Requirement for B Cell Linker Protein (BLNK) in B Cell Development

Pappu, Rajita; Cheng, Alec M.; Li, Bin; Gong, Qian; Chiu, Christopher; Griffin, Nancy; White, Michael A.; Sleckman, Barry P.; Chan, Andrew C.

doi:10.1126/science.286.5446.1949

Cited by 276 publications

(208 citation statements)

References 37 publications

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“…There are several genes whose level of expression in wild-type vs PU.1/Spi-B low/mutant cell lines is not predicted according to the hypothesis described above. BLNK (SLP-65) is an adaptor protein required for signaling through the BCR and for normal B cell development (58,59). There is no suggestion in the literature that BLNK transcription should be down-regulated during the pro-B to pre-B cell transition, but we found that BLNK transcription was reduced in PU.1/Spi-B low/mutant cells by an average of 3-fold.…”

Section: Discussionmentioning

confidence: 55%

Analysis of Gene Expression and Ig Transcription in PU.1/Spi-B-Deficient Progenitor B Cell Lines

Schweitzer

DeKoter

2004

The Journal of Immunology

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A number of presumptive target genes for the Ets-family transcription factor PU.1 have been identified in the B cell lineage. However, the precise function of PU.1 in B cells has not been studied because targeted null mutation of the PU.1 gene results in a block to lymphomyeloid development at an early developmental stage. In this study, we take advantage of recently developed PU.1−/−Spi-B−/− IL-7 and stromal cell-dependent progenitor B (pro-B) cell lines to analyze the function of PU.1 and Spi-B in B cell development. We show that contrary to previously published expectations, PU.1 and/or Spi-B are not required for Ig H chain (IgH) gene transcription in pro-B cells. In fact, PU.1−/−Spi-B−/− pro-B cells have increased levels of IgH transcription compared with wild-type pro-B cells. In addition, high levels of Igκ transcription are induced after IL-7 withdrawal of wild-type or PU.1−/−Spi-B−/− pro-B cells. In contrast, we found that Igλ transcription is reduced in PU.1−/−Spi-B−/− pro-B cells relative to wild-type pro-B cells after IL-7 withdrawal. These results suggest that Igλ, but not IgH or Igκ, transcription, is dependent on PU.1 and/or Spi-B. The PU.1−/−Spi-B−/− pro-B cells have other phenotypic changes relative to wild-type pro-B cells including increased proliferation, increased CD25 expression, decreased c-Kit expression, and decreased RAG-1 expression. Taken together, our observations suggest that reduction of PU.1 and/or Spi-B activity in pro-B cells promotes their differentiation to a stage intermediate between late pro-B cells and large pre-B cells.

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Section: Discussionmentioning

confidence: 55%

Analysis of Gene Expression and Ig Transcription in PU.1/Spi-B-Deficient Progenitor B Cell Lines

Schweitzer

DeKoter

2004

The Journal of Immunology

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“…Btk -/- [10], BLNK -/- [13], and PLCc2 -/- [17] mice on a mixed C57BL/6 Â 129 background were mated to generate Btk -/-PLCc2 -/-, BLNK -/-Btk -/-, and BLNK -/-PLCc2 -/-mice. Mice were genotyped by PCR and sacrificed if they developed signs of illness such as scruffy fur, hunched posture, lack of movement, hind limb paralysis, or visible tumors.…”

Section: Micementioning

confidence: 99%

“…A similar disease results from mutation of the human BLNK gene [9]. Mice lacking Btk [10][11][12], BLNK [13][14][15][16], or PLCc2 [17,18] have a milder phenotype with the predominant defect at the T2 to mature transition in the periphery. Btk -/-mice have a reduced frequency of Igk-expressing cells due to a role for Btk in Igk rearrangement in pre-B cells [19].…”

Section: Introductionmentioning

confidence: 99%

Btk and phospholipase Cγ2 can function independently during B cell development

et al. 2007

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The pre-BCR and the BCR regulate B cell development via a signalosome nucleated by the adaptor protein B cell linker protein (BLNK). Formation of this complex facilitates activation of phospholipase C (PLC) c2 by Bruton's tyrosine kinase (Btk). To determine whether Btk and PLCc2 also have separate functions, we generated Btk -/-PLCc2 -/-mice. They demonstrated a block in development at the pre-B stage and increased pre-BCR surface expression. This phenotype was more severe than that of Btk -/-or PLCc2 -/-mice. Although both Btk and PLCc2 were required for proliferation of splenic B cells in response to BCR cross-linking, they contributed differently to anti-IgM-induced phosphorylation of ERK. Btk -/-and PLCc2 -/-mice each had a reduced frequency of Igk-expressing B cells and impaired migration of pre-B cells towards stromal cellderived factor 1. However, the increase in pre-B cell malignancy that occurs in BLNK -/-mice in the absence of Btk was not observed in the absence of PLCc2. Thus, Btk and PLCc2 act both in concert and independently throughout B cell development. IntroductionSignals from the B cell antigen receptor (BCR) regulate multiple stages of B lymphopoiesis [1]. In pre-B cells, the pre-BCR signals for proliferation, allelic exclusion of the IgH locus, down-regulation of VpreB and k5 expression, and light chain rearrangement. The pre-BCR is predominantly intracellular and is believed to signal in a ligand-independent manner. The BCR is first expressed on the surface of immature B cells. Antigen encounter at this stage results in negative selection via deletion, anergy, or receptor editing. A tonic BCR signal is required for differentiation beyond the T2 stage in the periphery and survival of mature B cells. Mature B cells proliferate and differentiate upon stimulation with foreign antigen in the appropriate context. BCR and pre-BCR signaling is mediated by a signalosome composed of Syk, Bruton's tyrosine kinase (Btk), B cell linker protein (BLNK; also known as SLP65 and BASH), and phospholipase C (PLC) c2 [2-4]. Receptor cross-linking activates Syk, which phosphorylates the adaptor protein BLNK. Btk and PLCc2 bind to phosphorylated BLNK via their SH2 domains. Btk then phosphorylates and activates PLCc2, resulting in changes in intracellular Ca 2+ and the activation of protein kinase Cb. Btk can also mediate BCR-stimulated Ca 2+ flux [5] and B cell development [6] independently of its catalytic activity. Btk acts as an adaptor to recruit and activate phosphatidylinositol 4-phosphate 5-kinase (PIP5K) [5], which produces the substrate for PLCc2. Thus, Btk signals through PLCc2 directly, by phosphorylation, and indirectly, by increasing substrate availability.Loss of Btk function causes the human immunodeficiency X-linked agammaglobulinemia (XLA) [7,8], which is characterized by a block in B cell development at the pre-B stage. A similar disease results from

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“…These interactions are essential for the assembly of the signaling cascade ultimately leading to the release of Ca 2 þ from the endoplasmic reticulum (Fu et al, 1998). In the absence of SLP65, B cell development is arrested at the pro-B to pre-B cell transition (Minegishi et al, 1999;Pappu et al, 1999), which parallels complete breakdown of (pre-) B cell receptor signaling in SLP65-deficient B cells (Ishiai et al, 1999;Pappu et al, 1999;Hayashi et al, 2003). While one study reported normal expression of SLP65 in childhood acute lymphoblastic leukemia (Imai et al, 2004), recent work by us and others Hayashi et al, 2003;Jumaa et al, 2003;Kersseboom et al, 2003;Klein et al, 2004) indicated a role of SLP65 as a tumor suppressor in human and murine leukemia derived from pre-B cells.…”

Section: Introductionmentioning

confidence: 99%

The SRC family kinase LYN redirects B cell receptor signaling in human SLP65-deficient B cell lymphoma cells

et al. 2006

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SLP65 represents a critical component in (pre-) B cell receptor signal transduction but is compromised in a subset of pre-B cell-derived acute lymphoblastic leukemia. Based on these findings, we investigated (i.) whether SLP65-deficiency also occurs in mature B cell-derived lymphoma and (ii.) whether SLP65-deficient B cell lymphoma cells use an alternative B cell receptor signaling pathway in the absence of SLP65. Indeed, expression of SLP65 protein was also missing in a fraction of B cell lymphoma cases. While SLP65 is essential for B cell receptor-induced Ca 2 þ mobilization in normal B cells, B cell receptor engagement in SLP65-deficient as compared to SLP65-reconstituted B cell lymphoma cells resulted in an accelerated yet shortlived Ca 2 þ -signal. B cell receptor engagement of SLP65-deficient lymphoma cells involves SRC kinase activation, which is critical for B cell receptor-dependent Ca 2 þ -mobilisation in the absence but not in the presence of SLP65. As shown by RNA interference, the SRC kinase LYN is required for B cell receptor-induced Ca 2 þ release in SLP65-deficient B cell lymphoma cells but dispensable after SLP65-reconstitution. B cell receptor engagement in SLP65-deficient B cell lymphoma cells also resulted in tyrosine-phosphorylation of the proliferation-and survival-related MAPK1 and STAT5 molecules, which was sensitive to silencing of the SRC kinase LYN. Inhibition of SRC kinase activity resulted in growth arrest and cell death specifically in SLP65-deficient lymphoma cells. These findings indicate that LYN can short-circuit conventional B cell receptor signaling in SLP65-deficient B cell lymphoma cells and thereby promote activation of survival and proliferationrelated molecules.

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Requirement for B Cell Linker Protein (BLNK) in B Cell Development

Cited by 276 publications

References 37 publications

Analysis of Gene Expression and Ig Transcription in PU.1/Spi-B-Deficient Progenitor B Cell Lines

Analysis of Gene Expression and Ig Transcription in PU.1/Spi-B-Deficient Progenitor B Cell Lines

Btk and phospholipase Cγ2 can function independently during B cell development

The SRC family kinase LYN redirects B cell receptor signaling in human SLP65-deficient B cell lymphoma cells

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