Search

With the introduction of next generation sequencing methods, such as RNA sequencing, it has become apparent that alterations in the non-coding regions of our genome are important in the development of cancer. Particularly interesting is the class of long non-coding RNAs (lncRNAs), including the recently described subclass of circular RNAs (circRNAs), which display tissue- and cell-type specific expression patterns and exert diverse regulatory functions in the cells. B-cells undergo complex and tightly regulated processes in order to develop from antigen naïve cells residing in the bone marrow to the highly diverse and competent effector cells circulating in peripheral blood. These processes include V(D)J recombination, rapid proliferation, somatic hypermutation and clonal selection, posing a risk of malignant transformation at each step. The aim of this review is to provide insight into how lncRNAs including circRNAs, participate in normal B-cell differentiation, and how deregulation of these molecules is involved in the development of B-cell malignancies. We describe the prognostic value and functional significance of specific deregulated lncRNAs in diseases such as acute lymphoblastic leukemia, chronic lymphocytic leukemia, mantle cell lymphoma, diffuse large B-cell lymphoma, follicular lymphoma, Burkitt lymphoma and multiple myeloma, and we provide an overview of the current knowledge on the role of circRNAs in these diseases.

Long noncoding (Inc)RNAs have emerged as essential mediators of cellular biology, differentiation and malignant transformation. LncRNAs have a broad range of possible functions at the transcriptional, posttranscriptional and protein level and their aberrant expression significantly contributes to the hallmarks of cancer cell biology. In addition, their high tissue- and cell-type specificity makes lncRNAs especially interesting as biomarkers, prognostic factors or specific therapeutic targets. Here, we review current knowledge on lncRNA expression changes during normal B-cell development, indicating essential functions in the differentiation process. In addition we address lncRNA deregulation in B-cell malignancies, the putative prognostic value of this as well as the molecular functions of multiple deregulated lncRNAs. Altogether, the discussed work indicates major roles for lncRNAs in normal and malignant B cells affecting oncogenic pathways as well as the response to common therapeutics.

B-cell development in the bone marrow is followed by specification into functional subsets in the spleen, including marginal zone (MZ) B-cells. MZ B-cells are classically characterized by T-independent antigenic responses and require the elaboration of distinct gene expression programs for development. Given their role in gene regulation, it is not surprising that microRNAs are important factors in B-cell development. Recent work demonstrated that deficiency of the NFκB feedback regulator, miR-146a, led to a range of hematopoietic phenotypes, but B-cell phenotypes have not been extensively characterized. Here, we found that miR-146a-deficient mice demonstrate a reduction in MZ B-cells, likely from a developmental block. Utilizing high-throughput sequencing and comparative analysis of developmental stage-specific transcriptomes, we determined that MZ cell differentiation was impaired due to decreases in Notch2 signaling. Our studies reveal miR-146a-dependent B-cell phenotypes and highlight the complex role of miR-146a in the hematopoietic system.

Most serum immunoglobulin M (IgM) is "natural IgM", which is produced apparently spontaneously by a distinct subset of B cells requiring no exogenous antigenic or microbial stimuli. Natural IgM is an evolutionarily conserved molecule and reacts with a variety of epitopes expressed on both self- and non-self antigens. It has long been understood that secreted (s) IgM contributes to the removal of altered self-antigens, such as apoptotic and dying cells. As we outline in this review, it is thought that this sIgM housekeeping function removes potential triggers of autoresponse induction. However, we recently demonstrated an unexpected and distinct role for sIgM in the control of autoreactive B cells: the regulation of bone marrow B cell development. The absence of sIgM blocked pro- to pre- B-cell transition and greatly altered the BCR repertoire of the developing B cells and the peripheral B-cell pools in genetically engineered mice. This finding strongly suggests that IgM is critical for B-cell central tolerance induction. Given that treatment of sIgM-deficient mice with polyclonal IgM corrected these developmental defects, therapeutic application of IgM could be of clinical relevance in the treatment of some B-cell-mediated autoimmune diseases.

Background Severe combined immunodeficiency (SCID) represents congenital disorders characterized by a deficiency of T cells caused by arrested development in the thymus. Yet the nature of these developmental blocks has remained elusive because of the difficulty of taking thymic biopsy specimens from affected children. Objective We sought to identify the stages of arrest in human T-cell development caused by various major types of SCID. Methods We performed transplantation of SCID CD34 + bone marrow stem/progenitor cells into an optimized NSG xenograft mouse model, followed by detailed phenotypic and molecular characterization using flow cytometry, immunoglobulin and T-cell receptor spectratyping, and deep sequencing of immunoglobulin heavy chain (IGH) and T-cell receptor δ (TRD) loci. Results Arrests in T-cell development caused by mutations in IL-7 receptor α (IL7RA) and IL-2 receptor γ (IL2RG) were observed at the most immature thymocytes much earlier than expected based on gene expression profiling of human thymocyte subsets and studies with corresponding mouse mutants. T-cell receptor rearrangements were functionally required at the CD4 − CD8 − CD7 + CD5 + stage given the developmental block and extent of rearrangements in mice transplanted with Artemis-SCID cells. The xenograft model used is not informative for adenosine deaminase–SCID, whereas hypomorphic mutations lead to less severe arrests in development. Conclusion Transplanting CD34 + stem cells from patients with SCID into a xenograft mouse model provides previously unattainable insight into human T-cell development and functionally identifies the arrest in thymic development caused by several SCID mutations.

Interleukin-7 (IL-7) is essential for lymphocyte development. To identify the functional subdomains in the cytoplasmic tail of the IL-7 receptor (IL-7R) &alpha, chain, here, we constructed a series of IL-7R&alpha, deletion mutants. We found that IL-7R&alpha, deficient hematopoietic progenitor cells (HPCs) gave rise to B cells both in vitro and in vivo when a wild-type (WT) IL-7R&alpha, chain was introduced, however, no B cells were observed under the same conditions from IL-7R&alpha, deficient HPCs with introduction of the exogenous IL-7R&alpha, subunit, which lacked the amino acid region at positions 414&ndash, 441 (d414&ndash, 441 mutant). Signal transducer and activator of transcription 5 (STAT5) was phosphorylated in cells with the d414&ndash, 441 mutant, similar to that in WT cells, in response to IL-7 stimulation. In contrast, more truncated STAT5 (tSTAT5) was generated in cells with the d414&ndash, 441 mutant than in WT cells. Additionally, the introduction of exogenous tSTAT5 blocked B lymphopoiesis but not myeloid cell development from WT HPCs in vivo. These results suggested that amino acids 414&ndash, 441 in the IL-7R&alpha, chain formed a critical subdomain necessary for the supportive roles of IL-7 in B-cell development.

During the last three decades, a number of B-lymphocyte specific surface antigens have been defined some of which may also show activation/differentiation specific expression. Here, we review the various signaling events and the receptor-ligand interactions for B-cell development, activation and differentiation. Our discussion and presentation include reviewing the in vivo and in vitro mechanisms. Focus is on the experiments that give us valuable insights into the B cell signaling mechanisms in vitro. Three significant pathways in B-cell development – c-Kit, FLT-3 and IL-7 signaling pathways are elucidated upon. Both antigen dependent and antigen independent mechanisms of B cell stimulation are also reviewed.

It is well established that all camelids have unique antibodies circulating in their blood. Unlike antibodies from other species, these special antibodies are devoid of light chains and are composed of a heavy-chain homodimer. These so-called heavy-chain antibodies (HCAbs) are expressed after a V-D-J rearrangement and require dedicated constant gamma-genes. An immune response is raised in these so-called heavy-chain antibodies following classical immunization protocols. These HCAbs are easily purified from serum, and the antigen-binding fragment interacts with parts of the target that are less antigenic to conventional antibodies. Since the antigen-binding site of the dromedary HCAb is comprised in one single domain, referred to as variable domain of heavy chain of HCAb (VHH) or nanobody (Nb), we designed a strategy to clone the Nb repertoire of an immunized dromedary and to select the Nbs with specificity for our target antigens. The monoclonal Nbs are well produced in bacteria, are very stable and highly soluble, and bind their cognate antigen with high affinity and specificity. We have successfully developed recombinant Nbs for research purposes, as probe in biosensors, to diagnose infections, and to treat diseases like cancer or trypanosomosis.

The receptor-like tyrosine phosphatase CD45 regulates antigen receptor signaling by dephosphorylating the C-terminal inhibitory tyrosine of the src family kinases. However, despite its abundance, the function of the large, alternatively spliced extracellular domain of CD45 has remained elusive. We used normally spliced CD45 transgenes either incorporating a phosphatase-inactivating point mutation or lacking the cytoplasmic domain to uncouple the enzymatic and noncatalytic functions of CD45 in lymphocytes. Although these transgenes did not alter T-cell signaling or development irrespective of endogenous CD45 expression, both partially rescued the phenotype of CD45-deficient B cells. We identify a noncatalytic role for CD45 in regulating tonic, but not antigen-mediated, B-cell antigen receptor (BCR) signaling through modulation of the function of the inhibitory coreceptor CD22. This finding has important implications for understanding how naïve B cells maintain tonic BCR signaling while restraining inappropriate antigen-dependent activation to preserve clonal "ignorance.".

The bursa of Fabricius (BF) is a lymphoepithelial organ found only in birds. Differences in morphology of BF could play an important role in immune response. The objective of this study was to investigate the histological and histochemical characteristics of the bursa of Fabricius in the long-legged buzzard (Buteo rufinus). The material for the study comprised bursa samples obtained from three long-legged buzzards with permission of the General Directorate of Nature Protection and National Parks (Ankara, Turkey). Briefly, interfollicular epithelium (IFE) was shown to be columnar in shape and not to contain goblet cells. Reticular fibers were located in interfollicular septae. Each lymphoid follicle in the bursa of Fabricius in the long-legged buzzard was remarkably linked to the follicle associated epithelium (FAE). Namely, FAE has been reported to stimulate antibody production by transferring antigens to the medulla and have a leading role in developing of local immune response. Among the others, the species-specific differences in bursa of Fabricius morphology of long-legged buzzard (Buteo rufinus) also might support the continuity of this species in nature.

Mature B cells are grouped into two major subsets, B-1 and B-2, believed to derive from separate lineages. We have recently shown that B-1 cells, which are characterized by CD5 surface expression, specifically exhibit significant levels of the tyrosine kinase Lck in man. Here we show that also in mice Lck expression is restricted to B-1 cells and address the potential role of Lck in B-1 cell development and activation. Using as a model an Lck-/- mouse, we show that, while dispensable for B-1 cell development, Lck is required for full and sustained activation of the tyrosine phosphorylation and MAP kinase cascades triggered by the BCR in CD5+, B-1 cells. The data suggest a potential role for Lck in the achievement of the higher activation threshold required for productive BCR signaling in B-1 as compared to B-2 cells.

Integrin-mediated adhesion and B cell antigen receptor (BCR) signaling play a critical role in B cell development and function, including antigen-specific B cell differentiation. Here we show that the BCR controls integrin alpha4beta1 (VLA-4)-mediated adhesion of B cells to vascular cell adhesion molecule-1 and fibronectin. Molecular dissection of the underlying signaling mechanism by a combined biochemical, pharmacological, and genetic approach demonstrates that this BCR-controlled integrin-mediated adhesion requires the (consecutive) activation of Lyn, Syk, phosphatidylinositol 3-kinase, Bruton's tyrosine kinase (Btk), phospholipase C (PLC)gamma2, IP3R-mediated Ca2+ release, and PKC. In contrast, activation of mitogen-activated protein kinase kinase (MEK) or extracellular signal-regulated kinase (ERK) is not required, and simultaneous activation of MEK, ERK, and PKB is not sufficient either. Furthermore, Btk is also involved in the control of integrin-mediated adhesion of preB cells. The control of integrin alpha4beta1-mediated B cell adhesion by the BCR involves cytoskeletal reorganization and integrin clustering. These results reveal a novel function for the BCR and Btk, i.e., regulation of integrin alpha4beta1 activity, thereby providing new insights into the control of B cell development and differentiation, as well as into the pathogenesis of the immunodeficiency disease X-linked agammaglobulineamia (XLA).

BACKGROUND:Adenosine deaminase (ADA) deficiency causes severe cellular and humoral immune defects and dysregulation because of metabolic toxicity. Alterations in B-cell development and function have been poorly studied. Enzyme replacement therapy (ERT) and hematopoietic stem cell (HSC) gene therapy (GT) are therapeutic options for patients lacking a suitable bone marrow (BM) transplant donor. OBJECTIVE: We sought to study alterations in B-cell development in ADA-deficient patients and investigate the ability of ERT and HSC-GT to restore normal B-cell differentiation and function. METHODS:Flow cytometry was used to characterize B-cell development in BM and the periphery. The percentage of gene-corrected B cells was measured by using quantitative PCR. B cells were assessed for their capacity to proliferate and release IgM after stimulation. RESULTS:Despite the severe peripheral B-cell lymphopenia, patients with ADA-deficient severe combined immunodeficiency showed a partial block in central BM development. Treatment with ERT or HSC-GT reverted most BM alterations, but ERT led to immature B-cell expansion. In the periphery transitional B cells accumulated under ERT, and the defect in maturation persisted long-term. HSC-GT led to a progressive improvement in B-cell numbers and development, along with increased levels of gene correction. The strongest selective advantage for ADA-transduced cells occurred at the transition from immature to naive cells. B-cell proliferative responses and differentiation to immunoglobulin secreting IgM after B-cell receptor and Toll-like receptor triggering were severely impaired after ERT and improved significantly after HSC-GT. CONCLUSIONS:ADA-deficient patients show specific defects in B-cell development and functions that are differently corrected after ERT and HSC-GT.

Patients without chronic graft-versus-host disease (cGVHD) have robust B cell reconstitution and are able to maintain B cell homeostasis after allogeneic hematopoietic stem cell transplantation (HSCT). To determine whether B lymphopoiesis differs before cGVHD develops, we examined bone marrow (BM) biopsies for terminal deoxynucleotidyl transferase (TdT) and PAX5 immunostaining early post-HSCT at day 30 when all patients have been shown to have high B cell activating factor (BAFF) levels. We found significantly greater numbers of BM B cell precursors in patients who did not develop cGVHD compared with those who developed cGVHD (median = 44 vs 2 cells/high powered field [hpf]; respectively; P < .001). Importantly, a significant increase in precursor B cells was maintained when patients receiving high-dose steroid therapy were excluded (median = 49 vs 20 cells/hpf; P = .017). Thus, we demonstrate the association of BM B cell production capacity in human GVHD development. Increased BM precursor B cell number may serve to predict good clinical outcome after HSCT.

Due to a limited range of immunoglobulin (Ig) genes, cattle and several other domestic animals rely on postrecombinatorial amplification of the primary repertoire. We report that activation-induced cytidine deaminase (AID) is strongly expressed in the fetal bovine ileal Peyer's patch and spleen but not in fetal bone marrow. The numbers of IGHV (immunoglobulin heavy chain variable) mutations correlate with AID expression. The mutational profile in the fetuses is similar to postnatal and immunized calves, with targeting of complementarity-determining region (CDR) over framework region (FR), preference of replacement over silent mutations in CDRs but not in FRs, and targeting of the AID hotspot motif RGYW/WRCY. Statistical analysis indicates negative selection on FRs and positive selection on CDRs. Our results suggest that AID-mediated somatic hypermutation and selection take place in bovine fetuses, implying a role for AID in the diversification of the primary antibody repertoire in the absence of exogenous antigens.

International audience; B-cell fate and responses are modulated by soluble mediators and direct cellular interactions. Migration properties also vary during differentiation, commitment and activation. In many cells, modulation of responses to stimuli involves cell surface glycans, whose architecture depends on the simultaneous expression of multiple enzymes. By looking at the glycosylation-related gene expression patterns among B cell populations, we determined in this study that the strongest variations were observed for CSGalNAcT-1 and EXTL1. These are enzymes involved in the biosynthesis of alternative forms of glycosaminoglycans, namely chondroitin sulphate and heparan sulphate respectively. These two enzymes showed inverse fluctuations in progenitors, resting B cells and activated B cells, suggesting a developmentally regulated switch between chondroitin and heparan sulphate synthesis. To explore whether these variations contributed to optimal B cell differentiation, we over-expressed EXTL1 in the B-cell lineage of transgenic mice, yielding a partial differentiation blockade at the pro-B to pre-B transition. In the periphery, this defect was almost fully compensated for in vivo, with normal-size B-cell compartments and normal serum immunoglobulin levels in the transgenic EXTL1 mice. The peripheral B cells from EXTL1 transgenics were only affected with regard to their in vitro responses to polyclonal activation, showing reduced proliferation. Together the data suggest that, despite their low amounts in lymphocytes, the heparan sulphate chains decorating the endogenous glycosaminoglycans appear to be regulators of B-cell physiology.

Objective It is difficult to identify a single causative factor for inflammatory arthritis because of the multifactorial nature of the disease. This study was undertaken to dissect the molecular complexity of systemic inflammatory disease, utilizing a combined approach of mutagenesis and systematic phenotype screening in a murine model. Methods In a large-scale N-ethyl-N-nitrosourea mutagenesis project, the Ali14 mutant mouse strain was established because of dominant inheritance of spontaneous swelling and inflammation of the hind paws. Genetic mapping and subsequent candidate gene sequencing were conducted to find the causative gene, and systematic phenotyping of Ali14/+ mice was performed in the German Mouse Clinic. Results A novel missense mutation in the phospholipase Cγ2 gene (Plcg2) was identified in Ali14/+ mice. Because of the hyperreactive external entry of calcium observed in cultured B cells and other in vitro experiments, the Ali14 mutation is thought to be a novel gain-of-function allele of Plcg2. Findings from systematic screening of Ali14/+ mice demonstrated various phenotypic changes: an abnormally high T cell:B cell ratio, up-regulation of Ig, alterations in body composition, and a reduction in cholesterol and triglyceride levels in peripheral blood. In addition, spermatozoa from Ali14/+ mice failed to fertilize eggs in vitro, despite the normal fertility of the Ali14/+ male mice in vivo. Conclusion These results suggest that the Plcg2-mediated pathways play a crucial role in various metabolic and sperm functions, in addition to initiating and maintaining the immune system. These findings may indicate the importance of the Ali14/+ mouse strain as a model for systemic inflammatory diseases and inflammation-related metabolic changes in humans.

Ikaros is the founding member of a family of transcription factors. Ikaros proteins are required for the normal development of lymphocytes and other blood cell lineages, and have an important role in haematopoietic malignancies. Accumulating data link Ikaros to immune receptor rearrangement and signaling, and the balance between proliferation and differentiation of immune cells. The emerging picture is that Ikaros family members are critical for a functional immune system that confers protection against invading pathogens, while minimising the risk of leukaemic transformation, immune pathology, and autoimmunity.

Several HIV-1 neutralizing antibodies (e.g. 2F5, 4E10) have been shown to react with self-antigens, suggesting that effective humoral responses to HIV-1 may be constrained by the tolerization of HIV-reactive B cells that also recognize self-antigens. I have tracked the development of 2F5-like HIV-1 gp41 membrane proximal external region (MPER)-reactive B cells throughout ontogeny using B-cell tetramer reagents. In BL/6 mice, MPER-binding populations are lost during normal B-cell development and immunization with HIV-1 MPER antigen does not elicit robust humoral responses. I have identified Kynureninase as a self-antigen that is recognized by 2F5 antibody and, therefore, is a molecule that could mediate the developmental loss of B cells reactive to an epitope shared by HIV gp41 and Kynureninase. To recover these MPER-reactive cells, I describe and characterize a stromal-cell independent culture system that efficiently supports pro-B cell to IgM+ B-cell development with near normal levels of IgH and Igkappa diversity. B-cell development in vitro closely follows the patterns of development in vivo with culture derived (CD) B cells demonstrating characteristic patterns of surface antigen expression and gene activation. Immature and transitional B-cell compartments are reduced, due to the induction of tolerance, in the bone marrow of 3H9 IgH knockin mice ; however, cultures of 3H9 IgH knockin pro-B cells yields high frequencies of "forbidden", autoreactive IgM+ B cells. Furthermore, serum IgG autoantibody exceeded that present in autoimmune, C4-/- animals following the reconstitution of RAG-1-/- mice with IgM+ CD cells derived from BL/6 mice. I show that HIV-1 MPER-reactive B cells are recovered from both BL/6 and 2F5 IgH knockin bone marrow using this in vitro culture system. RAG-1-/- mice reconstituted with these culture-derived B and T cells generate strong germinal center and antibody responses to HIV-1 MPER antigens. These data demonstrate that the humoral immune response to this HIV-1 gp41 MPER antigen can be restored in mice when the constraints of B-cell tolerance have been relaxed.

Jejunal development occurs in a spatio-temporal pattern and is characterized by morphological and functional changes. To investigate jejunal development at the transcriptomic level, we performed microarray studies in 1–21-day-old chickens. Nine gene clusters were identified, each with a specific gene expression pattern. Subsequently, groups of genes with similar functions could be identified. Genes involved in morphological and functional development were highly expressed immediately after hatch with declining expression patterns afterwards. Immunological development can be roughly divided based on expression patterns into three processes over time; first innate response and immigration of immune cells, secondly differentiation and specialization, and thirdly maturation and immune regulation. We conclude that specific gene expression patterns coincide with the immunological, morphological, and functional development as measured by other methods. Our data show that transcriptomic approaches provide more detailed information on the biological processes underlying jejunal development.

Background The enhancer (Eμ3') of the immunoglobulin heavy chain locus (IGH) of the channel catfish (Ictalurus punctatus) has been well characterized. The functional core region consists of two variant Oct transcription factor binding octamer motifs and one E-protein binding μE5 site. An orthologue to the Oct2 transcription factor has previously been cloned in catfish and is a functionally active transcription factor. This study was undertaken to clone and characterize the Oct1 transcription factor, which has also been shown to be important in driving immunoglobulin gene transcription in mammals. Results An orthologue of Oct1, a POU family transcription factor, was cloned from a catfish macrophage cDNA library. The inferred amino acid sequence of the catfish Oct1, when aligned with other vertebrate Oct1 sequences, revealed clear conservation of structure, with the POU specific subdomain of catfish Oct1 showing 96% identity to that of mouse Oct1. Expression of Oct1 was observed in clonal T and B cell lines and in all tissues examined. Catfish Oct1, when transfected into both mammalian (mouse) and catfish B cell lines, unexpectedly failed to drive transcription from three different octamer-containing reporter constructs. These contained a trimer of octamer motifs, a fish V H promoter, and the core region of the catfish Eμ3' IGH enhancer, respectively. This failure of catfish Oct1 to drive transcription was not rescued by human BOB.1, a co-activator of Oct transcription factors that stimulates transcription driven by catfish Oct2. When co-transfected with catfish Oct2, Oct1 reduced Oct2 driven transcriptional activation. Electrophoretic mobility shift assays showed that catfish Oct1 (native or expressed in vitro) bound both consensus and variant octamer motifs. Putative N- and C-terminal activation domains of Oct1, when fused to a Gal4 DNA binding domain and co-transfected with Gal4-dependent reporter constructs were transcriptionally inactive, which may be due in part to a lack of residues associated with activation domain function. Conclusion An orthologue to mammalian Oct1 has been found in the catfish. It is similar to mammalian Oct1 in structure and expression. However, these results indicate that the physiological functions of catfish Oct1 differ from those of mammalian Oct1 and include negative regulation of transcription.

The m chain together with the surrogate light chain (lambda 5 + V pre-B) forms the pre-B cell receptor (pre-BCR)on the surface of pre-B cells before the assembly of the complete Ig molecule. The pre-BCR expression on the pre-B cells and formation of the surface IgM molecules, after the light chain rearragement on immature B cells are key checkpoints in the development of mature B lymphocytes. Ig class switch and the subsequent expresion of other Ig classes occurs only in mature B cells, after they have undergone germinal center reaction in the secondary lymphoid organs. Therefore, the disruption of the membrane exone of Ig m chain genes blocks the B cell development at the pre-B stage. C57BL/6 mice homozygous for this mutation (mMT/mMT C57BL/6)are unable to generate immature and mature B cells and subsequently lack Ig in their sera (Kitamura et al., Nature, 350, 423-426, 1991.). We have backcrossed the mMT mutation onto BALB/c mice. The offspring homozygous for the mutation (mMT/mMT BALB/c) were tested for the presence of immunoglobulins. Surprisingly, although unable to produce IgM, the mMT/mMT BALB/c mice had high levels of IgG in their sera. Further experiments showed the presence of different Ig classes and subclasses. Despite the IgG presence mMT/mMT BALB/c mice could not generate a specific antibody response. The spleen ad lymph node architecture of mMT/mMT C57BL/6 and mMT/mMT BALB/c mice was similar. Both strains lacked lymphatic follicles in the spleen and lymph nodes, yet anti mouse Ig-POD labeled antibodies stained cells scattered throughout the lymphatic organs of mMT/mMT BALB/c mice. In addition, the Ig positive cells were found also in the lamina propria of the intestinal mucosa. The presence of serum Ig and Ig positive cells strongly suggests that the developmental block at the pre-B cell stage is leaky in mMT/mMT BALB/c mice. The cause of this leakage could be attributed to occasional early class switch events. Our next aim is to clarify the underlaying mechanism of this phenotype.