the p100 precursor and any heterodimeric complexes containing p52. If these latter complexes have critical, nonredundant functions, we would expect to see defects independent of Bcl-3. Similarly, Bcl-3 may have critical, nonredundant functions, independent of p52 (see below). As shown here, Bcl-3 and p52 knockouts differ somewhat with respect to resistance to infection with T. gondii
and loss of MZMs and MMs (32
In contrast to mice deficient in Bcl-3 or p52, mice lacking p50 form apparently normal B cell follicles, normal germinal centers, normal follicular dendritic networks, and exhibit no loss of macrophages of the marginal zone (Fig. , and Franzoso, G., L. Poljak, and U. Siebenlist, unpublished observations; reference 33
). Therefore, the data have revealed several specific biologic functions that depend on both Bcl-3 and p52, but not on p50, despite the fact that p50 is quite homologous to p52, and despite the fact that Bcl-3 is known to interact with p50 homodimers (although this interaction may differ from that with p52 homodimers; see Introduction) (10
). Clear biologic contexts in which a complex between p50 homodimers and Bcl-3 is uniquely critical have yet to be described, although mice deficient in p50 or in Bcl-3 have been reported as impaired in their overall ability to contain certain bacterial infections (24
). It remains possible, of course, that p50 homodimers and Bcl-3 have antagonistic activities (see Introduction).
Given the present data, it appears that a complex of p52 and Bcl-3 may be particularly critical for some accessory cell functions during antigen-dependent stimulation of lymphocytes. Adoptively transferred lymphocytes lacking p52 (or lacking Bcl-3) can form germinal centers in RAG-1– deficient mice and generate at least some T-dependent antibody responses, even in the absence of adjuvants. This implies that the primary defects with respect to these biologic processes do not reside within the mature, p52–deficient B or T cell lineages, which implicates accessory cells. In this regard, the apparent loss of follicular dendritic cell networks in both p52– and Bcl-3–deficient mice is particularly noteworthy and could account for the impairment in B cell follicular structure and the lack of germinal center formation (Figs. and ; reference 32
). On the other hand, FDCs may not always be absolutely necessary for germinal center formation (48
). It remains to be shown if defects lie within FDCs themselves or if they lie elsewhere, for example, in some other long-lived non–T, non–B cells or in nonhematopoietic cells. Previous data suggest high expression of p52 in FDCs (49
), supporting the notion that this protein has important functions within FDCs. It is possible that p52 and Bcl-3 are needed for the development/differentiation of these cells, acting directly or indirectly, just like RelB is required for formation of mature interdigitating dendritic cells (IDCs; references 28
); (IDCs are present in both p52 and Bcl-3 knockout mice [Franzoso, G. and U. Siebenlist, unpublished observations].) It is worth noting also that the absence of p52 or Bcl-3 may well have direct effects in more than one cell type. In any case, a correlation between the failure to form FDC networks and impaired antibody response has also been noted in lymphotoxin (LT)α–deficient mice (50
Adjuvant-aided presentation of antigen in p52 knockout mice appears to be able to compensate for the dramatic failure to respond to TD antigens without adjuvants. Although even then, proper germinal centers were still not observed. (Antibody production in the absence of germinal centers has been previously observed in LTα knockout mice .) This indicates a largely normal behavior of mutant lymphocytes once they are stimulated, implying that defects in the mutant mice must lie in other cell types. Strong adjuvant-aided antigenic stimulation may be associated with inflammation and thus production of cytokines in the mutant mice, which may overcome what is otherwise a defective priming of lymphocytes by accessory cells.
Which genes could be the unique targets of p52 and Bcl-3? Such genes must be regulated, at least in part, by nonredundant functions of these proteins. One clue may come from mutant mice that present with partially related phenotypes. For example, mice deficient in TNF-α, TNF receptor I, LTα, or LTβ display disrupted lymphoid microarchitecture including lack of germinal centers and FDC networks (52– 56). An absence of germinal centers has also been noted in mutant mice deficient in various proteins known to be involved in costimulatory signaling during antigen presentation, such as CD40 and CD40 ligand, but these mice have apparently normal lymphoid architecture (47
). All of these null mutant mice do have additional phenotypes that are not shared with p52– and Bcl-3–deficient mice. Still, it is quite likely that specific aspects of the signaling cascades that occur during in vivo antigenic stimulations are impaired in p52– or Bcl-3–deficient mice and/or that the specific in vivo microenvironments are disrupted that are needed to set up proper antigen-stimulated signaling cascades between cells. It remains to be shown if p52 and Bcl-3 lie on some of the same signaling pathways disrupted in the knockouts discussed above. In this regard, it is important to note that NF-κB complexes control, as well as are controlled by various members of the TNF ligand and TNF receptor families (2
), allowing for multiple ways in which these particular molecules could be functionally connected. Identification of molecular targets involved will be essential to understanding how these proteins regulate gene expression and how antigen priming is accomplished in vivo.
Since both p52 and Bcl-3 have been found to be associated with recurrent translocations (9
), it is noteworthy that an absence of either protein leads to losses in B cell numbers, in particular with age and with immune challenges. This suggests that p52 and Bcl-3 normally help to maintain B cells. When overexpressed and/or inappropriately expressed, these properties of p52 and Bcl-3 may contribute to tumor formation.