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The adaptor protein SLP-65 acts as a tumor suppressor that limits pre-B cell expansion

Nature Immunologyvolume 4pages3843 (2003) | Download Citation



Mice deficient in the adaptor protein SLP-65 (also known as BLNK) have reduced numbers of mature B cells, but an increased pre-B cell compartment. We show here that compared to wild-type cells, SLP-65−/− pre-B cells show an enhanced ex vivo proliferative capacity. This proliferation requires interleukin 7 and expression of the pre-B cell receptor (pre-BCR). In addition, SLP-65−/− mice have a high incidence of pre-B cell lymphoma. Reintroduction of SLP-65 into SLP-65−/− pre-B cells led to pre-BCR down-regulation and enhanced differentiation. Our results indicate that SLP-65 regulates a developmental program that promotes differentiation and limits pre-B cell expansion, thereby acting as a tumor suppressor.

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  1. 1

    Hardy, R.R., Carmack, C.E., Shinton, S.A., Kemp, J.D. & Hayakawa, K. Resolution and characterization of pro-B and pre-pro-B cell stages in normal mouse bone marrow. J. Exp. Med. 173, 1213–1225 (1991).

  2. 2

    Li, Y.S., Hayakawa, K. & Hardy, R.R. The regulated expression of B lineage associated genes during B cell differentiation in bone marrow and fetal liver. J. Exp. Med. 178, 951–960 (1993).

  3. 3

    Rolink, A.G. et al. B cell development in the mouse from early progenitors to mature B cells. Immunol. Lett. 68, 89–93 (1999).

  4. 4

    Osmond, D.G., Rolink, A. & Melchers, F. Murine B lymphopoiesis: towards a unified model. Immunol. Today 19, 65–68 (1998).

  5. 5

    Rajewsky, K. Clonal selection and learning in the antibody system. Nature 381, 751–758 (1996).

  6. 6

    Karasuyama, H. et al. The roles of preB cell receptor in early B cell development and its signal transduction. Immunol. Cell Biol. 75, 209–216 (1997).

  7. 7

    Martensson, I.-L. & Ceredig, R. Role of the surrogate light chain and the pre-B-cell receptor in mouse B cell development. Immunology 101, 435–441 (2000).

  8. 8

    Conley, M.E., Rohrer, J., Rapalus, L., Boylin, E.C. & Minegishi, Y. Defects in early B-cell development: comparing the consequences of abnormalities in pre-BCR signaling in the human and the mouse. Immunol. Rev. 178, 75–90 (2000).

  9. 9

    ten Boekel, E., Melchers, F. & Rolink, A.G. Precursor B cells showing H chain allelic inclusion display allelic exclusion at the level of pre-B cell receptor surface expression. Immunity 8, 199–207 (1998).

  10. 10

    Kitamura, D., Roes, J., Kuhn, R. & Rajewsky, K.A. B cell-deficient mouse by targeted disruption of the membrane exon of the immunoglobulin μ chain gene. Nature 350, 423–426 (1991).

  11. 11

    Kitamura, D. et al. A critical role of λ5 protein in B cell development. Cell 69, 823–831 (1992).

  12. 12

    Mundt, C., Licence, S., Shimizu, T., Melchers, F. & Martensson, I.L. Loss of precursor B cell expansion but not allelic exclusion in VpreB1/VpreB2 double-deficient mice. J. Exp. Med. 193, 435–445 (2001).

  13. 13

    Tamir, I. & Cambier, J.C. Antigen receptor signaling - integration of protein tyrosine kinase functions. Oncogene 17, 1353–1364 (1998).

  14. 14

    Kurosaki, T. Genetic analysis of B cell antigen receptor signaling. Annu. Rev. Immunol. 17, 555–592 (1999).

  15. 15

    Benschop, R.J. & Cambier, J.C. B cell development: signal transduction by antigen receptors and their surrogates. Curr. Opin. Immunol. 11, 143–151 (1999).

  16. 16

    Kelly, M.E. & Chan, A.C. Regulation of B cell function by linker proteins. Curr. Opin. Immunol. 12, 267–275 (2000).

  17. 17

    Myung, P.S., Boerthe, N.J. & Koretzky, G.A. Adapter proteins in lymphocyte antigen-receptor signaling. Curr. Opin. Immunol. 12, 256–266 (2000).

  18. 18

    Tomlinson, M.G., Lin, J. & Weiss, A. Lymphocytes with a complex: adapter proteins in antigen receptor signaling. Immunol. Today 21, 584–591 (2000).

  19. 19

    Pivniouk, V. et al. Impaired viability and profound block in thymocyte development in mice lacking the adaptor protein Slp-76. Cell 94, 229–238 (1998).

  20. 20

    Clements, J.L. et al. Requirement for the leukocyte-specific adapter protein Slp-76 for normal T-cell development. Science 281, 416–419 (1998).

  21. 21

    Zhang, W.G. et al. Essential role of LAT in T cell development. Immunity 10, 323–332 (1999).

  22. 22

    Wienands, J. et al. Slp-65 - a new signaling component in B lymphocytes which requires expression of the antigen receptor for phosphorylation. J. Exp. Med. 188, 791–795 (1998).

  23. 23

    Fu, C., Turck, C.W., Kurosaki, T. & Chan, A.C. Blnk - a central linker protein in B cell activation. Immunity 9, 93–103 (1998).

  24. 24

    Goitsuka, R. et al. Cutting Edge - Bash, a novel signaling molecule preferentially expressed in B cells of the Bursa of Fabricius. J. Immunol. 161, 5804–5808 (1998).

  25. 25

    Su, Y.W. et al. Interaction of SLP adaptors with the SH2 domain of Tec family kinases. Eur. J. Immunol. 29, 3702–3711 (1999).

  26. 26

    Hashimoto, S. et al. Identification of the SH2 domain binding protein of Bruton's tyrosine kinase as BLNK - Functional significance of Btk-SH2 domain in B-cell antigen receptor-coupled calcium signaling. Blood 94, 2357–2364 (1999).

  27. 27

    Ishiai, M. et al. BLNK required for coupling Syk to PLC-γ2 and Rac1-JNK in B cells. Immunity 10, 117–125 (1999).

  28. 28

    Kurosaki, T. et al. Regulation of the phospholipase C-γ2 pathway in B cells. Immunol. Rev. 176, 19–29 (2000).

  29. 29

    Jumaa, H. et al. Abnormal development and function of B lymphocytes in mice deficient for the signaling adaptor protein SLP-65. Immunity 11, 547–554 (1999).

  30. 30

    Pappu, R. et al. Requirement for B cell linker protein (BLNK) in B cell development. Science 286, 1949–1954 (1999).

  31. 31

    Hayashi, K. et al. The B cell-restricted adaptor BASH is required for normal development and antigen receptor-mediated activation of B cells. Proc. Natl. Acad. Sci. USA 97, 2755–2760 (2000).

  32. 32

    Xu, S.L. et al. B cell development and activation defects resulting in xid-like immunodeficiency in BLNK/SLP-65-deficient mice. Int. Immunol. 12, 397–404 (2000).

  33. 33

    Rolink, A., Kudo, A., Karasuyama, H., Kikuchi, Y. & Melchers, F. Long-term proliferating early pre B cell lines and clones with the potential to develop to surface Ig-positive, mitogen reactive B cells in vitro and in vivo. EMBO J. 10, 327–336 (1991).

  34. 34

    Ray, R.J. et al. Stromal cell-independent maturation of Il-7-responsive pro-B cells. J. Immunol. 160, 5886–5897 (1998).

  35. 35

    Karasuyama, H. et al. The expression of Vpre-B/λ5 surrogate light chain in early bone marrow precursor B cells of normal and B cell-deficient mutant mice. Cell 77, 133–143 (1994).

  36. 36

    Winkler, T.H., Rolink, A., Melchers, F. & Karasuyama, H. Precursor B cells of mouse bone marrow express two different complexes with the surrogate light chain on the surface. Eur. J. Immunol. 25, 446–450 (1995).

  37. 37

    Marshall, A.J., Fleming, H.E., Wu, G.E. & Paige, C.J. Modulation of the IL-7 dose-response threshold during pro-B cell differentiation is dependent on pre-B cell receptor expression. J. Immunol. 161, 6038–6045 (1998).

  38. 38

    Stephan, R.P., Elgavish, E., Karasuyama, H., Kubagawa, H. & Cooper, M.D. Analysis of VpreB expression during B lineage differentiation in λ5-deficient mice. J. Immunol. 167, 3734–3739 (2001).

  39. 39

    Grawunder, U. et al. Down-regulation of Rag1 and Rag2 gene expression in pre-B cells after functional immunoglobulin heavy chain rearrangement. Immunity 3, 601–608 (1995).

  40. 40

    Craxton, A., Otipoby, K.L., Jiang, A.M. & Clark, A. Signal transduction pathways that regulate the fate of B lymphocytes. Adv. Immunol. 73, 79–152 (1999).

  41. 41

    Brazil, D.P. & Hemmings, B.A. Ten years of protein kinase B signalling: a hard Akt to follow. Trends Biochem. Sci. 26, 657–664 (2001).

  42. 42

    Ketteler, R., Glaser, S., Sandra, O., Martens, U.M. & Klingmüller, U. Enhanced transgene expression in primitive hematopoietic progenitor cells and embryonic stem cells efficiently transduced by optimized retroviral hybrid vectors. Gene Ther. 9, 477–487 (2002).

  43. 43

    Gugasyan, R. et al. Rel/NF-κB transcription factors: key mediators of B-cell activation. Immunol. Rev. 176, 134–140 (2000).

  44. 44

    Tan, J.E.L., Wong, S.C., Gan, S.K.E., Xu, S.L. & Lam, K.P. The adaptor protein BLNK is required for B cell antigen receptor-induced activation of nuclear factor-κB and cell cycle entry and survival of B lymphocytes. J. Biol. Chem. 276, 20055–20063 (2001).

  45. 45

    Shaw, A.C., Swat, W., Davidson, L. & Alt, F.W. Induction of Ig light chain gene rearrangement in heavy chain-deficient B cells by activated Ras. Proc. Natl. Acad. Sci. USA 96, 2239–2243 (1999).

  46. 46

    Shaw, A.C., Swat, W., Ferrini, R., Davidson, L. & Alt, F.W. Activated Ras signals developmental progression of recombinase-activating gene (RAG)-deficient pro-B lymphocytes. J. Exp. Med. 189, 123–129 (1999).

  47. 47

    Nagaoka, H. et al. Ras mediates effector pathways responsible for pre-B cell survival, which is essential for the developmental progression to the late pre-B cell stage. J. Exp. Med. 192, 171–181 (2000).

  48. 48

    Fleming, H.E. & Paige, C.J. Pre-B cell receptor signaling mediates selective response to IL-7 at the pro-B to pre-B cell transition via an Erk/MAP kinase-dependent pathway. Immunity 15, 521–531 (2001).

  49. 49

    Jumaa, H. & Nielsen, P.J. The splicing factor SRp20 modifies splicing of its own mRNA and ASF/SF2 antagonizes this regulation. EMBO J. 16, 5077–5085 (1997).

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We thank P. Nielsen for critically reading the manuscript; A. Wuerch for cell sorting, C. Eschbach and M. Mitterer for technical help; U. Klingmüller for providing retroviral constructs and expert advice; H. Eibel and P. Richter for help in establishing the in vitro culture system. Supported by the Deutsche Forschungs Gemeinschaft through SFB 364 and the Leibniz program.

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  1. Institute for Biology III, Albert-Ludwigs University of Freiburg and Max Planck Institute for Immunobiology, Stuebeweg 51, Freiburg, 79108, Germany

    • Alexandra Flemming
    • , Tilman Brummer
    • , Michael Reth
    •  & Hassan Jumaa


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The authors declare no competing financial interests.

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Correspondence to Michael Reth or Hassan Jumaa.

Supplementary information

  1. Web Fig. 1.

    The number of κ+ cells increases in SLP-65–/– , but not in WT pre-B cell cultures after IL-7 withdrawal. WT and SLP-65–/– pre-B cells from 1-week cultures were analyzed for κ expression and subsequently plated-out in medium lacking IL-7 for 2 days. The ratio of κ+ cells was determined before and after IL-7 withdrawal. (PDF 22 kb)

  2. Web Fig. 2.

    The enhanced proliferation capacity of SLP-65–/– BM-derived B cells depends on pre-BCR expansion. (a) c-Kit versus B220 FACS profiles of lymphocyte-gated cells derived from total BM littermates. (b) Sorted B220+ c-Kit+ cells were cultured in IL-7 (>1 ng/ml). Proliferation was assessed by counting at the indicated time points, values were averaged between duplicates. After 5 days of in vitro culture, the number of SLP-65–/– cells had increased 70-fold, whereas the number of WT cells had only increased 5.5-fold. (PDF 66 kb)

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