Defective immune responses in mice lacking LUBAC-mediated linear ubiquitination in B cells.

The linear ubiquitin chain assembly complex (LUBAC) plays a crucial role in activating the canonical NF-κB pathway, which is important for B-cell development and function. Here, we describe a mouse model (B-HOIP(Δlinear)) in which the linear polyubiquitination activity of LUBAC is specifically ablated in B cells. Canonical NF-κB and ERK activation, mediated by the tumour necrosis factor (TNF) receptor superfamily receptors CD40 and TACI, was impaired in B cells from B-HOIP(Δlinear) mice due to defective activation of the IKK complex; however, B-cell receptor (BCR)-mediated activation of the NF-κB and ERK pathways was unaffected. B-HOIP(Δlinear) mice show impaired B1-cell development and defective antibody responses to thymus-dependent and thymus-independent II antigens. Taken together, these data suggest that LUBAC-mediated linear polyubiquitination is essential for B-cell development and activation, possibly via canonical NF-κB and ERK activation induced by the TNF receptor superfamily, but not by the BCR.

Preferential localization of IgG memory B cells adjacent to contracted germinal centers.

It has long been presumed that after leaving the germinal centers (GCs), memory B cells colonize the marginal zone or join the recirculating pool. Here we demonstrate the preferential localization of nitrophenol-chicken gamma-globulin-induced CD38(+)IgG1(+) memory B cells adjacent to contracted GCs in the spleen. The memory B cells in this region proliferated after secondary immunization, a response that was abolished by depletion of CD4(+) T cells. We also found that these IgG1(+) memory B cells could present antigen on their surface, and that this activity was required for their activation. These results implicate this peri-GC region as an important site for survival and reactivation of memory B cells.

Augmented antibody response with premature germinal center regression in CD40L transgenic mice.

Although CD40 signaling is required for activation and differentiation of B cells, including germinal center (GC) formation and generation of memory B cells, in vivo generation of CD40 signaling augments plasma cell differentiation but disrupts GCs. Thus, CD40 signaling is thought to direct B cells to extrafollicular plasma cell fate rather than GC formation. In this study, we analyzed CD40L transgenic (CD40LTg) mice that constitutively express CD40L on B cells. After immunization, activation of B cells, but not dendritic cells, was augmented, although dendritic cells can be activated by CD40 ligation. Bone marrow chimera carrying CD40LTg and nontransgenic B cells showed increased Ab production from transgenic, but not from coexisting nontransgenic, B cells, suggesting that CD40L on a B cell preferentially stimulates the same B cell through an autocrine pathway, thereby augmenting Ab production. Although GCs rapidly regressed after day 5 of immunization and failed to generate late-appearing high-affinity Ab, CD40LTg mice showed normal GC formation up to day 5, as well as normal generation of long-lived plasma cells and memory B cell responses. This observation suggests that CD40 signaling does not block GC formation or differentiation of GC B cells, but it inhibits sustained expansion of GC B cells and augments B cell differentiation.

Regulation of B-cell development by BCAP and CD19 through their binding to phosphoinositide 3-kinase.

Despite the importance of phosphoinositide 3-kinase (PI3K) in B-cell development, its activation mechanism still remains elusive. In this study, we show that deletion of both BCAP and CD19 leads to an almost complete block of BCR-mediated Akt activation and to severe defects in generation of immature and mature B cells. The YXXM motifs in BCAP and CD19 are crucial for regulating B-cell development in that mutation of these motifs abrogated their ability to induce BCR-mediated Akt activation as well as to promote B-cell development. Furthermore, the developmental defect in CD19(-/-)BCAP(-/-) B cells was partly relieved by introducing a constitutively active form of PI3K or PDK1. Together, our data suggest that BCAP and CD19 have complementary roles in BCR-mediated PI3K activation, thereby, at least in part, contributing to B-cell development.

Extracellular signal-regulated protein kinase 2 is required for efficient generation of B cells bearing antigen-specific immunoglobulin G.

Activation of extracellular signal-regulated protein kinase (ERK) has been implicated in proliferation as well as differentiation in a wide variety of cell types. Using B-cell-specific gene-targeted mice, we report here that in T-cell-dependent immune responses, ERK2 is required to generate efficient immunoglobulin G (IgG) production. In its absence, the proportion of antigen-specific surface IgG1-bearing cells and the subsequent number of IgG1 antibody-secreting cells were decreased, despite apparently unimpaired class switch recombination. Notably, this defect was countered by overexpression of the antiapoptotic factor Bcl-2. Together, our results suggest that ERK2 plays a key role in efficient generation of antigen-specific IgG-bearing B cells by promoting their survival.

BANK negatively regulates Akt activation and subsequent B cell responses.

BANK is an adaptor protein that is highly expressed in B cells. To investigate its physiological role, we generated BANK-deficient mice. BANK-deficient mice displayed enhanced germinal center formation and IgM production in response to T-dependent antigens, whereas this phenotype was blocked in CD40-BANK double knockout mice. Involvement of BANK in CD40 signaling was further demonstrated by in vitro analysis. CD40-mediated proliferation and survival were significantly increased in BANK-deficient B cells, with enhanced Akt activation, whereas introduction of dominant-negative Akt into BANK-deficient B cells suppressed the augmented CD40-mediated responses. Together, our findings suggest that BANK attenuates CD40-mediated Akt activation, thereby preventing hyperactive B cell responses.

Ectopic CD40 ligand expression on B cells triggers intestinal inflammation.

Several studies indicate that CD4(+) T cells, macrophages, and dendritic cells initially mediate intestinal inflammation in murine models of human inflammatory bowel disease. However, the initial role of B cells in the development of intestinal inflammation remains unclear. In this study we present evidence that B cells can trigger intestinal inflammation using transgenic (Tg) mice expressing CD40 ligand (CD40L) ectopically on B cells (CD40L/B Tg). We demonstrated that CD40L/B Tg mice spontaneously developed severe transmural intestinal inflammation in both colon and ileum at 8-15 wk of age. In contrast, CD40L/B TgxCD40(-/-) double-mutant mice did not develop colitis, indicating the direct involvement of CD40-CD40L interaction in the development of intestinal inflammation. The inflammatory infiltrates consisted predominantly of massive aggregated, IgM-positive B cells. These mice were also characterized by the presence of anti-colon autoantibodies and elevated IFN-gamma production. Furthermore, although mice transferred with CD4(+) T cells alone or with both CD4(+) T and B220(+) B cells, but not B220(+) cells alone, from diseased CD40L/B Tg mice, develop colitis, mice transferred with B220(+) B cells from diseased CD40L/B Tg mice and CD4(+) T cells from wild-type mice also develop colitis, indicating that the Tg B cells should be a trigger for this colitis model, whereas T cells are involved as effectors. As it has been demonstrated that CD40L is ectopically expressed on B cells in some autoimmune diseases, the present study suggests the possible contribution of B cells in triggering intestinal inflammation in human inflammatory bowel disease.

Cutting Edge: Ectopic expression of CD40 ligand on B cells induces lupus-like autoimmune disease.

CD40 ligand (CD40L) is ectopically expressed on B cells in patients with systemic lupus erythematosus (SLE) and lupus-prone BXSB mice. To assess the role of the ectopic CD40L expression in development of SLE, we have established transgenic mice expressing CD40L on B cells. Some of the 12- to 14-mo-old CD40L-transgenic mice spontaneously produced autoantibodies such as antinuclear Abs, anti-DNA Abs, and antihistone Abs. Moreover, approximately half of the transgenic mice developed glomerulonephritis with immune-complex deposition, whereas the kidneys of the normal littermates showed either no pathological findings or only mild histological changes. These results indicate that CD40L on B cells causes lupus-like disease in the presence of yet unknown environmental factors that by themselves do not induce the disease. Thus, ectopic CD40L expression on B cells may play a crucial role in development of SLE.