Mcl-1 is essential for the survival of plasma cells.

The long-term survival of plasma cells is entirely dependent on signals derived from their environment. These extrinsic factors presumably induce and sustain the expression of antiapoptotic proteins of the Bcl-2 family. It is uncertain whether there is specificity among Bcl-2 family members in the survival of plasma cells and whether their expression is linked to specific extrinsic factors. We found here that deletion of the gene encoding the antiapoptotic protein Mcl-1 in plasma cells resulted in rapid depletion of this population in vivo. Furthermore, we found that the receptor BCMA was needed to establish high expression of Mcl-1 in bone marrow but not spleen plasma cells and that establishing this survival pathway preceded the component of plasma cell differentiation that depends on the transcriptional repressor Blimp-1. Our results identify a critical role for Mcl-1 in the maintenance of plasma cells.

Cutaneous Exposure to Clinically Relevant Lone Star Ticks Promotes IgE Production and Hypersensitivity through CD4+ T Cell- and MyD88-Dependent Pathways in Mice.

Tick-borne allergies are a growing public health concern and have been associated with the induction of IgE-mediated food allergy to red meat. However, despite the increasing prevalence of tick bite-induced allergies, the mechanisms by which cutaneous exposure to ticks leads to sensitization and the production of IgE Abs are poorly understood. To address this question, an in vivo approach was used to characterize the IgE response to lone star tick proteins administered through the skin of mice. The results demonstrated that tick sensitization and challenge induced a robust production of IgE Abs and supported a role for IgE-mediated hypersensitivity reactions in sensitized animals following oral administration of meat. The induction of IgE responses was dependent on cognate CD4+ T cell help during both the sensitization phase and challenge phase with cutaneous tick exposure. In addition, IgE production was dependent on B cell-intrinsic MyD88 expression, suggesting an important role for TLR signaling in B cells to induce IgE responses to tick proteins. This model of tick-induced IgE responses could be used to study the factors within tick bites that cause allergies and to investigate how sensitization to food Ags occurs through the skin that leads to IgE production.

An integrated framework using high-dimensional mass cytometry and fluorescent flow cytometry identifies discrete B cell subsets in patients with red meat allergy.

BACKGROUND: B cells play a critical role in the development and maintenance of food allergy by producing allergen-specific IgE. Despite the importance of B cells in IgE-mediated food allergy, the identity of sIgE-producing human B cells and how IgE is regulated are poorly understood. OBJECTIVE: To identify the immunophenotypes of circulating B cells associated with the production of galactose-alpha-1,3-galactose-specific IgE production in patients with red meat allergy. METHODS: B cells in PBMC samples obtained from 19 adults with physician-diagnosed red meat allergy and 20 non-meat allergic healthy controls were assessed by mass cytometry along with a bioinformatics analysis pipeline to identify discrete B cell phenotypes that associated with serum sIgE. Fluorescent flow cytometry was then applied to sort purify discrete B cell subsets, and B cells were functionally evaluated on an individual cell level for the production of sIgE by ELISPOT. RESULTS: Discrete B cell phenotypes abundant in meat allergic subjects compared to non-meat allergic controls were found in peripheral blood that do not share typical characteristics of classical isotype-switched memory B cells that express high levels of CD27. These B cell subsets shared higher IgD and lower IgM expression levels coupled with CXCR4, CCR6 and CD25 expression. In vitro polyclonal stimulation of purified B cell subsets from meat allergic subjects demonstrated that these subsets were enriched for cells induced to secrete sIgE. CONCLUSIONS AND CLINICAL RELEVANCE: Circulating B cells display increased abundance of discrete B cell subsets in meat allergic subjects. This observation, coupled with the capacity of individual B cell subsets to produce sIgE following activation, implicates these novel B cell phenotypes in promoting IgE in meat allergy.

Vancomycin Treatment Alters Humoral Immunity and Intestinal Microbiota in an Aged Mouse Model of Clostridium difficile Infection.

BACKGROUND: The elderly host is highly susceptible to severe disease and treatment failure in Clostridium difficile infection (CDI). We investigated how treatment with vancomycin in the aged host influences systemic and intestinal humoral responses and select intestinal microbiota. METHODS: Young (age, 2 months) and aged (age, 18 months) C57BL/6 mice were infected with VPI 10463 after exposure to broad-spectrum antibiotics. Vancomycin was given 24 hours after infection, and treatment was continued for 5 days. At select time points, specimens of serum and intestinal tissue and contents were collected for histopathologic analysis, to measure antibody levels and the pathogen burden, and to determine the presence and levels of select intestinal microbiota and C. difficile toxin. RESULTS: Levels of disease severity, relapse, and mortality were increased, and recovery from infection was slower in aged mice compared to young mice. Serum levels of immunoglobulin M, immunoglobulin A, and immunoglobulin G against C. difficile toxin A were depressed in aged mice, and vancomycin treatment reduced antibody responses in both age groups. While baseline levels of total bacterial load, Bacteroidetes, Firmicutes, and Enterobacteriaceae were mostly similar, aged mice had a significant change in the Firmicutes to Bacteroidetes ratio with vancomycin treatment. CONCLUSIONS: Vancomycin treatment decreases the systemic humoral response to CDI. Increased mortality from and recurrence of CDI in the aged host are associated with an impaired humoral response and a greater susceptibility to vancomycin-induced alteration of intestinal microbiota.

Delayed anaphylaxis to alpha-gal, an oligosaccharide in mammalian meat.

IgE-mediated hypersensitivity refers to immune reactions that can be rapidly progressing and, in the case of anaphylaxis, are occasionally fatal. To that end, identification of the associated allergen is important for facilitating both education and allergen avoidance that are essential to long-term risk reduction. As the number of known exposures associated with anaphylaxis is limited, discovery of novel causative agents is crucial to evaluation and management of patients with idiopathic anaphylaxis. Within the last 10 years several apparently separate observations were recognized to be related, all of which resulted from the development of antibodies to a carbohydrate moiety on proteins. Interestingly, the exposure differed from airborne allergens but was nevertheless capable of producing anaphylactic and hypersensitivity reactions. Our recent work has identified these responses as being due to a novel IgE antibody directed against a mammalian oligosaccharide epitope, galactose-alpha-1,3-galactose ("alpha-gal"). This review will present the historical summary of the identification of cetuximab hypersensitivity due to alpha-gal IgE and discuss the non-primate mammalian meat food allergy as well as current goals and directions of our research programs.

Distinct regulation of murine lupus susceptibility genes by the IRF5/Blimp-1 axis.

Genome-wide association studies have identified lupus susceptibility genes such as IRF5 and PRDM1 (encoding for IFN regulatory factor 5 [IRF]5 and Blimp-1) in the human genome. Accordingly, the murine Irf5 and Prdm1 genes have been shown to play a role in lupus susceptibility. However, it remains unclear how IRF5 and Blimp-1 (a transcriptional target of IRF5) contribute to lupus susceptibility. Given that the murine lupus susceptibility locus Nba2 includes the IFN-regulated genes Ifi202 (encoding for the p202 protein), Aim2 (encoding for the Aim2 protein), and Fcgr2b (encoding for the FcγRIIB receptor), we investigated whether the IRF5/Blimp-1 axis could regulate the expression of these genes. We found that an Irf5 deficiency in mice decreased the expression of Blimp-1 and reduced the expression of the Ifi202. However, the deficiency increased the expression of Aim2 and Fcgr2b. Correspondingly, increased expression of IRF5 in cells increased levels of Blimp-1 and p202 protein. Moreover, Blimp-1 expression increased the expression of Ifi202, whereas it reduced the expression of Aim2. Interestingly, an Aim2 deficiency in female mice increased the expression of IRF5. Similarly, the Fcgr2b-deficient mice expressed increased levels of IRF5. Moreover, increased expression of IRF5 and Blimp-1 in lupus-prone C57BL/6.Nba2, New Zealand Black, and C57BL/6.Sle123 female mice (as compared with age-matched C57BL/6 female mice) was associated with increased levels of the p202 protein. Taken together, our observations demonstrate that the IRF5/Blimp-1 axis differentially regulates the expression of Nba2 lupus susceptibility genes, and they suggest an important role for the IRF5/Blimp-1/p202 axis in murine lupus susceptibility.

Loss of TET2 increases B-1 cell number and IgM production while limiting CDR3 diversity.

Recent studies have demonstrated a role for Ten-Eleven Translocation-2 (TET2), an epigenetic modulator, in regulating germinal center formation and plasma cell differentiation in B-2 cells, yet the role of TET2 in regulating B-1 cells is largely unknown. Here, B-1 cell subset numbers, IgM production, and gene expression were analyzed in mice with global knockout of TET2 compared to wildtype (WT) controls. Results revealed that TET2-KO mice had elevated numbers of B-1a and B-1b cells in their primary niche, the peritoneal cavity, as well as in the bone marrow (B-1a) and spleen (B-1b). Consistent with this finding, circulating IgM, but not IgG, was elevated in TET2-KO mice compared to WT. Analysis of bulk RNASeq of sort purified peritoneal B-1a and B-1b cells revealed reduced expression of heavy and light chain immunoglobulin genes, predominantly in B-1a cells from TET2-KO mice compared to WT controls. As expected, the expression of IgM transcripts was the most abundant isotype in B-1 cells. Yet, only in B-1a cells there was a significant increase in the proportion of IgM transcripts in TET2-KO mice compared to WT. Analysis of the CDR3 of the BCR revealed an increased abundance of replicated CDR3 sequences in B-1 cells from TET2-KO mice, which was more clearly pronounced in B-1a compared to B-1b cells. V-D-J usage and circos plot analysis of V-J combinations showed enhanced usage of VH11 and VH12 pairings. Taken together, our study is the first to demonstrate that global loss of TET2 increases B-1 cell number and IgM production and reduces CDR3 diversity, which could impact many biological processes and disease states that are regulated by IgM.

Regulatory roles of the tumor necrosis factor receptor BCMA.

B cell maturation antigen (BCMA) is a tumor necrosis family receptor (TNFR) member that is predominantly expressed on terminally differentiated B cells and, upon binding to its ligands B cell activator of the TNF family (BAFF) and a proliferation inducing ligand (APRIL), delivers pro-survival cell signals. Thus, BCMA is mostly known for its functional activity in mediating the survival of plasma cells that maintain long-term humoral immunity. The expression of BCMA has also been linked to a number of cancers, autoimmune disorders, and infectious diseases that suggest additional roles for BCMA activity. Despite recent advances in our understanding of the roles for the related TNFR members BAFF-R and transmembrane activator and calcium-modulator and cyclophilin ligand interactor (TACI), the signaling pathway used by BCMA for mediating plasma cell survival as well as its putative function in certain disease states are not well understood. By examining the expression, regulation, and signaling targets of BCMA, we may gain further insight into this receptor and how it operates within cells in both health and disease. This information is important for identifying new therapeutic targets that may be relevant in treating diseases that involve the BAFF/APRIL cytokine network.

B cell maturation antigen deficiency exacerbates lymphoproliferation and autoimmunity in murine lupus.

Systemic lupus erythematosus and its preclinical lupus-prone mouse models are autoimmune disorders involving the production of pathogenic autoantibodies. Genetic predisposition to systemic lupus erythematosus results in B cell hyperactivity, survival of self-reactive B cells, and differentiation to autoantibody-secreting plasma cells (PCs). These corrupt B cell responses are, in part, controlled by excess levels of the cytokine BAFF that normally maintains B cell homeostasis and self-tolerance through limited production. B cell maturation Ag (BCMA) is a receptor for BAFF that, under nonautoimmune conditions, is important for sustaining enduring Ab protection by mediating survival of long-lived PCs but is not required for B cell maturation and homeostasis. Through analysis of two different lupus-prone mouse models deficient in BCMA, we identify BCMA as an important factor in regulating peripheral B cell expansion, differentiation, and survival. We demonstrate that a BCMA deficiency combined with the lpr mutation or the murine lupus susceptibility locus Nba2 causes dramatic B cell and PC lymphoproliferation, accelerated autoantibody production, and early lethality. This study unexpectedly reveals that BCMA works to control B cell homeostasis and self-tolerance in systemic autoimmunity.

Aim2 deficiency in mice suppresses the expression of the inhibitory Fcgamma receptor (FcgammaRIIB) through the induction of the IFN-inducible p202, a lupus susceptibility protein.

Murine Aim2 and Ifi202 genes (encoding for the Aim2 and p202 proteins) are members of the IFN-inducible Ifi200 gene family. The Aim2 deficiency in mice activates IFN signaling and stimulates the expression of the lupus susceptibility gene, the Ifi202, located within the NZB autoimmunity 2 (Nba2) interval. Given that the deficiency in the expression of the Fcgr2b gene (encoding for the inhibitory FcγRIIB receptor) is associated with increased lupus susceptibility in mice, we investigated whether the Aim2 protein could regulate the expression of Fcgr2b gene. In this article, we report that Aim2 deficiency in mice suppresses the expression of the FcγRIIB receptor. Interestingly, the Fcgr2b-deficient cells expressed increased levels of the IFN-ß, activated IFN signaling, and expressed reduced levels of the Aim2 protein. Treatment of splenic cells with IFN-α or -γ reduced levels of the FcγRIIB mRNA and protein and also decreased the activity of the FcγRIIB p(-729/+585) Luc reporter. Moreover, levels of the FcγRIIB receptor were significantly higher in the Stat1-deficient splenic cells than in the wild-type cells. Accordingly, increased expression of IFN-ß in lupus-prone B6.Nba2-ABC mice, as compared with non-lupus-prone C57BL/6 (B6) or B6.Nba2-C mice, was associated with reduced expression of the FcγRIIB receptor. Notably, overexpression of the p202 protein in cells decreased the expression of the Aim2 gene, activated the IFN response, and suppressed the expression of the Fcgr2b gene. These observations demonstrate that the expression of Aim2 protein is required to maintain the expression of the Fcgr2b gene and also predict epistatic interactions between the Ifi200 genes and the Fcgr2b gene within the Nba2 interval.

Optimized protocol for the isolation of spleen-resident murine neutrophils.

Neutrophils are an important cellular component of the innate immune system that provides immediate protection to the host from infection. Neutrophil infiltration into inflamed peripheral tissues during infection is beneficial for immunity through phagocytosis of microbes, the release of antimicrobial factors, and secretion of proinflammatory cytokines. Recent reports further suggest that spleen-infiltrating neutrophils play a role in the adaptive immune response by providing survival signals to B cells. However, neutrophils may have detrimental effects on immunity in inflammatory diseases where their recruitment to lymphoid tissues and activation occur abnormally. To determine the contribution of neutrophils that reside in secondary lymphoid tissues to adaptive immunity, direct evaluation of the functional properties of tissue-resident neutrophils is required. We have developed a modified magnetic bead isolation approach for purifying neutrophils from inflamed spleens of autoimmune-prone mice by negative selection. Using this approach, we yielded neutrophils with greater than 90% purity without compromising cell viability. Equally important, the isolation procedure had little effect on the activation of neutrophils and did not impair phagocytic function. Thus, isolation of spleen-resident neutrophils by this optimized approach could be useful for interrogating the functional role of murine neutrophils in normal and abnormal immune responses.

Aim2 deficiency stimulates the expression of IFN-inducible Ifi202, a lupus susceptibility murine gene within the Nba2 autoimmune susceptibility locus.

Murine Aim2 and p202 proteins (encoded by the Aim2 and Ifi202 genes) are members of the IFN-inducible p200 protein family. Both proteins can sense dsDNA in the cytoplasm. However, upon sensing dsDNA, only the Aim2 protein through its pyrin domain can form an inflammasome to activate caspase-1 and induce cell death. Given that the p202 protein has been predicted to inhibit the activation of caspase-1 by the Aim2 protein and that increased levels of the p202 protein in female mice of certain strains are associated with lupus susceptibility, we compared the expression of Aim2 and Ifi202 genes between Aim2-deficient and age-matched wild-type mice. We found that the Aim2 deficiency in immune cells stimulated the expression of Ifi202 gene. The increased levels of the p202 protein in cells were associated with increases in the expression of IFN-ß, STAT1, and IFN-inducible genes. Moreover, after knockdown of Aim2 expression in the murine macrophage cell line J774.A1, IFN-ß treatment of cells robustly increased STAT1 protein levels (compared with those of control cells), increased the activating phosphorylation of STAT1 on Tyr-701, and stimulated the activity of an IFN-responsive reporter. Notably, the expression of Aim2 in non-lupus-prone (C57BL/6 and B6.Nba2-C) and lupus-prone (B6.Nba2-ABC) splenic cells and in a murine macrophage cell line that overexpressed p202 protein was found to be inversely correlated with Ifi202. Collectively, our observations demonstrate an inverse correlation between Aim2 and p202 expressions. We predict that defects in Aim2 expression within immune cells contribute to increased susceptibility to lupus.

Development of murine lupus involves the combined genetic contribution of the SLAM and FcgammaR intervals within the Nba2 autoimmune susceptibility locus.

Autoantibodies are of central importance in the pathogenesis of Ab-mediated autoimmune disorders. The murine lupus susceptibility locus Nba2 on chromosome 1 and the syntenic human locus are associated with a loss of immune tolerance that leads to antinuclear Ab production. To identify gene intervals within Nba2 that control the development of autoantibody-producing B cells and to determine the cellular components through which Nba2 genes accomplish this, we generated congenic mice expressing various Nba2 intervals where genes for the FcgammaR, SLAM, and IFN-inducible families are encoded. Analysis of congenic strains demonstrated that the FcgammaR and SLAM intervals independently controlled the severity of autoantibody production and renal disease, yet are both required for lupus susceptibility. Deregulated homeostasis of terminally differentiated B cells was found to be controlled by the FcgammaR interval where FcgammaRIIb-mediated apoptosis of germinal center B cells and plasma cells was impaired. Increased numbers of activated plasmacytoid dendritic cells that were distinctly CD19+ and promoted plasma cell differentiation via the proinflammatory cytokines IL-10 and IFNalpha were linked to the SLAM interval. These findings suggest that SLAM and FcgammaR intervals act cooperatively to influence the clinical course of disease through supporting the differentiation and survival of autoantibody-producing cells.

Lessons in Innate and Allergic Immunity From Dust Mite Feces and Tick Bites.

Allergic diseases represent a major cause of morbidity in modern industrialized and developing countries. The origins and development of allergic immune responses have proven difficult to unravel and remain an important scientific objective. House dust mites (HDM) and ticks represent two important causes of allergic disease. Investigations into HDM fecal particles and tick bites have revealed insights which have and will continue to shape our understanding of allergic immunity. In the present review, focus is given to the role of innate immunity in shaping the respective responses to HDM and ticks. The HDM fecal particle represents a rich milieu of molecules that can be recognized by pathogen-recognition receptors of the innate immune system. Factors in tick saliva and/or tissue damage resultant from tick feeding are thought to activate innate immune signaling that promotes allergic pathways. Recent evidence indicates that innate sensing involves not only the direct recognition of allergenic agents/organisms, but also indirect sensing of epithelial barrier disruption. Although fecal particles from HDM and bites from ticks represent two distinct causes of sensitization, both involve a complex array of molecules that contribute to an innate response. Identification of specific molecules will inform our understanding of the mechanisms that contribute to allergic immunity, however the key may lie in the combination of molecules delivered to specific sites in the body.

Proliferating CD8+ T Cell Infiltrates Are Associated with Improved Survival in Glioblastoma.

BACKGROUND: tumor-infiltrating lymphocytes are prognostic in many human cancers. However, the prognostic value of lymphocytes infiltrating glioblastoma (GBM), and roles in tumor control or progression are unclear. We hypothesized that B and T cell density, and markers of their activity, proliferation, differentiation, or function, would have favorable prognostic significance for patients with GBM. METHODS: initial resection specimens from 77 patients with IDH1/2 wild type GBM who received standard-of-care treatment were evaluated with multiplex immunofluorescence histology (mIFH), for the distribution, density, differentiation, and proliferation of T cells and B cells, as well as for the presence of tertiary lymphoid structures (TLS), and IFNγ expression. Immune infiltrates were evaluated for associations with overall survival (OS) by univariate and multivariate Cox proportional hazards modeling. RESULTS: in univariate analyses, improved OS was associated with high densities of proliferating (Ki67+) CD8+ cells (HR 0.36, p = 0.001) and CD20+ cells (HR 0.51, p = 0.008), as well as CD8+Tbet+ cells (HR 0.46, p = 0.004), and RORγt+ cells (HR 0.56, p = 0.04). Conversely, IFNγ intensity was associated with diminished OS (HR 0.59, p = 0.036). In multivariable analyses, adjusting for clinical variables, including age, resection extent, Karnofsky Performance Status (KPS), and MGMT methylation status, improved OS was associated with high densities of proliferating (Ki67+) CD8+ cells (HR 0.15, p < 0.001), and higher ratios of CD8+ cells to CD4+ cells (HR 0.31, p = 0.005). Diminished OS was associated with increases in patient age (HR 1.21, p = 0.005) and higher mean intensities of IFNγ (HR 2.13, p = 0.027). CONCLUSIONS: intratumoral densities of proliferating CD8 T cells and higher CD8/CD4 ratios are independent predictors of OS in patients with GBM. Paradoxically, higher mean intensities of IFNγ in the tumors were associated with shorter OS. These findings suggest that survival may be enhanced by increasing proliferation of tumor-reactive CD8+ T cells and that approaches may be needed to promote CD8+ T cell dominance in GBM, and to interfere with the immunoregulatory effects of IFNγ in the tumor microenvironment.

Chemokine Receptor Activation Enhances Memory B Cell Class Switching Linked to IgE Sensitization to Alpha Gal and Cardiovascular Disease.

Background: Recent studies have suggested that IgE sensitization to α-gal is associated with coronary artery disease (CAD). However, the B cell subtype(s) responsible for production of IgE to α-gal and mechanisms mediating this production remain elusive. Methods: Single cell multi-omics sequencing, was utilized to phenotype B cells obtained from 60 subjects that had undergone coronary angiography in whom serum IgE was evaluated by ImmunoCAP. Bioinformatics approaches were used to identify B cell subtype(s) and transcriptomic signatures associated with α-gal sensitization. In vitro characterization of chemokine/chemokine receptor pairs on switched memory B cells associated with IgE to α-gal was performed. Results: Of the 60 patients, 17 (28%) were positive for IgE to α-gal. CITESeq identified CCR6+ class-switched memory (SWM) B cells and CXCR4 expresssion on these CCR6+ SWM B cells as significantly associated with IgE sensitization to α-gal but not to other common allergens (peanut or inhalants). In vitro studies of enriched human B cells revealed significantly greater IgE on SWM B cells with high CCR6 and CXCR4 expression 10 days after cells were treated with IL-4 and CD40 to stimulate class switch recombination. Both CCL20 (CCR6 ligand) and CXCL12 (ligand for CXCR4) increased the expression of IgE on SWM B cells expressing their receptors. However, they appeared to have unique pathways mediating this effect as only CCL20 increased activation-induced cytidine deaminase (AID), while CXCL12 drove proliferation of CXCR4+ SWM B cells. Lastly, correlation analysis indicated an association between CAD severity and the frequency of both CCR6+ SWM and CXCR4+ SWM B cells. Conclusions: CCR6+ SWM B cells were identified as potential producers of IgE to α-gal in CAD patients. Additionally, our findings highlighted non-chemotaxis roles of CCL20/CCR6 and CXCL12/CXCR4 signaling in mediating IgE class switching and cell proliferation of SWM B cells respectively. Results may have important implications for a better understanding and better therapeutic approaches for subjects with IgE sensitization to α-gal.

BCMA is essential for the survival of long-lived bone marrow plasma cells.

Long-lived humoral immunity is manifested by the ability of bone marrow plasma cells (PCs) to survive for extended periods of time. Recent studies have underscored the importance of BLyS and APRIL as factors that can support the survival of B lineage lymphocytes. We show that BLyS can sustain PC survival in vitro, and this survival can be further enhanced by interleukin 6. Selective up-regulation of Mcl-1 in PCs by BLyS suggests that this alpha-apoptotic gene product may play an important role in PC survival. Blockade of BLyS, via transmembrane activator and cyclophilin ligand interactor-immunoglobulin treatment, inhibited PC survival in vitro and in vivo. Heightened expression of B cell maturation antigen (BCMA), and lowered expression of transmembrane activator and cyclophilin ligand interactor and BAFF receptor in PCs relative to resting B cells suggests a vital role of BCMA in PC survival. Affirmation of the importance of BCMA in PC survival was provided by studies in BCMA-/- mice in which the survival of long-lived bone marrow PCs was impaired compared with wild-type controls. These findings offer new insights into the molecular basis for the long-term survival of PCs.

Affinity of antigen encounter and other early B-cell signals determine B-cell fate.

Three possible effector fates await the naïve follicular B cell following antigen stimulation in thymus-dependent reactions. Short-lived plasma cells produce an initial burst of germline-encoded protective antibodies, and long-lived plasma cells and memory B cells arise from the germinal center and function to enhance and sustain the humoral immune response. The inherent B-cell receptor affinity of naïve follicular B cells and the contribution of other early B-cell signals pre-determines the pattern of transcription factor expression and the differentiation path taken by these cells. High initial B-cell receptor affinity shunts naïve follicular B-cell clones towards the short-lived plasma cell fate, whereas modest-affinity clones are skewed towards a plasma cell fate and low-affinity clones are recruited into the germinal center and are selected for both long-lived plasma cells and memory B cell pathways. In the germinal center reaction, increased levels of the transcription factor interferon regulatory factor-4 drive the molecular program that dictates differentiation into the long-lived plasma cell phenotype but has no impact on the memory B cell compartment. We hypothesize that graded interferon regulatory factor-4 levels driven by signals to B cells, including B-cell receptor signal strength, are responsible for this branch point in the B-cell terminal differentiation pathway.

Deregulation of c-Myc Confers distinct survival requirements for memory B cells, plasma cells, and their progenitors.

Deregulation of the c-Myc oncogene is tightly associated with human and murine plasma cell (PC) neoplasms. Through the analysis of Ag-specific B cell responses in mice where Myc is targeted to the Igh Calpha locus, we show here that c-Myc dramatically impairs the primary and secondary Ab response. This impairment is differentiation stage specific, since germinal center B cell formation, affinity maturation, and class switch recombination were intact. Examination of PC viability revealed that c-Myc triggered apoptosis only upon final maturation when Ab is secreted and is resistant to the survival factor BAFF (B cell-activating factor belonging to the TNF family). In contrast, PC precursors (PC(pre)) that ultimately give rise to mature PCs survived normally and vigorously expanded with BAFF signaling. We further show that c-Myc also facilitates the apoptosis of memory B cells. Thus, Calpha-Myc controls both cellular arms of long-lived B cell immunity than previously anticipated. Only when deregulation of c-Myc was combined with enforced Bcl-x(L) expression were mature PCs able to survive in response to BAFF. These data indicate that the survival requirements for tumor-susceptible PC(pre) and PCs are distinct and that tumor progression likely develops as PC(pre) transition to functional PCs when apoptotic pathways such as members of the Bcl-2 family are disabled.