Autonomous membrane IgE signaling prevents IgE-memory formation.

Aberrant production of IgE antibodies can lead to allergic diseases. Normally, IgE(+) B cells rarely differentiate into memory B cells (Bmem) or long-lived plasma cells (LLPCs), as they only transiently participate in the germinal center (GC), but the mechanism behind this remains elusive. We found that membrane IgE (mIgE) autonomously triggered rapid plasma-cell differentiation and apoptosis independently of antigen or cellular context, predominantly through the mutually independent CD19-PI3K-Akt-IRF4 and BLNK-Jnk/p38 pathways, respectively, and we identified the ectodomains of mIgE as being responsible. Accordingly, deregulated GC IgE(+) B cell proliferation and prolonged IgE production with exaggerated anaphylaxis were observed in CD19- and BLNK-deficient mice. Our findings reveal an autonomous mIgE signaling mechanism that normally prevents IgE(+) Bmem and LLPC formation, providing insights into the molecular pathogenesis of allergic diseases.

Commensal Bacteria and the Lung Environment Are Responsible for Th2-Mediated Memory Yielding Natural IgE in MyD88-Deficient Mice.

IgE Abs are a common mediator of allergic responses and are generally produced in type 2 immune responses to allergens. Allergen stimulation of IgE-bound FcεRI on mast cells or basophils induces the production of chemical mediators and cytokines. In addition, IgE binding to FcεRI without allergen promotes the survival or proliferation of these and other cells. Thus, spontaneously produced natural IgE can increase an individual's susceptibility to allergic diseases. Mice deficient in MyD88, a major TLR signaling molecule, have high serum levels of natural IgE, the mechanism for which remains unknown. In this study, we demonstrated that the high serum IgE levels were maintained from weaning by memory B cells (MBCs). IgE from plasma cells and sera from most Myd88-/- mice, but none of the Myd88+/- mice, recognized Streptococcus azizii, a commensal bacterium overrepresented in the lungs of Myd88-/- mice. IgG1+ MBCs from the spleen also recognized S. azizii. The serum IgE levels declined with the administration of antibiotics and were boosted by challenge with S. azizii in Myd88-/- mice, indicating the contribution of S. azizii-specific IgG1+ MBCs to the natural IgE production. Th2 cells were selectively increased in the lungs of Myd88-/- mice and were activated upon addition of S. azizii in the lung cells ex vivo. Finally, lung nonhematopoietic cells, and CSF1 overproduced therefrom, were responsible for natural IgE production in Myd88-/- mice. Thus, some commensal bacteria may prime the Th2 response and natural IgE production in the MyD88-defective lung environment in general.

B cell-intrinsic DNase1L3 is essential for the T cell-independent type II response in mice.

T cell independent type II (TI-II) antigens, such as capsular polysaccharides, have multivalent epitopes, which induce B cell activation, plasma cell differentiation and antibody production by strongly cross-linking B cell receptors. However, the mechanism of B cell activation by TI-II antigens remains unclear. In this study, we demonstrate that DNA endonuclease DNase1L3 (also termed DNase γ) is required for the TI-II response. The production of antigen-specific antibodies was severely diminished in DNase1L3-deficient mice upon immunization with TI-II antigens, but not with T cell dependent (TD) antigens. Bone marrow chimeric mice and B cell transfer experiments revealed that B cell-intrinsic DNase1L3 was required for the TI-II response. DNase1L3-deficient B cells were defective in cell proliferation and plasma cell differentiation in the TI-II response in vivo as well as in vitro, which was not rescued by co-culture with DNase1L3-sufficient B cells in vitro, disproving an involvement of a secretory DNase1L3. In vitro stimulation with TI-II antigen transiently increased expression of DNase1L3 and its translocation into the nucleus. RNA-seq analysis of ex vivo B cells that had responded to TI-II antigen in vivo revealed a marked reduction of Myc-target gene sets in DNase1L3-deficient B cells. Expression of IRF4, a gene that Myc targets, was diminished in the ex vivo DNase1L3-deficient B cells, in which forced expression of IRF4 restored the TI-II response in vivo. These data revealed an unexpected role of DNase1L3 in a missing link between B cell receptor signaling and B cell activation in the TI-II response, giving a valuable clue to molecularly dissect this response.

gp49B-mediated negative regulation of antibody production by memory and marginal zone B cells.

The rapid Ab responses observed after primary and secondary immunizations are mainly derived from marginal zone (MZ) and memory B cells, respectively, but it is largely unknown how these responses are negatively regulated. Several inhibitory receptors have been identified and their roles have been studied, but mainly on follicular B cells and much less so on MZ B, and never on memory B cells. gp49B is an Ig superfamily member that contains two ITIMs in its cytoplasmic tail, and it has been shown to negatively regulate mast cell, macrophage, and NK cell responses. In this study, we demonstrate that gp49B is preferentially expressed on memory and MZ B cells. We show that gp49B(-/-) mice produce more IgM after a primary immunization and more IgM and IgG1 after a secondary immunization than gp49B(+/+) mice in T cell-dependent immune responses. Memory and MZ B cells from gp49B(-/-) mice also produce more Abs upon in vitro stimulation with CD40 than those from gp49B(+/+) mice. The in vitro IgM production by MZ B cells from gp49B(+/+), but not gp49B(-/-), mice is suppressed by interaction with a putative gp49B ligand, the integrin αvß3 heterodimer. In addition, gp49B(-/-) mice exhibited exaggerated IgE production in the memory recall response. These results suggest that plasma cell development from memory and MZ B cells, as well as subsequent Ab production, are suppressed via gp49B. In memory B cells, this suppression also prevents excessive IgE production, thus curtailing allergic diseases.

Protein kinase Cδ is essential for the IgG response against T-cell-independent type 2 antigens and commensal bacteria.

Antigens (Ags) with multivalent and repetitive structure elicit IgG production in a T-cell-independent manner. However, the mechanisms by which such T-cell-independent type-2 (TI-2) Ags induce IgG responses remain obscure. Here, we report that B-cell receptor (BCR) engagement with a TI-2 Ag but not with a T-cell-dependent (TD) Ag was able to induce the transcription of Aicda encoding activation-induced cytidine deaminase (AID) and efficient class switching to IgG3 upon costimulation with IL-1 or IFN-α in mouse B cells. TI-2 Ags strongly induced the phosphorylation of protein kinase C (PKC)δ and PKCδ mediated the Aicda transcription through the induction of BATF, the key transcriptional regulator of Aicda. In PKCδ-deficient mice, production of IgG was intact against TD Ag but abrogated against typical TI-2 Ags as well as commensal bacteria, and experimental disruption of the gut epithelial barrier resulted in fatal bacteremia. Thus, our results have revealed novel molecular requirements for class switching in the TI-2 response and highlighted its importance in homeostatic commensal-specific IgG production.

When the human body faces a potentially harmful microorganism, the immune system responds by finding and destroying the pathogen. This involves the coordination of several different parts of the immune system. B cells are a type of white blood cell that is responsible for producing antibodies: large proteins that bind to specific targets such as pathogens. B cells often need help from other immune cells known as T cells to complete antibody production. However, T cells are not required for B cells to produce antibodies against some bacteria. For example, when certain pathogenic bacteria coated with a carbohydrate called a capsule ­ such as pneumococcus, which causes pneumonia, or salmonella ­ invade our body, B cells recognize a repetitive structure of the capsule using a B-cell antigen receptor. This recognition allows B cells to produce antibodies independently of T cells. It is unclear how B cells produce antibodies in this situation or what proteins are required for this activity. To understand this process, Fukao et al. used genetically modified mice and their B cells to study how they produce antibodies independently of T cells. They found that a protein called PKCδ is critical for B cells to produce antibodies, especially of an executive type called IgG, in the T-cell-independent response. PKCδ became active when B cells were stimulated with the repetitive antigen present on the surface of bacteria like salmonella or pneumococcus. Mice that lack PKCδ were unable to produce IgG independently of T cells, leading to fatal infections when bacteria reached the tissues and blood. Understanding the mechanism behind the T cell-independent B cell response could lead to more effective antibody production, potentially paving the way for new vaccines to prevent fatal diseases caused by pathogenic bacteria.

Metabolic Reprogramming Induces Germinal Center B Cell Differentiation through Bcl6 Locus Remodeling.

The germinal center (GC) reaction is essential for long-lived humoral immunity. However, molecular requirements for the induction of Bcl6, the master regulator for GC B cell differentiation, remain unclear. Through screening for cytokines and other stimuli that regulate Bcl6 expression, we identify IL-4 as the strongest inducer. IL-4 signaling alters the metabolomic profile in activated B cells and induces accumulation of the TCA cycle intermediate α-ketoglutarate (αKG), which is required for activation of the Bcl6 gene locus. Mechanistically, after IL-4 treatment, STAT6 bound to the known enhancers in the Bcl6 locus recruits UTX, a demethylase for the repressive histone mark H3K27me3 that requires αKG as a cofactor. In turn, the H3K27me3 demethylation activates the enhancers and transcription of the Bcl6 gene. We propose that IL-4-mediated metabolic reprogramming in B cells is pivotal for epigenomic activation of Bcl6 expression to promote GC B cell differentiation.