HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Goitsuka, R. Articles by Kitamura, D. Search for Related Content PubMed PubMed Citation Articles by Goitsuka, R. Articles by Kitamura, D.

Cutting Edge: BASH, A Novel Signaling Molecule Preferentially Expressed in B Cells of the Bursa of Fabricius¹

Ryo Goitsuka^2,*,, Yu-ichi Fujimura, Hiroshi Mamada, Akiko Umeda, Toshifumi Morimura, Koji Uetsuka, Kunio Doi, Sachiyo Tsuji and Daisuke Kitamura

^* Inheritance and Variation Group, PREST, JST, Kyoto, Japan; Division of Molecular Biology, Research Institute for Biological Sciences, Science University of Tokyo, Chiba, Japan; and Department of Veterinary Pathology, University of Tokyo, Tokyo, Japan

	Abstract

Top Abstract Introduction Materials and Methods Results and Discussion References

 
The bursa of Fabricius is a gut-associated lymphoid organ that is essential for the generation of a diversified B cell repertoire in the chicken. We describe here a novel gene preferentially expressed in bursal B cells. The gene encodes an 85-kDa protein, designated BASH (B cell adaptor containing SH2 domain), that contains N-terminal acidic domains with SH2 domain-binding phosphotyrosine-based motifs, a proline-rich domain, and a C-terminal SH2 domain. BASH shows a substantial sequence similarity to SLP-76, an adaptor protein functioning in TCR-signal transduction. BASH becomes tyrosine-phosphorylated with the B cell Ag receptor (BCR) cross-link or by coexpression with Syk and Lyn and associates with signaling molecules including Syk and a putative chicken Shc homologue. Overexpression of BASH results in suppression of the NF-AT activation induced by BCR-cross-linking. These findings suggest that BASH is involved in BCR-mediated signal transduction and could play a critical role in B cell development in the bursa.

	Introduction

Top Abstract Introduction Materials and Methods Results and Discussion References

 
The avian bursa of Fabricius is an outgrowth of the cloaca that is essential for the generation of a peripheral B cell pool with diversified Ag receptors (1, 2). Within the bursa, a small number of committed progenitors extensively proliferate and produce diversified B cell progeny through a somatic Ig gene conversion mechanism. The majority of these B cells undergo programmed cell death in the bursa and only 1–5% of the cells survive to emigrate into the periphery (3, 4). As has been demonstrated in the mouse (5), the B cell Ag receptor (BCR)³ may be required for the survival and maturation of B cells in the bursa, because it has been reported that loss of surface Ig precedes the induction of apoptosis in rapidly dividing bursal B cells (6). This finding suggests that continuous expression of surface BCR or the signals thereof might be required for the survival of bursal B cells. Consistent with a role in activation and proliferation of bursal B cells, the BCR has been shown to be capable of transducing signals, such as rapid induction of calcium influx and phospholipid hydrolysis upon cross-linking (7).

Cross-linking of the BCR induces a cascade of biochemical events mediated by transient phosphorylations and interactions of various signaling proteins, including protein tyrosine kinases, phosphatases, and adaptor molecules (8). However, it is still unclear how these intracellular biochemical events are regulated to induce different cellular responses, e.g., proliferation or apoptosis. Particularly, BCR-mediated signal transduction mechanisms in bursal B cells have been poorly characterized.

To better understand mechanisms for the induction and regulation of B cell proliferation, diversification, death, or survival in the bursa, we adopted a strategy of identifying proteins selectively expressed in bursal B cells. Here we describe the molecular cloning of one such protein, BASH, a B cell adaptor containing an SH2 domain, and provide evidence suggesting that BASH is a signaling component downstream of BCR-associated tyrosine kinases.

	Materials and Methods

Top Abstract Introduction Materials and Methods Results and Discussion References

 
Antibodies

The anti-chicken IgM Ab, M4, was provided from Dr. C.-L. H. Chen (University of Alabama at Birmingham, AL). The anti-porcine Syk polyclonal antiserum, which was shown to cross-react with chicken Syk (9), was a gift from Dr. T. Kurosaki (Kansai Medical University, Osaka, Japan). The anti-T7 epitope Ab, anti-phosphotyrosine (pY) Ab (RC20), and polyclonal anti-Shc Ab directed against the SH2 domain of human Shc were purchased from Novagen (Madison, WI), Signal Transduction Laboratories (Lexington, KY), and Upstate Biotechnology (Lake Placid, NY), respectively.

Isolation of chicken BASH cDNA and construction of an expression plasmid

The cDNA from chicken bursal B cells was subtracted by the cDNA from spleen lymphocytes using the PCR-select cDNA subtraction kit (Clontech, Palo Alto, CA), and the subtracted PCR products were cloned into pCR2.1 vector (Invitrogen, San Diego, CA). To further screen differentially expressed products, the subtracted fragments were run on an agarose gel and immobilized on duplicate nylon membrane filters, and then hybridized to cDNA probes derived from either a DT40 B cell line or a 132B T cell line poly(A)⁺ RNA. One of the subtracted cDNA fragments hybridized with the DT40 cell cDNA probe but not to the 132B cell cDNA probe was used to screen a DT40 cDNA library constructed in ZAP-XR (Stratagene, La Jolla, CA). The positive clones were subjected to sequencing by the dideoxy-chain termination method with an automatic DNA sequencer (Applied Biosystems, Foster City, CA). A full-length cDNA encoding the short-form of BASH was ligated in-frame with N-terminal double T7 epitope tag in pAT7neo expression vector, which was driven by the chicken ß-actin promoter (pAT7neo-BASH). Details of restriction maps and cloning sites of the construct are available upon request.

Northern blot and in situ hybridization analysis

Total RNA samples (10 µg) extracted from tissues and cell lines were electrophoresed in 1.2 M formaldehyde/1.2% agarose gel and transferred to nylon membranes. The blots were hybridized with a ³²P-labeled 900-bp chicken BASH cDNA fragment initially isolated by the subtraction. Whole-mount in situ hybridization was performed essentially as described (10). Digoxigenin-labeled sense and anti-sense 900-bp BASH riboprobes were produced with the Digoxigenin RNA Labeling Kit (Boehringer Mannheim, Mannheim, Germany) according to the manufacturer’s instruction. After whole-mount in situ hybridization, tissues were embedded in Tissue-Tek (Miles, Elkhart, IN), frozen in liquid nitrogen, and sectioned at 10 µm on a Leiz cryostat. Sections were mounted with Mowiol, and photographed on a Olympass BX30 Nomarwsky microscope (Tokyo, Japan).

Immunoprecipitations and immunoblot analysis

DT40 cells were electroporated with pAT7neo-BASH and then stable clones expressing T7-tagged BASH protein were selected by G418 (2 mg/ml). These clones were stimulated with the anti-chicken IgM Ab (15 µg/ml) at 40°C for the indicated period of time and then lysed at 2 x 10⁷ cells/ml in the lysis buffer containing 1% NP40, 10 mM Tris, pH 7.8, 150 mM NaCl, 2 mM EDTA, and protease and phosphatase inhibitors. The lysates were immunoprecipitated with the indicated Abs and then analyzed by Western blotting with the indicated Abs and secondary Abs conjugated with horseradish peroxidase. Immunoreactive proteins were detected by ECL kit (Amersham, Arlington Heights, IL). COS7 cells (5 x 10⁵) were cotransfected with 1 µg of pAT7neo-BASH with either 1 µg of pME-Lyn, pME-Syk (gifts from Dr. H. Nishizumi, Institute of Medical Science, University of Tokyo, Tokyo, Japan), or both plasmids using a TransIT-LT1 transfection reagent (Pan Vera, Masison, WI). After the incubation for 48 h, cell lysates were prepared and analyzed for tyrosine phosphorylation as described above.

Luciferase assay

DT40 cells were cotransfected with 10 µg of a luciferase reporter plasmid driven by seven tandem copies of the NF of activated T cells (NF-AT) response element from the mouse IL-2 gene promoter (NF-AT-Luc; a gift from Dr. K. Arai, Institute of Medical Science, University of Tokyo), together with 15 µg of either empty pAT7neo or pAT7neo-BASH in serum-free RPMI 1640 at a density of 10⁷ cells/400 µl per cuvette with a gene pulser (Bio-Rad Laboratories, Richmond, CA) set at 250 V and 975 µF. After electroporation, the cells were transferred to complete RPMI 1640 and incubated at 40°C for 48 h. Triplicates of 5 x 10⁵ viable cells were then stimulated with anti-IgM Ab and subsequently assayed for luciferase activity, as described previously (11). Light emission was measured in a Lumat LB9501 luminometer (Berthold, Wildbad, Germany).

	Results and Discussion

Top Abstract Introduction Materials and Methods Results and Discussion References

 
Isolation of a cDNA fragment whose mRNA is preferentially expressed in B cells of the bursa of Fabricius

We sought to clarify the molecular basis of the unique bursal differentiation process by identifying genes selectively expressed in bursal B cells through PCR-based subtraction combined with differential hybridization. One positive cDNA clone, designated B-1-2, hybridized to a 2.5-kb mRNA transcript that was abundantly expressed in the bursa, not in thymus, bone marrow, or other tissues, except for a low level of expression in spleen and ovary (Fig. 1A). All chicken B cell lines, but no T cell lines expressed this mRNA transcript, although the level of the expression was highly variable among B cell lines. High levels of transcripts were detectable in immature B cell lines such as DT40 and CL18, in which gene conversion persists (12), and TLT-1, whose Ig light chain gene is in germline configuration. In contrast, very low levels of transcripts were detected in relatively mature B cell lines, such as 293B9 and 249L4 (Fig. 1B). In situ hybridization analysis revealed a strong positive signal in B cell follicles in the bursa (Fig. 1C), especially in medullar B cells surrounded by the follicular epithelium (Fig. 1D).

View larger version (72K): [in this window] [in a new window]   FIGURE 1. The expression pattern of BASH mRNA. A, Northern blot analysis of total RNA from chicken tissues. The blots were hybridized with the B-1-2 cDNA probe (top) and reprobed with the chicken ß-actin cDNA (bottom). B, Northern blot analysis of total RNA from chicken cell lines. B cell lines are CL-18, DT40, TLT1, 293 B9, and 249L4; T cell lines are RP1, JP2, and MSB1. C, Whole-mount in situ hybridization analysis of the bursa of Fabricius, showing that B-1-2 transcripts are localized at B cell follicles. D, Transverse section of the sample in C.

Using a B-1-2 cDNA fragment as a probe, we isolated two full-length cDNA clones with different lengths (GenBank/EMBL/DDBJ accession number AB015289). The long-form cDNA contains the longest open reading frame encoding a 553-amino acid protein, with a calculated molecular mass of 62 kDa (Fig. 2). The shorter form of cDNA lacks 57 nucleotides corresponding to the 19 amino acids located at the N terminus of the long-form protein. These two forms of mRNA transcripts were detectable in the bursa as well as DT40 cells by PCR using a pair of primers flanking the deleted portion (data not shown), although we could not assess the corresponding protein species expressed in B cells due to the lack of a discriminating Ab. The two predicted proteins contain N-terminal acidic domains with seven potential phosphotyrosine-based binding motifs for SH2 domains (13, 14) and a putative SH2 domain in the C-terminal region. The intervening region between the two domains is rich in proline residues and may represent binding sites for proteins containing an SH3 domain (15). Based on these molecular characteristics, this protein was termed BASH (B cell-specific adaptor containing an SH2 domain).

View larger version (95K): [in this window] [in a new window]   FIGURE 2. Deduced amino acid sequences of chicken BASH (B), aligned with mouse SLP-76 (S). Identical and similar amino acids are indicated by black and gray shading, respectively. Dashes denote gaps introduced to optimize similarity. The numbers in the left column indicate the position of amino acid residues. The amino acid residues missing in the short-form of BASH are indicated by a dashed line above the sequence. Tyrosine residues in the potential SH2 domain-binding motifs are shown by closed circles above the sequence. The SH2 domain is underlined.

BASH is tyrosine-phosphorylated after cross-linking of surface-IgM on B cells

Because SLP-76 is known to be tyrosine-phosphorylated upon TCR-cross-linking (16), we examined whether BASH is phosphorylated upon BCR stimulation. The T7-epitope tagged BASH cDNA was transfected into DT40 cells, and then the BASH protein was analyzed by immunoprecipitation with anti-tag Ab. The BASH protein was found to migrate on SDS-PAGE with an apparent molecular mass of 85 kDa, which is larger than the predicted molecular mass of 60 kDa plus the additional 4 kDa of the T7-epitope tag. The anomalous migration may result from posttranslational modification or the abundance of charged amino acids present in the protein. Maximal BCR-induced tyrosine phosphorylation of BASH was detectable after 1 min and was sustained for at least 15 min after BCR stimulation (Fig. 3A). In addition, we found that lysates of the stimulated DT40 cells contained several additional tyrosine-phosphorylated proteins (70 kDa, 52 kDa, and 46 kDa), which were specifically coprecipitated with BASH.

View larger version (43K): [in this window] [in a new window]   FIGURE 3. BASH is tyrosine-phosphorylated after BCR stimulation and associates with Shc and Syk. A, Tyrosine phosphorylation of BASH. DT40 cells stably transfected with a T7-tagged BASH cDNA (DT40/BASH) were stimulated with anti-IgM Ab for the indicated periods of time. Whole-cell lysates were immunoprecipitated with an anti-T7 Ab, immunoblotted with an anti-phosphotyrosine Ab (top) and reprobed with an anti-T7 Ab (bottom). B, Coimmunoprecipitation of Shc with BASH. Lysates of DT40/BASH cells, either unstimulated or stimulated with anti-IgM mAb for 1 min, were immunoprecipitated with an anti-T7 Ab and then probed with anti-phosphotyrosine Ab (top) or anti-Shc Ab (bottom). C, Coimmunoprecipitation of Syk with BASH. Immunoprecipitates of unstimulated and stimulated (anti-IgM, 1 min) DT40/BASH cells with anti-T7 (bottom) or anti-Syk (top) Abs were reciprocally probed with the same Abs. D, Lysates from COS7 cells transfected with expression vectors for T7-tagged BASH, Lyn, or Syk in the indicated combinations were immunoprecipitated with anti-T7 Ab and immunoblotted with an anti-phosphotyrosine Ab (top) or an anti-T7 Ab (bottom).

By blotting the BASH immunoprecipitate with Abs against known tyrosine-phosphorylated proteins, we found that the 52-kDa BASH-associated phosphoprotein reacted with an anti-human Shc Ab, suggesting an association of BASH with the chicken Shc homologue. Although this association was seen before stimulation, it was significantly enhanced after BCR cross-linking (Fig. 3B). The 70-kDa phosphoprotein bound to BASH was identified as Syk; BASH was detectable in the anti-Syk immunoprecipitate, and Syk was coimmunoprecipitated with BASH. The BASH-Syk association was enhanced by BCR-stimulation (Fig. 3C).

Both Syk and Lyn are required for the maximal phosphorylation of BASH

To identify the tyrosine kinases responsible for the phosphorylation of BASH following BCR cross-linking, COS7 cells were cotransfected with a BASH plasmid and plasmids encoding Syk and Lyn. Anti-phosphotyrosine immunoblots of the BASH-immunoprecipitate from these cell lysates revealed that coexpression of either Syk or Lyn induced only a very weak tyrosine phosphorylation of BASH (Fig. 3D). In contrast, coexpression of Syk and Lyn dramatically increased the tyrosine phosphorylation of BASH, indicating that both kinases are required for maximal activity.

Overexpression of BASH interferes with NF-AT activation upon BCR-stimulation

Because SLP-76, a potential T-cell counterpart of BASH, has been demonstrated to augment TCR signals leading to IL-2 promoter and NF-AT activation (17, 18), we examined whether overexpression of BASH influences the activation of NF-AT, which is also involved in BCR-mediated transcriptional events (19). In contrast to the positive effect of SLP-76 on NF-AT activation in T cells, overexpression of BASH resulted in significant suppression of NF-AT activation when compared with transfection of the control vector (Fig. 4). This finding suggests that BASH may play an inhibitory role in a BCR-mediated signaling pathway leading to the activation of NF-AT.

View larger version (24K): [in this window] [in a new window]   FIGURE 4. Overexpession of BASH inhibits BCR-mediated NF-AT activation. DT40 cells were cotransfected with NF-AT-Luc plasmid and either the empty vector or a BASH expression vector. After 48 h, cells were either left unstimulated or stimulated with anti-IgM Ab for 6 h and subsequently assayed for luciferase activity. The results are shown as the fold induction of luciferase activity as compared with the activity in unstimulated cells transfected with empty plasmid. Luciferase activity was determined in triplicate in the experiment.

Very recently, novel mammalian B cell-specific adaptor proteins, BLNK (20) and SLP-65 (21), have been described. These proteins are very homologous to BASH, particularly in the N-terminal and SH2 domains, although the length and homology of the intervening region have diverged considerably between BLNK/SLP-65 and BASH. In contrast to BASH, BLNK does not associate with Shc and augments NF-AT activation following BCR-stimulation (20), suggesting that BASH may not simply be a homologue of BLNK/SLP-65. Moreover, the expression patterns of the proteins are quite different. BASH is found primarily in immature B cell lines and is expressed only weakly in mature B cells, while the converse is true of BLNK/SLP-65. Therefore, BASH may have evolved to fulfill unique functions in developing B cells in the avian bursa.

	Acknowledgments

 
We thank Dr. H. Nishizumi for providing pME-Lyn and pME-Syk plasmids; Dr. K. Arai for a gift of NF-AT luciferase plasmid; Dr. T. Kurosaki for anti-Syk antisera; Dr. C.-L. H. Chen for M4 mAb; and Dr. F. P. Zavala, Dr. P. D. Burrows and his colleague for critical reading of the manuscript.

	Footnotes

 
¹ This work has been supported by grants from the Japan Science Technology Corporation and the Human Frontier Science Program (RG-366/96).

² Address correspondence and reprint requests to Dr. Ryo Goitsuka, Division of Molecular Biology, Research Institute for Biological Sciences, Science University of Tokyo, 2669 Yamazaki, Noda-city, Chiba 278, Japan; E-mail:

³ Abbreviations used in this paper: BCR; B cell Ag receptor, NF-AT; Nuclear factor of activated T cells, pY; phosphotyrosine; BASH, B cell adaptor containing an SH2 domain.

Received for publication August 6, 1998. Accepted for publication September 25, 1998.

	References

Top Abstract Introduction Materials and Methods Results and Discussion References

 

1. Masteller, E. L., G. T. Pharr, P. E. Funk, C. B. Thompson. 1997. Avian B cell development. Int. Rev. Immunol. 15:185.[Medline]
2. Reynaud, C.-A., B. Bertocci, A. Dahan, J.-C. Weill. 1994. Formation of the chicken B-cell repertoire: ontogenesis, regulation of Ig gene rearrangement, and diversification by gene conversion. Adv. Immunol. 57:353.[Medline]
3. Lassila, O.. 1989. Emigration of B cells from chicken bursa of Fabricius. Eur. J. Immunol. 19:955.[Medline]
4. Motyka, B., J. D. Reynold. 1991. Apoptosis is associated with the extensive B cell death in the sheep ileal Peyer’s patch and the chicken bursa of Fabricius: possible role in B cell selection. Eur. J. Immunol. 21:1951.[Medline]
5. Lam, K. P., R. Kuhn, K. Rajewsky. 1997. In vivo ablation of surface immunoglobulin on mature B cells by inducible gene targeting results in rapid cell death. Cell 90:1073.[Medline]
6. Paramithiotis, E., K. A. Jacobsen, M. J. Ratcliffe. 1995. Loss of surface immunoglobulin expression precedes B cell death by apoptosis in the bursa of Fabricius. J. Exp. Med. 181:105.[Abstract/Free Full Text]
7. Ratcliffe, M. J. H., L. Tkalec. 1990. Cross-linking of the surface immunoglobulin on lymphocytes from the bursa of Fabricius results in second messenger generation. Eur. J. Immunol. 20:1073.[Medline]
8. DeFranco, A. L.. 1997. The complexity of signaling pathways activated by the BCR. Curr. Opin. Immunol. 9:296.[Medline]
9. Takata, M., H. Sabe, A. Hata, T. Inazu, Y. Homma, T. Nukada, H. Yamamura, T. Kurosaki. 1994. Tyrosine kinases Lyn and Syk regulate B cell receptor-coupled Ca²⁺ mobilization through distinct pathways. EMBO J. 13:1341.[Medline]
10. Nieto, M. A., K. Patel, D. G. Wilkinson. 1996. In situ hybridization analysis of chick embryos in whole mount and tissue sections. Methods Cell Biol. 51:219.[Medline]
11. Katsuta, H., S. Tsuji, Y. Niho, T. Kurosaki, D. Kitamura. 1998. Lyn-mediated down-regulation of B cell antigen receptor signaling: inhibition of protein kinase C activation by Lyn in a kinase-independent fashion. J. Immunol. 160:1547.[Abstract/Free Full Text]
12. Bueerstedde, J., C.-A. Reynaud, E. H. Humphries, W. Olson, D. L. Ewert, J.-C. Weil. 1990. Light chain gene conversion continues at high rate in an ALV-induced cell line. EMBO J. 9:921.[Medline]
13. Songyang, Z., S. E. Shoelson, J. McGlade, P. Olivier, T. Pawson, X. R. Bustelo, M. Barbacid, H. Sabe, H. Hanafusa, T. Yi, R. Ren, D. Baltimore, S. Ratnofsky, R. A. Feldman, L. C. Cantley. 1994. Specific motifs recognized by the SH2 domains of Csk, 3BP2, fps/fes, GRB-2, HCP, SHC, Syk, and Vav. Mol. Cell. Biol. 14:2777.[Abstract/Free Full Text]
14. Songyang, Z., S. E. Shoelson, M. Chaudhuri, G. Gish, T. Pawson, W. G. Haser, F. King, T. Roberts, S. Ratnofsky, R. J. Lechleider, B. G. Neel, R. B. Birge, J. E. Fajardo, M. M. Chou, H. Hanafusa, B. Schaffhausen, L. C. Cantley. 1993. SH2 domains recognize specific phosphopeptide sequences. Cell 72:767.[Medline]
15. Sparks, A. B., J. E. Rider, N. G. Hoffman, D. M. Fowlkes, L. A. Quilliam, B. K. Kay. 1996. Distinct ligand preferences of Src homology 3 domains from Src, Yes, Abl, Cortactin, p53bp2, PLC, Crk, and Grb2. Proc. Natl. Acad. Sci. USA 93:1540.[Abstract/Free Full Text]
16. Jackman, J. K., D. G. Motto, Q. Sun, M. Takemoto, C. W. Turck, G. A. Peltz, G. A. Koretzky, P. R. Findell. 1995. Molecular cloning of SLP-76, a 76-kDa tyrosine phosphoprotein associated with Grb2 in T cells. J. Biol. Chem. 270:7029.[Abstract/Free Full Text]
17. Fang, N., D. G. Motto, S. E. Ross, G. A. Koretzky. 1996. Tyrosines 113, 128, 145 of SLP-76 are required for optimal augmentation of NF-AT promoter activity. J. Immunol. 157:3769.[Abstract]
18. Motto, D. G., S. E. Ross, J. Wu, L. R. Hendricks-Taylor, G. A. Koretzky. 1996. Implication of the Grb2-associated phosphoprotein SLP-76 in T cell receptor-mediated interleukin 2 production. J. Exp. Med. 183:1937.[Abstract/Free Full Text]
19. Venkataraman, L., D. A. Francis, Z. Wang, J. Liu, T. L. Rothstein, R. Sen. 1994. Cyclosporin-A sensitive induction of NF-AT in murine B cells. Immunity 1:189.[Medline]
20. Fu, C., C. W. Turck, T. Kurosaki, A. C. Chan. 1998. BLNK: a central linker protein in B cell activation. Immunity 9:93.[Medline]
21. Wienands, J., J. Schweikert, B. Wollscheid, H. Jumaa, P. J. Nielsen, M. Reth. 1998. SLP-65: a new signaling component in B lymphocytes which requires expression of the antigen receptor for phosphorylation. J. Exp. Med. 188:791.[Abstract/Free Full Text]

This article has been cited by other articles:

				T. Oellerich, M. Gronborg, K. Neumann, H.-H. Hsiao, H. Urlaub, and J. Wienands SLP-65 Phosphorylation Dynamics Reveals a Functional Basis for Signal Integration by Receptor-proximal Adaptor Proteins Mol. Cell. Proteomics, July 1, 2009; 8(7): 1738 - 1750. [Abstract] [Full Text] [PDF]

				Y. Imamura, A. Oda, T. Katahira, K. Bundo, K. A. Pike, M. J. H. Ratcliffe, and D. Kitamura BLNK Binds Active H-Ras to Promote B Cell Receptor-mediated Capping and ERK Activation J. Biol. Chem., April 10, 2009; 284(15): 9804 - 9813. [Abstract] [Full Text] [PDF]

				M. Weber, B. Treanor, D. Depoil, H. Shinohara, N. E. Harwood, M. Hikida, T. Kurosaki, and F. D. Batista Phospholipase C-{gamma}2 and Vav cooperate within signaling microclusters to propagate B cell spreading in response to membrane-bound antigen J. Exp. Med., April 14, 2008; 205(4): 853 - 868. [Abstract] [Full Text] [PDF]

				A. Abudula, A. Grabbe, M. Brechmann, C. Polaschegg, N. Herrmann, I. Goldbeck, K. Dittmann, and J. Wienands SLP-65 Signal Transduction Requires Src Homology 2 domain-mediated Membrane Anchoring and a Kinase-independent Adaptor Function of Syk J. Biol. Chem., September 28, 2007; 282(39): 29059 - 29066. [Abstract] [Full Text] [PDF]

				R. Goitsuka, C.-l. H. Chen, L. Benyon, Y. Asano, D. Kitamura, and M. D. Cooper Chicken cathelicidin-B1, an antimicrobial guardian at the mucosal M cell gateway PNAS, September 18, 2007; 104(38): 15063 - 15068. [Abstract] [Full Text] [PDF]

				A. Grabbe and J. Wienands Human SLP-65 isoforms contribute differently to activation and apoptosis of B lymphocytes Blood, December 1, 2006; 108(12): 3761 - 3768. [Abstract] [Full Text] [PDF]

				V. G. Karur, C. A. Lowell, P. Besmer, V. Agosti, and D. M. Wojchowski Lyn kinase promotes erythroblast expansion and late-stage development Blood, September 1, 2006; 108(5): 1524 - 1532. [Abstract] [Full Text] [PDF]

				N. Fujiwara, S. Hidano, H. Mamada, K. Ogasawara, D. Kitamura, M. D Cooper, N. Hozumi, C.-l. H Chen, and R. Goitsuka A novel avian homologue of CD72, chB1r, down modulates BCR-mediated activation signals Int. Immunol., May 1, 2006; 18(5): 775 - 783. [Abstract] [Full Text] [PDF]

				T. Katahira, Y. Imamura, and D. Kitamura The BASH/BLNK/SLP-65-associated protein BNAS1 regulates antigen-receptor signal transmission in B cells Int. Immunol., April 1, 2006; 18(4): 545 - 553. [Abstract] [Full Text] [PDF]

				H. Oshiumi, K. Shida, R. Goitsuka, Y. Kimura, J. Katoh, S. Ohba, Y. Tamaki, T. Hattori, N. Yamada, N. Inoue, et al. Regulator of Complement Activation (RCA) Locus in Chicken: Identification of Chicken RCA Gene Cluster and Functional RCA Proteins J. Immunol., August 1, 2005; 175(3): 1724 - 1734. [Abstract] [Full Text] [PDF]

				K. Hayashi, T. Nojima, R. Goitsuka, and D. Kitamura Impaired Receptor Editing in the Primary B Cell Repertoire of BASH-Deficient Mice J. Immunol., November 15, 2004; 173(10): 5980 - 5988. [Abstract] [Full Text] [PDF]

				M. Yamamoto, T. Nojima, K. Hayashi, R. Goitsuka, K. Furukawa, T. Azuma, and D. Kitamura BASH-deficient mice: limited primary repertoire and antibody formation, but sufficient affinity maturation and memory B cell generation, in anti-NP response Int. Immunol., August 1, 2004; 16(8): 1161 - 1171. [Abstract] [Full Text] [PDF]

				Y. Imamura, T. Katahira, and D. Kitamura Identification and Characterization of a Novel BASH N Terminus-associated Protein, BNAS2 J. Biol. Chem., June 18, 2004; 279(25): 26425 - 26432. [Abstract] [Full Text] [PDF]

				A. Reichlin, A. Gazumyan, H. Nagaoka, K. H. Kirsch, M. Kraus, K. Rajewsky, and M. C. Nussenzweig A B Cell Receptor with Two Ig{alpha} Cytoplasmic Domains Supports Development of Mature But Anergic B Cells J. Exp. Med., March 15, 2004; 199(6): 855 - 865. [Abstract] [Full Text] [PDF]

				K. E. Nichols, K. Haines, P. S. Myung, S. Newbrough, E. Myers, H. Jumaa, D. J. Shedlock, H. Shen, and G. A. Koretzky Macrophage activation and Fc{gamma} receptor-mediated signaling do not require expression of the SLP-76 and SLP-65 adaptors J. Leukoc. Biol., March 1, 2004; 75(3): 541 - 552. [Abstract] [Full Text] [PDF]

				Y.-w. Su, A. Flemming, T. Wossning, E. Hobeika, M. Reth, and H. Jumaa Identification of a Pre-BCR Lacking Surrogate Light Chain J. Exp. Med., December 1, 2003; 198(11): 1699 - 1706. [Abstract] [Full Text] [PDF]

				R. L. Wange The Missing Link(er): A Return to Symmetry in Antigen Receptor Signaling? Mol. Interv., March 1, 2003; 3(2): 75 - 78. [Abstract] [Full Text]

				S. Xu and K.-P. Lam Delayed Cellular Maturation and Decreased Immunoglobulin {kappa} Light Chain Production In Immature B Lymphocytes Lacking B Cell Linker Protein J. Exp. Med., July 15, 2002; 196(2): 197 - 206. [Abstract] [Full Text] [PDF]

				K. Mizuno, Y. Tagawa, K. Mitomo, N. Watanabe, T. Katagiri, M. Ogimoto, and H. Yakura Src Homology Region 2 Domain-Containing Phosphatase 1 Positively Regulates B Cell Receptor-Induced Apoptosis by Modulating Association of B Cell Linker Protein with Nck and Activation of c-Jun NH2-Terminal Kinase J. Immunol., July 15, 2002; 169(2): 778 - 786. [Abstract] [Full Text] [PDF]

				S. Kabak, B. J. Skaggs, M. R. Gold, M. Affolter, K. L. West, M. S. Foster, K. Siemasko, A. C. Chan, R. Aebersold, and M. R. Clark The Direct Recruitment of BLNK to Immunoglobulin {alpha} Couples the B-Cell Antigen Receptor to Distal Signaling Pathways Mol. Cell. Biol., April 15, 2002; 22(8): 2524 - 2535. [Abstract] [Full Text] [PDF]

				D. Watanabe, S. Hashimoto, M. Ishiai, M. Matsushita, Y. Baba, T. Kishimoto, T. Kurosaki, and S. Tsukada Four Tyrosine Residues in Phospholipase C-gamma 2, Identified as Btk-dependent Phosphorylation Sites, Are Required for B Cell Antigen Receptor-coupled Calcium Signaling J. Biol. Chem., October 12, 2001; 276(42): 38595 - 38601. [Abstract] [Full Text] [PDF]

				J. Yu, C. Riou, D. Davidson, R. Minhas, J. D. Robson, M. Julius, R. Arnold, F. Kiefer, and A. Veillette Synergistic Regulation of Immunoreceptor Signaling by SLP-76-Related Adaptor Clnk and Serine/Threonine Protein Kinase HPK-1 Mol. Cell. Biol., September 15, 2001; 21(18): 6102 - 6112. [Abstract] [Full Text] [PDF]

				S. Tsuji, M. Okamoto, K. Yamada, N. Okamoto, R. Goitsuka, R. Arnold, F. Kiefer, and D. Kitamura B Cell Adaptor Containing Src Homology 2 Domain (BASH) Links B Cell Receptor Signaling to the Activation of Hematopoietic Progenitor Kinase 1 J. Exp. Med., August 20, 2001; 194(4): 529 - 540. [Abstract] [Full Text] [PDF]

				N. Engels, M. Merchant, R. Pappu, A. C. Chan, R. Longnecker, and J. Wienands Epstein-Barr Virus Latent Membrane Protein 2A (LMP2A) Employs the SLP-65 Signaling Module J. Exp. Med., July 30, 2001; 194(3): 255 - 264. [Abstract] [Full Text] [PDF]

				S. Xu, S.-C. Wong, and K.-P. Lam Cutting Edge: B Cell Linker Protein Is Dispensable for the Allelic Exclusion of Immunoglobulin Heavy Chain Locus But Required for the Persistence of CD5+ B Cells J. Immunol., October 15, 2000; 165(8): 4153 - 4157. [Abstract] [Full Text] [PDF]

				, , , A. , , and Involvement of LAT, Gads, and Grb2 in Compartmentation of SLP-76 to the Plasma J. Exp. Med., September 18, 2000; 192(6): 847 - 856. [Abstract] [Full Text] [PDF]

				R. Muller, J. Wienands, and M. Reth The serine and threonine residues in the Ig-alpha cytoplasmic tail negatively regulate immunoreceptor tyrosine-based activation motif-mediated signal transduction PNAS, July 18, 2000; 97(15): 8451 - 8454. [Abstract] [Full Text] [PDF]

				R. Goitsuka, H. Kanazashi, H. Sasanuma, Y.-i. Fujimura, Y. Hidaka, A. Tatsuno, C. Ra, K. Hayashi, and D. Kitamura A BASH/SLP-76-related adaptor protein MIST/Clnk involved in IgE receptor-mediated mast cell degranulation Int. Immunol., April 1, 2000; 12(4): 573 - 580. [Abstract] [Full Text] [PDF]

				S. Xu, J. E.-L. Tan, E. P.-Y. Wong, A. Manickam, S. Ponniah, and K.-P. Lam B cell development and activation defects resulting in xid-like immunodeficiency in BLNK/SLP-65-deficient mice Int. Immunol., March 1, 2000; 12(3): 397 - 404. [Abstract] [Full Text] [PDF]

				T. Yasuda, A. Maeda, M. Kurosaki, T. Tezuka, K. Hironaka, T. Yamamoto, and T. Kurosaki Cbl Suppresses B Cell Receptor-mediated Phospholipase C (PLC)-{gamma}2 Activation by Regulating B Cell Linker Protein-PLC-{gamma}2 Binding J. Exp. Med., February 21, 2000; 191(4): 641 - 650. [Abstract] [Full Text] [PDF]

				R. Pappu, A. M. Cheng, B. Li, Q. Gong, C. Chiu, N. Griffin, M. White, B. P. Sleckman, and A. C. Chan Requirement for B Cell Linker Protein (BLNK) in B Cell Development Science, December 3, 1999; 286(5446): 1949 - 1954. [Abstract] [Full Text]

				Y. Minegishi, J. Rohrer, E. Coustan-Smith, H. M. Lederman, R. Pappu, D. Campana, A. C. Chan, and M. E. Conley An Essential Role for BLNK in Human B Cell Development Science, December 3, 1999; 286(5446): 1954 - 1957. [Abstract] [Full Text]

				M. Y. Cao, D. Davidson, J. Yu, S. Latour, and A. Veillette Clnk, a Novel SLP-76-related Adaptor Molecule Expressed in Cytokine-stimulated Hemopoietic Cells J. Exp. Med., November 15, 1999; 190(10): 1527 - 1534. [Abstract] [Full Text] [PDF]

				M. RETH and J. WIENANDS The Maintenance and the Activation Signal of the B-cell Antigen Receptor Cold Spring Harb Symp Quant Biol, January 1, 1999; 64(0): 323 - 328. [Abstract] [PDF]

				J. Wong, M. Ishiai, T. Kurosaki, and A. C. Chan Functional Complementation of BLNK by SLP-76 and LAT Linker Proteins J. Biol. Chem., October 13, 2000; 275(42): 33116 - 33122. [Abstract] [Full Text] [PDF]

				J. E.-L. Tan, S.-C. Wong, S. K.-E. Gan, S. Xu, and K.-P. Lam The Adaptor Protein BLNK Is Required for B Cell Antigen Receptor-induced Activation of Nuclear Factor-kappa B and Cell Cycle Entry and Survival of B Lymphocytes J. Biol. Chem., June 1, 2001; 276(23): 20055 - 20063. [Abstract] [Full Text] [PDF]

				R. Goitsuka, A. Tatsuno, M. Ishiai, T. Kurosaki, and D. Kitamura MIST Functions through Distinct Domains in Immunoreceptor Signaling in the Presence and Absence of LAT J. Biol. Chem., September 14, 2001; 276(38): 36043 - 36050. [Abstract] [Full Text] [PDF]

				K. Sauer, J. Liou, S. B. Singh, D. Yablonski, A. Weiss, and R. M. Perlmutter Hematopoietic Progenitor Kinase 1 Associates Physically and Functionally with the Adaptor Proteins B Cell Linker Protein and SLP-76 in Lymphocytes J. Biol. Chem., November 21, 2001; 276(48): 45207 - 45216. [Abstract] [Full Text] [PDF]

				K. Hayashi, R. Nittono, N. Okamoto, S. Tsuji, Y. Hara, R. Goitsuka, and D. Kitamura The B cell-restricted adaptor BASH is required for normal development and antigen receptor-mediated activation of B cells PNAS, March 14, 2000; 97(6): 2755 - 2760. [Abstract] [Full Text] [PDF]

				Y. Baba, S. Hashimoto, M. Matsushita, D. Watanabe, T. Kishimoto, T. Kurosaki, and S. Tsukada BLNK mediates Syk-dependent Btk activation PNAS, February 27, 2001; 98(5): 2582 - 2586. [Abstract] [Full Text] [PDF]

This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Goitsuka, R. Articles by Kitamura, D. Search for Related Content PubMed PubMed Citation Articles by Goitsuka, R. Articles by Kitamura, D.