Differential Regulation of B Cell Development, Activation, and Death by the Src Homology 2 Domain–Containing 5′ Inositol Phosphatase (Ship) (original) (raw)
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Although the Src homology 2 domain-containing 5 Ј inositol phosphatase (SHIP) is a wellknown mediator of inhibitory signals after B cell antigen receptor (BCR) coaggregation with the low affinity Fc receptor, it is not known whether SHIP functions to inhibit signals after stimulation through the BCR alone. Here, we show using gene-ablated mice that SHIP is a crucial regulator of BCR-mediated signaling, B cell activation, and B cell development. We demonstrate a critical role for SHIP in termination of phosphatidylinositol 3,4,5-triphosphate (PI[3,4,5]P 3 ) signals that follow BCR aggregation. Consistent with enhanced PI(3,4,5)P 3 signaling, we find that splenic B cells from SHIP-deficient mice display enhanced sensitivity to BCR-mediated induction of the activation markers CD86 and CD69. We further demonstrate that SHIP regulates the rate of B cell development in the bone marrow and spleen, as B cell precursors from SHIP-deficient mice progress more rapidly through the immature and transitional developmental stages. Finally, we observe that SHIP-deficient B cells have increased resistance to BCR-mediated cell death. These results demonstrate a central role for SHIP in regulation of BCR signaling and B cell biology, from signal driven development in the bone marrow and spleen, to activation and death in the periphery. 1 Abbreviations used in this paper: 7AAD, 7-amino-actinomycin D; BCR, B cell antigen receptor; Btk, Bruton's tyrosine kinase; [Ca 2 ϩ ], intracellular free calcium; HSA, heat-stable antigen; IP 3 , inositol 1,4,5-triphosphate; MAP, mitogen-activated protein; mIg, membrane-bound Ig; NF, nuclear factor; PI3-K, phosphatidylinositol 3-kinase; PI(3,4)P 2 , phosphatidylinositol 3,4-biphosphate; PI(3,4,5)P 3 , phosphatidylinositol 3,4,5-triphosphate; PLC, phospholipase C; SHIP, Src homology 2 domain-containing 5 Ј inositol phosphatase; sIg, surface Ig.
2010
Ship is an Src homology 2 domain containing inositol polyphosphate 5-phosphatase which has been implicated as an important signaling molecule in hematopoietic cells. In B cells, Ship becomes associated with Fc ␥ receptor IIB (Fc ␥ RIIB), a low affinity receptor for the Fc portion of immunoglobulin (Ig)G, and is rapidly tyrosine phosphorylated upon B cell antigen receptor (BCR)-Fc ␥ RIIB coligation. The function of Ship in lymphocytes was investigated in Ship Ϫ / Ϫ recombination-activating gene (Rag) Ϫ / Ϫ chimeric mice generated from gene-targeted Ship Ϫ / Ϫ embryonic stem cells. Ship Ϫ / Ϫ Rag Ϫ / Ϫ chimeras showed reduced numbers of B cells and an overall increase in basal serum Ig. Ship Ϫ / Ϫ splenic B cells displayed prolonged Ca 2 ϩ influx, increased proliferation in vitro, and enhanced mitogen-activated protein kinase (MAPK) activation in response to BCR-Fc ␥ RIIB coligation. These results demonstrate that Ship plays an essential role in Fc ␥ RIIB-mediated inhibition of BCR signaling, and that Ship is a crucial negative regulator of Ca 2 ϩ flux and MAPK activation.
Journal of Experimental Medicine, 2000
In this report, we demonstrate that the Src homology 2 domain–containing inositol-5-phosphatase (SHIP) plays a critical role in regulating both B cell development and responsiveness to antigen stimulation. SHIP−/− mice exhibit a transplantable alteration in B lymphoid development that results in reduced numbers of precursor B (fraction C) and immature B cells in the bone marrow. In vitro, purified SHIP−/− B cells exhibit enhanced proliferation in response to B cell receptor stimulation in both the presence and absence of Fcγ receptor IIB coligation. This enhancement is associated with increased phosphorylation of both mitogen-activated protein kinase and Akt, as well as with increased survival and cell cycling. SHIP−/− mice manifest elevated serum immunoglobulin (Ig) levels and an exaggerated IgG response to the T cell–independent type 2 antigen trinitrophenyl Ficoll. However, only altered B cell development was apparent upon transplantation into nonobese diabetic–severe combined im...
Journal of autoimmunity, 2015
Many self-reactive B cells exist in the periphery in a rapidly reversible state of unresponsiveness referred to as anergy. Reversibility of anergy indicates that chronically occupied BCR must transduce non-durable regulatory signals that maintain unresponsiveness. Consistent with such a mechanism, studies of immunoglobulin transgenic, as well as naturally occurring polyclonal autoreactive B cells demonstrate activation of the inositol 5-phosphatase SHIP-1 in anergic cells, and low affinity chromatin autoantigen-reactive B cells have been shown to require expression of this phosphatase to maintain anergy. However, it has been reported that anergy of B cells recognizing high affinity soluble antigen may not require SHIP-1, and is instead mediated by upregulation of the inositol 3-phosphatase PTEN. To further explore this apparent difference in mechanism we analyzed the effect of B cell-targeted SHIP-1 deletion on immune tolerance of high affinity anti-HEL B cells in mice expressing so...
Immunity, 2011
Anergic B cells are characterized by impaired signaling and activation after aggregation of their antigen receptors (BCR). The molecular basis of this impairment is not understood. In studies reported here, Src homology-2 (SH2)-containing inositol 5-phosphatase SHIP-1 and its adaptor Dok-1 were found to be constitutively phosphorylated in anergic B cells, and activation of this inhibitory circuit was dependent on Src-family kinase activity and consequent to biased BCR immunoreceptor tyrosine-based activation motif (ITAM) monophosphorylation. B cell-targeted deletion of SHIP-1 caused severe lupus-like disease. Moreover, absence of SHIP-1 in B cells led to loss of anergy as indicated by restoration of BCR signaling, loss of anergic surface phenotype, and production of autoantibodies. Thus, chronic BCR signals maintain anergy in part via ITAM monophosphorylation-directed activation of an inhibitory signaling circuit involving SHIP-1 and Dok-1. Immunity SHIP-1 Is Required for B Cell Anergy
Immunology Letters, 2006
FcgRIIB are single-chain low-affinity receptors for IgG that bear an immunoreceptor tyrosine-based inhibition motif (ITIM) in their intracytoplasmic domain and that negatively regulate immunoreceptor tyrosine-based activation motif (ITAM)-dependent cell activation. In B cells, coaggregation of the B cell receptor (BCR) and FcgRIIB leads to an inhibition of B cell activation. Inhibitory properties of FcgRIIB have been related to the recruitment of SHIP, an SH2 domain-containing inositol 5-phosphatase (referred to as SHIP1), via ITIM phosphorylated FcgRIIB. Here, we demonstrate that the second SH2 domain-containing inositol 5-phosphatase SHIP2 could also bind to the FcgRIIB ITIM. As a model, a FcgRIIB deficient B cell line (IIA1.6), transfected with a cDNA encoding either w.t. FcgRIIB1% or FcgRIIB1% whose ITIM tyrosine was mutated has been used. SHIP2 tyrosine phosphorylation and association to the adaptator protein Shc were only found in transfectants expressing w.t. FcgRIIB1%. SHIP2 was also found to bind to a phosphopeptide corresponding to the ITIM sequence of FcgRIIB. There was no binding to the nonphosphorylated peptide. Finally, both SHIP2 and SHIP1 were coprecipitated with FcgRIIB1% upon coaggregation with BCR in IIA1.6 transfectants.
Journal of Biological Chemistry, 1999
Src homology 2 domain-containing inositol 5-phosphatase (SHIP) mediates inhibitory signals that attenuate intracellular Ca 2؉ mobilization in B cells upon B cell receptor (BCR) stimulation. To clarify the mechanisms affected by SHIP, we analyzed Ca 2؉ mobilization in the DT40 B cell line in which the SHIP gene was disrupted. In SHIP-deficient cells, Ca 2؉ transient elicited by BCR stimulation was more prolonged than that in control cells both in the presence and absence of extracellular Ca 2؉. Inositol 1,4,5-trisphosphate production following BCR stimulation was enhanced in SHIP-deficient cells. In SHIP-deficient cells in comparison with the control cells, BCR stimulation in the absence of extracellular Ca 2؉ induced a greater degree of Ca 2؉ store depletion and the Ca 2؉ influx upon re-addition of extracellular Ca 2؉ was also greater. However, store-operated Ca 2؉ influx (SOC) elicited by thapsigargin-induced store depletion was not affected by SHIP. These results indicate that the primary target pathway of SHIP is the Ca 2؉ release from the stores, and that Ca 2؉ influx by the SOC mechanism is secondarily controlled by the level of Ca 2؉ in the stores without direct inhibition of SOC. In this way, SHIP may play an important role in ensuring the robust tuning of Ca 2؉ signaling in B cells.
Immunology Letters, 1999
Co-clustering B cell receptors (BCR) and type II receptors binding the Fc part of IgG (FcgRIIb) inhibits B cell activation and antibody production. Tyrosine phosphorylation of an intracellular motif of FcgRIIb has been shown to be a prerequisite of the inhibition. After being phosphorylated by BCR-activated tyrosine kinases, the immunoreceptor tyrosine-based inhibitory motif (P-ITIM) of FcgRIIb recruits SH2 domain containing protein tyrosine phosphatase(s) (PTPs) and polyphosphoinositol 5-phosphatase (SHIP) to the vicinity of BCR, which in turn dephosphorylate their specific substrates. This leads to the interruption of signal transduction, consequently to the anergy and/or apoptosis of the cell. The downstream signaling pathways affected by FcgRIIb-BCR co-clustering are not clarified yet, neither the substrates of PTPs are known. We have studied the FcgRIIb mediated B cell inhibition on human Burkitt lymphoma cell line (BL41). From the lysates of BL41 cells SHP-2 and phosphatidylinositol 3-kinase (PI3-K), as well as the protein tyrosine kinase (PTK) Lyn bind both to the BCR-co-clustered FcgRIIb and to its P-ITIM peptide. Lyn hyperphosphorylates the P-ITIM associated molecules, including SHIP in the in vitro protein tyrosine kinase activity assay. The P-ITIM-compelled multi-phosphoprotein complex binds to and activates SHP-2, which in turn dephosphorylates SHIP and Shc and probably other substrates. Subcellular localisation of these signaling molecules is regulated by the phosphotyrosin-SH2 domain interactions, thus dephosphorylation may result in the re-direction of Shc and SHIP within the cell, consequently, in the modulation of their activity. Finally, co-clustering FcgRIIb and BCR or FcgRIIb and CD19 on the intact cells inhibited PI3-K activity as detected in the anti-phosphotyrosine (anti-PY) precipitates. The results indicate that SHP-2 bound to and activated by the BCR co-clustered FcgRIIb, may down-regulate PI3-K activity by dephosphorylating a yet unidentified regulatory molecule, which recruits PI3-K to the cell membrane.
Molecular and cellular biology, 1997
Recently, we and others have demonstrated that negative signaling in B cells selectively induces the tyrosine phosphorylation of a novel inositol polyphosphate phosphatase, p145SHIP. In this study, we present data indicating that p145SHIP binds directly a phosphorylated motif, immunoreceptor tyrosine-based inhibition motif (ITIM), present in the cytoplasmic domain of Fc gammaRIIB1. Using recombinant SH2 domains, we show that binding is mediated via the Src homology region 2 (SH2)-containing inositol phosphatase (SHIP) SH2 domain. SHIP also bound to a phosphopeptide derived from CD22, raising the possibility that SHIP contributes to negative signaling by this receptor as well as Fc gammaRIIB1. The association of SHIP with the ITIM phosphopeptide was activation independent, while coassociation with Shc was activation dependent. Furthermore, experiments with Fc gammaRIIB1-deficient B cells demonstrated a genetic requirement for expression of Fc gammaRIIB1 in the induction of SHIP phosp...