Csnk2β, the Regulatory Subunit of Protein Kinase CK2, modulates Peripheral B Cell Development Repressing Notch2 Signaling and Promoting a Proper B-Cell Receptor Signal Transmission

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 566-566
Author(s):  
Fortunato Zaffino ◽  
Paolo Macaccaro ◽  
Alessandro Casellato ◽  
Elisa Mandato ◽  
Sabrina Manni ◽  
...  
Keyword(s):  
B Cells ◽  
Protein Kinase ◽  
B Cell ◽  
Protein Kinase Ck2 ◽  
Marginal Zone ◽  
Cell Development ◽  
B Cell Development ◽  
Class Switch ◽  
Kinase Ck2

Abstract Background. Serine-threonine protein kinase CK2 has been recently involved in the pathogenesis of B-cell tumors, such as B acute lymphoblastic leukemia, B chronic lymphocytic leukemia, mantle cell lymphoma and multiple myeloma. CK2 acts through a “non-oncogene” addiction mechanism to propel tumor growth, protecting from apoptosis by a phosphorylation-dependent “shielding” mechanism of pro-survival molecules and stimulating oncogenic kinases by helping folding and enzymatic activity. In addition, CK2 has been shown to enhance the transactivation potential of several transcription factors, such as STAT3, NF-κB and c-Myc. The existing data on CK2 function in B cell tumors suggest that this kinase might act as a “hub” downstream signals from surface membrane molecules, like the B-cell (BCR), growth factor and cytokine receptors, as well as from cell-intrinsic pathways – like proteotoxic and DNA-damage-related stress cascades. Aims and methods. To gain insights into the role of CK2 in B-lymphopoiesis and, consequently, in B-cell tumors, we generated CK2β conditional knockout (KO) mice in B-cells by crossing Csnk2β-Flox/Flox mice with CD19-CRE transgenic mice. Results. CK2 kinase activity was decreased in Csnk2β KO B cells. In the bone marrow (BM), Csnk2β KO mice displayed a reduction of B-cells, especially of the B220high IgMint-high recirculating population of transitional and follicular (FO) B cells. Pro-B and pre-B-cell progenitors were slightly reduced in number. In peripheral blood, lymph-nodes, spleen and peritoneal cavity the number of B-cells was markedly reduced. Csnk2β KO mice had lower levels of all the immunoglobulin classes in the serum. The splenic IgDlow IgMhigh B-cell subset was increased whereas the IgDhigh IgMint-low population was decreased. An imbalance between the amount of FO and marginal zone (MZ) B-cells was found with an absolute reduction of FO B cells by approximately 2-folds and an increase of MZ B-cells and MZB cell precursors by up to three folds, on average. Histological and immunofluorescence (IF) analysis revealed a change of size/shape of spleen follicles and a significant expansion of the inter-follicular, marginal zone areas, which appeared to invade the follicle with larger cells. In vitro class-switch recombination assays demonstrated impairment in IgG1 and IgG3 class-switch and a marked reduction of the generation of antibody-producing cells. Anti-IgM stimulation was uncoupled to Ca++ mobilization, indicating a disrupted transmission of the signal from the BCR to the release of Ca++ stores in the endoplasmic reticulum. In vivo sheep red blood cells (SRBC) treatment (T-cell dependent response) showed a conserved up-regulation of GC markers, such as CD38, GL7 and PNA. Nonetheless, the architecture of the reactive follicles was found markedly changed. The analysis of FO, GC and MZ-associated genes showed normal levels of Bcl6, elevated levels of Lrf mRNA and, more significantly, a marked up-regulation of Notch2 target genes, such as Hes1 and Deltex1, in Csnk2β KO B cells. In vivo Notch2 blockage with neutralizing antibodies markedly reduced the MZB cell number in Csnk2β KO mice, indicating a Notch2-dependent MZB expansion associated with Csnk2β loss. High throughput RNAseq analysis was also performed and revealed significant alteration in FOB and MZB-regulating pathways. Conclusions. Here, we found that the β subunit of protein kinase CK2 is a novel regulator of peripheral B cell differentiation. CK2β sustains a proper BCR signal, controls the GC reaction and negatively regulates Notch2 signaling, acting as a master regulator of follicular/marginal zone architecture and terminal homeostasis of FOB and MZB cells. On one side our data enrich the knowledge on the mechanisms regulating B cell development, on the other side they inform about the potential mechanisms altered by CK2 during B-cell tumorigenesis. Disclosures No relevant conflicts of interest to declare.

scholarly journals Protein kinase CK2� regulates peripheral B cell development

2013 ◽  
Vol 4 ◽  
Author(s):  
Zaffino Fortunato ◽  
Manni Sabrina ◽  
Mandato Elisa ◽  
Quotti Tubi Laura ◽  
Brancalion Alessandra ◽  
...  
Keyword(s):  
Protein Kinase ◽  
B Cell ◽  
Protein Kinase Ck2 ◽  
Cell Development ◽  
B Cell Development ◽  
Kinase Ck2

scholarly journals A critical role of Rap1b in B-cell trafficking and marginal zone B-cell development

Blood ◽  
2008 ◽  
Vol 111 (9) ◽  
pp. 4627-4636 ◽  
Author(s):  
Yuhong Chen ◽  
Mei Yu ◽  
Andrew Podd ◽  
Renren Wen ◽  
Magdalena Chrzanowska-Wodnicka ◽  
...  
Keyword(s):  
B Cells ◽  
Lymph Nodes ◽  
B Cell ◽  
Marginal Zone ◽  
Cell Development ◽  
B Cell Development ◽  
Cell Trafficking

Abstract B-cell development is orchestrated by complex signaling networks. Rap1 is a member of the Ras superfamily of small GTP-binding proteins and has 2 isoforms, Rap1a and Rap1b. Although Rap1 has been suggested to have an important role in a variety of cellular processes, no direct evidence demonstrates a role for Rap1 in B-cell biology. In this study, we found that Rap1b was the dominant isoform of Rap1 in B cells. We discovered that Rap1b deficiency in mice barely affected early development of B cells but markedly reduced marginal zone (MZ) B cells in the spleen and mature B cells in peripheral and mucosal lymph nodes. Rap1b-deficient B cells displayed normal survival and proliferation in vivo and in vitro. However, Rap1b-deficient B cells had impaired adhesion and reduced chemotaxis in vitro, and lessened homing to lymph nodes in vivo. Furthermore, we found that Rap1b deficiency had no marked effect on LPS-, BCR-, or SDF-1–induced activation of mitogen-activated protein kinases and AKT but clearly impaired SDF-1–mediated activation of Pyk-2, a key regulator of SDF-1–mediated B-cell migration. Thus, we have discovered a critical and distinct role of Rap1b in mature B-cell trafficking and development of MZ B cells.

scholarly journals Wiskott-Aldrich syndrome protein (WASP) and N-WASP are critical for peripheral B-cell development and function

Blood ◽  
2012 ◽  
Vol 119 (17) ◽  
pp. 3966-3974 ◽  
Author(s):  
Lisa S. Westerberg ◽  
Carin Dahlberg ◽  
Marisa Baptista ◽  
Christopher J. Moran ◽  
Cynthia Detre ◽  
...  
Keyword(s):  
T Cell ◽  
B Cells ◽  
B Cell ◽  
Marginal Zone ◽  
Cell Development ◽  
B Cell Development ◽  
Wild Type ◽  
And Function ◽  
Syndrome Protein

Abstract The Wiskott-Aldrich syndrome protein (WASP) is a key cytoskeletal regulator of hematopoietic cells. Although WASP-knockout (WKO) mice have aberrant B-cell cytoskeletal responses, B-cell development is relatively normal. We hypothesized that N-WASP, a ubiquitously expressed homolog of WASP, may serve some redundant functions with WASP in B cells. In the present study, we generated mice lacking WASP and N-WASP in B cells (conditional double knockout [cDKO] B cells) and show that cDKO mice had decreased numbers of follicular and marginal zone B cells in the spleen. Receptor-induced activation of cDKO B cells led to normal proliferation but a marked reduction of spreading compared with wild-type and WKO B cells. Whereas WKO B cells showed decreased migration in vitro and homing in vivo compared with wild-type cells, cDKO B cells showed an even more pronounced decrease in the migratory response in vivo. After injection of 2,4,6-trinitrophenol (TNP)–Ficoll, cDKO B cells had reduced antigen uptake in the splenic marginal zone. Despite high basal serum IgM, cDKO mice mounted a reduced immune response to the T cell–independent antigen TNP-Ficoll and to the T cell–dependent antigen TNP–keyhole limpet hemocyanin. Our results reveal that the combined activity of WASP and N-WASP is required for peripheral B-cell development and function.

Protein Kinase CK2 Regulates B Cell Development and Differentiation

2021 ◽  
pp. ji2100059
Author(s):  
Hairong Wei ◽  
Wei Yang ◽  
Huixian Hong ◽  
Zhaoqi Yan ◽  
Hongwei Qin ◽  
...  
Keyword(s):  
Protein Kinase ◽  
B Cell ◽  
Protein Kinase Ck2 ◽  
Cell Development ◽  
B Cell Development ◽  
Development And Differentiation ◽  
Kinase Ck2

scholarly journals Differential Regulation of Mouse B Cell Development by Transforming Growth Factor β1

2002 ◽  
Vol 9 (2) ◽  
pp. 86-95 ◽  
Author(s):  
Denise A. Kaminski ◽  
John J. Letterio ◽  
Peter D. Burrows
Keyword(s):  
B Cells ◽  
Growth Factor ◽  
B Cell ◽  
Transforming Growth Factor ◽  
Plasma Cells ◽  
Population Analysis ◽  
Cell Development ◽  
B Cell Development

Transforming growth factor β (TGFβ) can inhibit thein vitroproliferation, survival and differentiation of B cell progenitors, mature B lymphocytes and plasma cells. Here we demonstrate unexpected, age-dependent reductions in the bone marrow (BM) B cell progenitors and immature B cells in TGFβ1-/-mice. To evaluate TGFβ responsiveness during normal B lineage development, cells were cultured in interleukin 7 (IL7)±TGFβ. Picomolar doses of TGFβ1 reduced pro-B cell recoveries at every timepoint. By contrast, the pre-B cells were initially reduced in number, but subsequently increased compared to IL7 alone, resulting in a 4-fold increase in the growth rate for the pre-B cell population. Analysis of purified BM sub-populations indicated that pro-B cells and the earliest BP1-pre-B cells were sensitive to the inhibitory effects of TGFβ1. However, the large BP1+pre-B cells, although initially reduced, were increased in number at days 5 and 7 of culture. These results indicate that TGFβ1 is important for normal B cell developmentin vivo, and that B cell progenitors are differentially affected by the cytokine according to their stage of differentiation.

scholarly journals Ligand-independent Signaling Functions for the B Lymphocyte Antigen Receptor and Their Role in Positive Selection during B Lymphopoiesis

2001 ◽  
Vol 194 (11) ◽  
pp. 1583-1596 ◽  
Author(s):  
Gregory Bannish ◽  
Ezequiel M. Fuentes-Pananá ◽  
John C. Cambier ◽  
Warren S. Pear ◽  
John G. Monroe
Keyword(s):  
B Cells ◽  
B Cell ◽  
B Lymphocyte ◽  
Antigen Receptor ◽  
Cell Development ◽  
B Cell Development ◽  
B Lymphopoiesis ◽  
Biologically Relevant ◽  
Developmental Progression

Signal transduction through the B cell antigen receptor (BCR) is determined by a balance of positive and negative regulators. This balance is shifted by aggregation that results from binding to extracellular ligand. Aggregation of the BCR is necessary for eliciting negative selection or activation by BCR-expressing B cells. However, ligand-independent signaling through intermediate and mature forms of the BCR has been postulated to regulate B cell development and peripheral homeostasis. To address the importance of ligand-independent BCR signaling functions and their regulation during B cell development, we have designed a model that allows us to isolate the basal signaling functions of immunoglobulin (Ig)α/Igβ-containing BCR complexes from those that are dependent upon ligand-mediated aggregation. In vivo, we find that basal signaling is sufficient to facilitate pro-B → pre-B cell transition and to generate immature/mature peripheral B cells. The ability to generate basal signals and to drive developmental progression were both dependent on plasma membrane association of Igα/Igβ complexes and intact immunoregulatory tyrosine activation motifs (ITAM), thereby establishing a correlation between these processes. We believe that these studies are the first to directly demonstrate biologically relevant basal signaling through the BCR where the ability to interact with both conventional as well as nonconventional extracellular ligands is eliminated.

scholarly journals Identification of an Essential Cytoplasmic Region of Interleukin-7 Receptor α Subunit in B-Cell Development

2018 ◽  
Vol 19 (9) ◽  
pp. 2522 ◽  
Author(s):  
Hirotake Kasai ◽  
Taku Kuwabara ◽  
Yukihide Matsui ◽  
Koichi Nakajima ◽  
Motonari Kondo
Keyword(s):  
B Cells ◽  
B Cell ◽  
Myeloid Cell ◽  
Cell Development ◽  
B Cell Development ◽  
Α Subunit ◽  
Interleukin 7 ◽  
In Cells

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) α chain, here, we constructed a series of IL-7Rα deletion mutants. We found that IL-7Rα-deficient hematopoietic progenitor cells (HPCs) gave rise to B cells both in vitro and in vivo when a wild-type (WT) IL-7Rα chain was introduced; however, no B cells were observed under the same conditions from IL-7Rα-deficient HPCs with introduction of the exogenous IL-7Rα subunit, which lacked the amino acid region at positions 414–441 (d414–441 mutant). Signal transducer and activator of transcription 5 (STAT5) was phosphorylated in cells with the d414–441 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 d414–441 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 414–441 in the IL-7Rα chain formed a critical subdomain necessary for the supportive roles of IL-7 in B-cell development.

B Cell Lineage-Specific Deletion of Icsbp/IRF8 Reveals Roles for IRF8 in the Regulation of Marginal Zone and Follicular B Cell Development.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1334-1334
Author(s):  
Hongsheng Wang ◽  
Jianxun Feng ◽  
Chang Hoon Lee ◽  
Herbert Morse
Keyword(s):  
B Cells ◽  
B Cell ◽  
Marginal Zone ◽  
Cell Development ◽  
B Cell Development ◽  
Conditional Knockout ◽  
Myelogenous Leukemia ◽  
Exon 2 ◽  
B Lineage ◽  
B Lineage Cells

Abstract Interferon regulatory factor 8 (IRF8), also known as interferon consensus sequence-binding protein (ICSBP), is a transcription factor that expresses in T cells, B cells and macrophages and plays a role in myeloid development. Targeted deletion of IRF8 in mice (IRF8−/−) induced progressive increase in the numbers of granulocytes in various lymphoid organs and development of a syndrome similar to human chronic myelogenous leukemia. In addition to defective development of macrophages and dendritic cells, B cell development was also impaired in IRF8−/− mice. This includes decreased numbers of early B cells, expanded marginal zone (MZ) B cells and diminished follicular (OF) B2 cells. Because abnormal myeloid cells could alter microenvironment required for normal B cell development, we have generated IRF8 conditional knockout mice to specifically investigate the function of IRF8 in B lineage cells. Mice were engineered to have exon 2, encoding the DNA binding domain of IRF8, flanked by loxP sites (designated IRF8f/+). These mice were then crossed with the CD19Cre strain in which the expression of Cre-recombinase is controlled by the endogenous CD19 locus. Homozygous mice (designated (IRF8f/f x Cre)F1) underwent germline excision of IRF8 in CD19+ B lineage cells. As a result, there was no detectable mRNA and protein of IRF8 in their splenic B cells. Flow cytometry analysis revealed expanded MZ B cells and reduced OF B2 cells in the spleen of (IRF8f/f x Cre)F1 mice. Interestingly, the expression level of CD23 on OF B cells was significantly decreased in (IRF8f/f x Cre)F1 mice, indicating that IRF8 is required for maintaining a normal OF phenotype. In the peritoneum of (IRF8f/f x Cre)F1 mice, while the numbers of B1a and B2 cells were slightly decreased, the number of B1b cells was slightly increased. Furthermore, BXH2 mice carrying a mutation (C915T) in the Icsbp1 gene exhibited similar expansion of MZ B cells and low expression of CD23 in OF B cells. Taken together, these analyses indicate that IRF8 is required for development of normal MZ and B2 cells.

The Proto-Oncogene LRF Is Essential for Normal Mature B Cell Development and Germinal Center Formation.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1788-1788
Author(s):  
Nagisa Sakurai ◽  
Manami Maeda ◽  
Sung-UK Lee ◽  
Julie Teruya-Feldstein ◽  
Takahiro Maeda
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cell Development ◽  
B Cell Development ◽  
Cell Stage ◽  
Transitional B Cells ◽  
Notch Pathways ◽  
Secondary Lymphoid Organs

Abstract LRF (Leukemia/Lymphoma Related Factor, also known as Pokemon, FBI-1, OCZF and ZBTB7a) was originally identified as an interaction partner of the oncoprotein BCL6. LRF can act as a proto-oncogene by repressing the tumor suppressor ARF and cooperates with BCL6 in MEF (mouse embryonic fibroblasts) immortalization. It is highly expressed in human Non-Hodgkin Lymphoma (NHL) cases, in the pathogenesis of which BCL6 is known to be involved (Maeda et al. Nature 2005). Inducible inactivation of the LRF gene in mouse Hematopoietic Stem Cells (HSCs) results in complete block of early B cell development at the HSC/progenitor stages and concomitant development of double positive (DP) T cells in the bone marrow (BM) (Maeda et al. Science 2007). While these findings clearly illustrate key roles of LRF in normal and malignant B cell development, it is not fully identified as to which B cell stages LRF is required during normal B cell development. To elucidate the role of LRF in B cells in vivo, we established and characterized B cell-specific LRF conditional knockout (KO) mice. We took advantage of mb-1 Cre knock-in mice, in which Cre expression is restricted to the B cells after the ProB cell stage. B cell compartments in the BM (PreProB, ProB, PreB and immatureB) are grossly normal in LRFF/ Fmb1-Cre mice. The LRF gene was efficiently eliminated in BM CD19+ B cells revealed by quantitative real-time PCR assay. Furthermore, LRF protein was not detected in purified CD19+ B cells, but seen in CD19-non-B cells, confirming the specific inactivation of the LRF gene in B cells. Thus, despite its critical role at the HSC/progenitor stages, LRF was found to be dispensable for the survival of normal BM B cells. These findings are consistent with the fact that GSI treatment (Maeda et al. Science 2007) or Notch1 loss (Lee and Maeda, unpublished) rescues the defects in early B cell development seen in LRFF/FMx1-Cre+ mice. Notch signaling is necessary for the transitional B cells to commit to the marginal zone B cells (MZB). Inactivation of the component of the Notch pathways in mice results in no MZB development. On the contrary, deletion of the MINT/SHARP gene, a suppressor of Notch signaling, leads to increase of MZB cells and concomitant reduction of follicular B (FOB) cells, indicating that Notch induces MZB cell fate at the transitional B cell stage. Given that LRF is a potent Notch suppressor at the HSC/progenitor stages, we hypothesized that LRF opposes Notch pathway in mature B cells as well. To test this hypothesis, we characterized mature B cell development in LRFF/Fmb1-Cre mice. While transitional B cells were largely unaffected in LRFF/Fmb1-Cre mice, we observed a slight but statistically significant reduction of follicular (FO) B cells (B220+CD19+AA4.1-CD1d-CD23+) and concomitant increase of MZB cells (B220+CD19+AA4.1-CD1d+CD23-) as seen in MINT/SHARP knockout mice. Thus, LRF may also oppose Notch pathways at the branching point for the FOB vs. MZB fate decision. Finally, to determine the role of LRF in Germinal Center (GC) formation in vivo, we characterized secondary lymphoid organs of LRFF/Fmb1-Cre mice after antigen stimulation. Both spleen and Peyer’s Patches were analyzed two weeks after immunization with Chicken Gamma Globulin (NP-CGG). While a GC reaction was robustly induced in control mice upon immunization, GC formation was significantly impaired in LRFF/Fmb1-Cre mice as revealed by immuno-histochemical analysis (IHC) and FACS. Only few GC cells (B220+CD19+FAS+CD38-PNA+) were observed in spleens, and the absolute numbers of GC cells were drastically reduced in LRFF/Fmb1-Cre mice. Residual LRF-deficient GC B cells were mostly negative for CXCR4, which is predominantly expressed in proliferating centroblasts within GCs, suggesting that LRF-deficient GC B cells may have defects in cellular proliferation in response to antigen stimuli. Our data indicates that LRF plays key roles in mature B cell development in the secondary lymphoid organs, but dispensable for the maintenance of early BM B cells.

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