BLIMP-1 is insufficient to induce antibody secretion in the absence of IRF4 in DT40 cells.

Differentiation of B cells into antibody-secreting cells (ASCs), plasmablasts and plasma cells is regulated by a network of transcription factors. Within this network, factors including PAX5 and BCL6 prevent ASC differentiation and maintain the B cell phenotype. In contrast, BLIMP-1 and high IRF4 expression promote plasma cell differentiation.

Bach2 regulates AID-mediated immunoglobulin gene conversion and somatic hypermutation in DT40 B cells.

The transcription factor Bach2 is required for germinal center formation, somatic hypermutation (SHM), and class-switch recombination (CSR) of immunoglobulins. SHM and CSR are initiated by activation-induced cytidine deaminase (AID) which has potential to induce human B cell lymphoma. To understand the role of Bach2 in AID-mediated immunoglobulin gene diversification processes, we established a BACH2-deficient DT40 B cell line.

IRF4 Deficiency Leads to Altered BCR Signalling Revealed by Enhanced PI3K Pathway, Decreased SHIP Expression and Defected Cytoskeletal Responses.

The graded expression of transcription factor interferon regulatory factor 4 (IRF4) regulates B cell development and is critical for plasma cell differentiation. However, the mechanisms, by which IRF4 elicits its crucial tasks, are largely unknown. To characterize the molecular targets of IRF4 in B cells, we established an IRF4-deficient DT40 B cell line.

Regulation of B Cell to Plasma Cell Transition within the Follicular B Cell Response.

Persistent humoral immunity depends on the follicular B cell response and on the generation of somatically mutated high-affinity plasma cells and memory B cells. Upon activation by an antigen, cognately activated follicular B cells and follicular T helper (TFH ) cells initiate germinal centre (GC) reaction during which high-affinity effector cells are generated. The differentiation of activated follicular B cells into plasma cells and memory B cells is guided by complex selection events, both at the cellular and molecular level.

Cartilage-hair hypoplasia: follow-up of immunodeficiency in two patients.

Purpose: To study the changes in the immunological status in 2 children with cartilage hair hypoplasia (CHH).

Methods: A 4-6 year immunological follow-up from infancy.

Results: In infancy the children presented a combined T cell and B cell immunodeficiency which partly resolved in time.

Alternate pathways for Bcl6-mediated regulation of B cell to plasma cell differentiation.

The transcription factor Bcl6 regulates germinal center formation and differentiation of B cells into high-affinity antibody-producing plasma cells. The direct double-negative regulatory circuit between Bcl6 and Blimp-1 is well established. We now reveal alternative mechanisms for Bcl6-mediated regulation of B-cell differentiation to plasma cells and show with DT40 cells that Bcl6 directly promotes the expression of Bach2, a known suppressor of Blimp-1.

Concerted action of Helios and Ikaros controls the expression of the inositol 5-phosphatase SHIP.

Ikaros family transcription factors have a key role in lymphoid development, and their aberrant function contributes to a multitude of lymphoid malignancies. Ikaros and Helios bind to similar DNA sequences, and Helios associates with Ikaros-containing chromatin remodeling complexes. Previously, we have shown that loss of Ikaros leads to diminished BCR-signaling strength.

Avian model for B-cell immunology--new genomes and phylotranscriptomics.

The purpose of this review is to discuss the use of chicken and other model organisms in the study of B-cell development and function as well as to highlight the opportunities afforded by the expanded genome-sequencing efforts. A brief introduction on chicken B-cell biology is followed by discussion of somatic cell reverse genetic approaches using the DT40 cell line. The unique advantages of the DT40 system are emphasized with discussion on B-cell receptor signalling research as well as on DNA repair and mechanisms of immunoglobulin diversification.

DT40 mutants: a model to study transcriptional regulation of B cell development and function.

A key issue in understanding the hematopoietic system and B cell biology is to define the function of transcription factors. B lymphocyte development and function is controlled by a hierarchy of transcription factors including PU.1, Ikaros, E2A, EBF, Pax5 and Aiolos.

Aiolos controls gene conversion and cell death in DT40 B cells.

The Ikaros family transcription factor Aiolos is important for B cell function, since B cells of Aiolos-null mutant mice exhibit an activated phenotype, enhanced B-cell receptor (BCR) signalling response and develop a systemic lupus erythematosus (SLE) type autoimmune disease. Aiolos has also been reported to interact with anti-apoptotic Bcl-2 and Bcl-x(L) in T cells, but whether Aiolos regulates cell death has not been studied in B cells. Here we show that the disruption of Aiolos in the DT40 B cell line induces a cell death sensitive phenotype, as the Aiolos(-/-) cells are more prone to apoptosis by nutritional stress, BCR cross-linking, UV- or gamma-irradiation.

Pax5--a critical inhibitor of plasma cell fate.

Paired box protein 5 (Pax5) is essential for early B cell commitment as well as for B cell development, and continuous expression of Pax5 is required throughout the B cell lineage to maintain the functional identity of B cells. During B cell activation, Pax5 is downregulated before terminal differentiation into antibody-secreting plasma cells, and enforced expression of Pax5 prevents plasmacytic development. Recently, loss of Pax5 was shown to result in the substantial transition to a plasma cell state, demonstrating a functionally significant role for Pax5 in the regulation of terminal B cell differentiation.

Novel method for cell debris removal in the flow cytometric cell cycle analysis using carboxy-fluorescein diacetate succinimidyl ester.

Background: Cell cycle analysis with flow cytometry using propidium iodide (PI) can be difficult in some cases because of the cell debris. Here, we introduce debris removal using intranuclear protein staining (DRIPS), a novel method for separating intact nuclei and cell debris to different populations using carboxy-fluorescein diacetate succinimidyl ester (CFSE).

Methods: To study the apoptosis-sensitivity, chicken DT40 B cell lymphoma cell line was gamma irradiated.

Loss of Pax5 promotes plasma cell differentiation.

Pax5 is indispensable for the commitment of early lymphoid progenitors to the B cell lineage as well as for the development of B cells. To better understand the functional importance of Pax5 at the later stages of B cell differentiation, we established a Pax5-deficient DT40 B cell line. The Pax5(-/-) cells exhibited slower growth, decreased surface IgM expression, and total loss of B cell receptor signaling.

Ikaros has a crucial role in regulation of B cell receptor signaling.

The transcription factor Ikaros, a key regulator of hematopoiesis, has an essential role in lymphocyte development. In mice, fetal lymphoid differentiation is blocked in the absence of Ikaros, and whereas T cells develop postnatally, B cells are totally absent. The significance of Ikaros in the B cell development is evident, but how Ikaros regulates B cell function has neither been established nor previously been studied with B cells that lack Ikaros expression.

Development of early PCLP1-expressing haematopoietic cells within the avian dorsal aorta.

The first haematopoietic stem cells (HSC) develop in the dorsal aorta as haematopoietic intra-aortic clusters (HIAC). To evaluate the initial steps of definitive haematopoiesis, we have studied the emergence and the expression profile of podocalyxin-like protein 1 (PCLP1)-expressing cells in early chick embryos. Here we demonstrate that at embryonic day 2 (E2), the PCLP1+ cells are present in the splanchnic mesoderm and in the ventral lining of the paired dorsal aorta.

CD4+ CD25+ T cells with the phenotypic and functional characteristics of regulatory T cells are enriched in the synovial fluid of patients with rheumatoid arthritis.

CD4(+) CD25(+) regulatory T (T(reg)) cells play a critical role in the maintenance of peripheral tolerance and the prevention of autoimmunity. In the present study, we have explored the characteristics of CD4(+) CD25(+) T(reg) cells in patients with rheumatoid arthritis (RA). The frequency and phenotype of CD4(+) CD25(+) T cells in paired samples of synovial fluid (SF) and peripheral blood (PB) from patients with RA and PB from normal controls were analysed.

Avian Helios and evolution of the Ikaros family.

Abstract Helios (Znfn1a2) is an Ikaros-related lymphoid regulatory protein with possible involvement in T-cell development and function as well as in the early events of haematopoietic stem cell differentiation. To evaluate the role of Helios in avian haemato/lymphopoiesis, we have characterized the avian Helios gene. In contrast to studies in mouse and human, we have found that the highly conserved avian Helios encodes a novel exon and three isoforms.

Chicken B-cell-activating factor: regulator of B-cell survival in the bursa of fabricius.

Previous studies on mammals have demonstrated that a tumour necrosis factor family member, B-cell-activating factor (BAFF) (BlyS, TALL-1), is mainly produced by myeloid and dendritic cells and that BAFF promotes B-cell differentiation and survival in a paracrine fashion. We have recently shown that BAFF is upregulated at the bursal stage of the avian B-cell development. We now show that the avian bursal B cells and B-cell lines, RP-9, RP-13 and DT40, express chicken BAFF (cBAFF).

Phenotypic characterization of regulatory T cells in the human decidua.

Pregnancy is a unique situation for the maternal immune system. We have studied and identified a CD4+CD25+ regulatory T (Treg) cell population isolated from the human decidua. This mucosal surface in the uterus is in direct contact with semiallogenic fetal cells.

Identification of a novel cytokine-like transcript differentially expressed in avian gammadelta T cells.

Chicken represents a species with a high frequency of gammadelta T cells in peripheral blood, suggesting an important function. To elucidate the genes specific for the avian gammadelta T cells, the suppression subtractive hybridization (SSH) between gammadelta and alphabeta T cells was used. The SSH library, which was successfully enriched for the TCR gamma and delta (both V and C region) sequences, provided gammadelta T-cell-specific genes, including, for example, the ribosomal proteins, signaling and structural molecules, and molecules related to transcription and translation.

Insight into lymphoid development by gene expression profiling of avian B cells.

The avian immune system provides an excellent model to track B-cell development from prebursal stem cells throughout B-cell differentiation and maturation. Bursal B cells are uniquely positioned at the crossroads of B-cell development, having properties of both stem cells and of mature B cells, as demonstrated by their ability to reconstruct the bursal B-cell compartment and to express and diversify the B-cell receptor at their cell surface. To understand avian B-cell development better, we determined the gene expression profile of different B-cell stages using a bursal expressed sequence tag array.

Interaction between Borrelia burgdorferi and immature human dendritic cells.

Antigen uptake and the following maturation of dendritic cells (DCs) are pivotal to the initiation of specific antimicrobial immune responses. DCs also play an important role in the recruitment and activation of the cells of the innate immune system. We have examined the interactions of DCs with Borrelia burgdorferi to find explanations for the difficulties the human immune system has in dealing with the bacterium.