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. 1995 Jul 17;14(14):3403–3414. doi: 10.1002/j.1460-2075.1995.tb07346.x

LckBP1, a proline-rich protein expressed in haematopoietic lineage cells, directly associates with the SH3 domain of protein tyrosine kinase p56lck.

Y Takemoto 1, M Furuta 1, X K Li 1, W J Strong-Sparks 1, Y Hashimoto 1
PMCID: PMC394407  PMID: 7628441

Abstract

The Lck tyrosine kinase molecule plays an essential role in T cell activation and T cell development. Using the expression cloning technique, we have isolated a gene that encodes a molecule, LckBP1, able to associate with murine Lck. Analysis of full-length LckBP1 cDNA indicates at least four potentially important segments: a four tandem 37 amino acid repeat motif with a potential helix-turn-helix DNA binding motif; a proline-rich region; a proline-glutamate repeat; and an SH3 domain. These four regions are very similar to the human haematopoietic-specific protein 1 (HS1). Deletion mutant analysis of LckBP1 revealed two proline-rich regions that permit association with Lck SH3. One region contains prolines conserved among HS1 and cortactin, and the other region contains a potential MAP kinase recognition site. In vivo association between Lck and LckBP1 was confirmed by immunoprecipitation of lysates from a pre-T cell line and adult thymocytes using antibodies specific for Lck and LckBP1. LckBP1 is tyrosine phosphorylated after T-cell receptor stimulation. The SH3 domain and the potential helix-turn-helix motif in LckBP1 suggest that this molecule may associate with various molecules and function as a DNA binding molecule. The data also suggest that LckBP1 mediates intracellular signalling through Lck in T cells.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Anderson S. J., Levin S. D., Perlmutter R. M. Protein tyrosine kinase p56lck controls allelic exclusion of T-cell receptor beta-chain genes. Nature. 1993 Oct 7;365(6446):552–554. doi: 10.1038/365552a0. [DOI] [PubMed] [Google Scholar]
  2. Autero M., Saharinen J., Pessa-Morikawa T., Soula-Rothhut M., Oetken C., Gassmann M., Bergman M., Alitalo K., Burn P., Gahmberg C. G. Tyrosine phosphorylation of CD45 phosphotyrosine phosphatase by p50csk kinase creates a binding site for p56lck tyrosine kinase and activates the phosphatase. Mol Cell Biol. 1994 Feb;14(2):1308–1321. doi: 10.1128/mcb.14.2.1308. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bergman M., Mustelin T., Oetken C., Partanen J., Flint N. A., Amrein K. E., Autero M., Burn P., Alitalo K. The human p50csk tyrosine kinase phosphorylates p56lck at Tyr-505 and down regulates its catalytic activity. EMBO J. 1992 Aug;11(8):2919–2924. doi: 10.1002/j.1460-2075.1992.tb05361.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Blackwood E. M., Eisenman R. N. Max: a helix-loop-helix zipper protein that forms a sequence-specific DNA-binding complex with Myc. Science. 1991 Mar 8;251(4998):1211–1217. doi: 10.1126/science.2006410. [DOI] [PubMed] [Google Scholar]
  5. Brzeska H., Lynch T. J., Martin B., Korn E. D. The localization and sequence of the phosphorylation sites of Acanthamoeba myosins I. An improved method for locating the phosphorylated amino acid. J Biol Chem. 1989 Nov 15;264(32):19340–19348. [PubMed] [Google Scholar]
  6. Chou P. Y., Fasman G. D. Empirical predictions of protein conformation. Annu Rev Biochem. 1978;47:251–276. doi: 10.1146/annurev.bi.47.070178.001343. [DOI] [PubMed] [Google Scholar]
  7. Chow L. M., Fournel M., Davidson D., Veillette A. Negative regulation of T-cell receptor signalling by tyrosine protein kinase p50csk. Nature. 1993 Sep 9;365(6442):156–160. doi: 10.1038/365156a0. [DOI] [PubMed] [Google Scholar]
  8. Cicchetti P., Mayer B. J., Thiel G., Baltimore D. Identification of a protein that binds to the SH3 region of Abl and is similar to Bcr and GAP-rho. Science. 1992 Aug 7;257(5071):803–806. doi: 10.1126/science.1379745. [DOI] [PubMed] [Google Scholar]
  9. Clark S. G., Stern M. J., Horvitz H. R. C. elegans cell-signalling gene sem-5 encodes a protein with SH2 and SH3 domains. Nature. 1992 Mar 26;356(6367):340–344. doi: 10.1038/356340a0. [DOI] [PubMed] [Google Scholar]
  10. Drubin D. G., Mulholland J., Zhu Z. M., Botstein D. Homology of a yeast actin-binding protein to signal transduction proteins and myosin-I. Nature. 1990 Jan 18;343(6255):288–290. doi: 10.1038/343288a0. [DOI] [PubMed] [Google Scholar]
  11. Duplay P., Thome M., Hervé F., Acuto O. p56lck interacts via its src homology 2 domain with the ZAP-70 kinase. J Exp Med. 1994 Apr 1;179(4):1163–1172. doi: 10.1084/jem.179.4.1163. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Ettehadieh E., Sanghera J. S., Pelech S. L., Hess-Bienz D., Watts J., Shastri N., Aebersold R. Tyrosyl phosphorylation and activation of MAP kinases by p56lck. Science. 1992 Feb 14;255(5046):853–855. doi: 10.1126/science.1311128. [DOI] [PubMed] [Google Scholar]
  13. Franz W. M., Berger P., Wang J. Y. Deletion of an N-terminal regulatory domain of the c-abl tyrosine kinase activates its oncogenic potential. EMBO J. 1989 Jan;8(1):137–147. doi: 10.1002/j.1460-2075.1989.tb03358.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Garnier J., Osguthorpe D. J., Robson B. Analysis of the accuracy and implications of simple methods for predicting the secondary structure of globular proteins. J Mol Biol. 1978 Mar 25;120(1):97–120. doi: 10.1016/0022-2836(78)90297-8. [DOI] [PubMed] [Google Scholar]
  15. Gonzalez F. A., Raden D. L., Davis R. J. Identification of substrate recognition determinants for human ERK1 and ERK2 protein kinases. J Biol Chem. 1991 Nov 25;266(33):22159–22163. [PubMed] [Google Scholar]
  16. Harrison S. C., Aggarwal A. K. DNA recognition by proteins with the helix-turn-helix motif. Annu Rev Biochem. 1990;59:933–969. doi: 10.1146/annurev.bi.59.070190.004441. [DOI] [PubMed] [Google Scholar]
  17. Hashimoto Y., Blank K. J. T cell receptor genes and T cell development in virus-transformed early T cell lines. J Immunol. 1990 Feb 15;144(4):1518–1525. [PubMed] [Google Scholar]
  18. Hatakeyama M., Kono T., Kobayashi N., Kawahara A., Levin S. D., Perlmutter R. M., Taniguchi T. Interaction of the IL-2 receptor with the src-family kinase p56lck: identification of novel intermolecular association. Science. 1991 Jun 14;252(5012):1523–1528. doi: 10.1126/science.2047859. [DOI] [PubMed] [Google Scholar]
  19. Hirai H., Varmus H. E. Site-directed mutagenesis of the SH2- and SH3-coding domains of c-src produces varied phenotypes, including oncogenic activation of p60c-src. Mol Cell Biol. 1990 Apr;10(4):1307–1318. doi: 10.1128/mcb.10.4.1307. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Iseki R., Mukai M., Iwata M. Regulation of T lymphocyte apoptosis. Signals for the antagonism between activation- and glucocorticoid-induced death. J Immunol. 1991 Dec 15;147(12):4286–4292. [PubMed] [Google Scholar]
  21. Jackson P., Baltimore D. N-terminal mutations activate the leukemogenic potential of the myristoylated form of c-abl. EMBO J. 1989 Feb;8(2):449–456. doi: 10.1002/j.1460-2075.1989.tb03397.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Kato J. Y., Takeya T., Grandori C., Iba H., Levy J. B., Hanafusa H. Amino acid substitutions sufficient to convert the nontransforming p60c-src protein to a transforming protein. Mol Cell Biol. 1986 Dec;6(12):4155–4160. doi: 10.1128/mcb.6.12.4155. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Kitamura D., Kaneko H., Miyagoe Y., Ariyasu T., Watanabe T. Isolation and characterization of a novel human gene expressed specifically in the cells of hematopoietic lineage. Nucleic Acids Res. 1989 Nov 25;17(22):9367–9379. [PMC free article] [PubMed] [Google Scholar]
  24. Koch C. A., Anderson D., Moran M. F., Ellis C., Pawson T. SH2 and SH3 domains: elements that control interactions of cytoplasmic signaling proteins. Science. 1991 May 3;252(5006):668–674. doi: 10.1126/science.1708916. [DOI] [PubMed] [Google Scholar]
  25. Kubo M., Kincaid R. L., Ransom J. T. Activation of the interleukin-4 gene is controlled by the unique calcineurin-dependent transcriptional factor NF(P). J Biol Chem. 1994 Jul 29;269(30):19441–19446. [PubMed] [Google Scholar]
  26. Kubo M., Kincaid R. L., Webb D. R., Ransom J. T. The Ca2+/calmodulin-activated, phosphoprotein phosphatase calcineurin is sufficient for positive transcriptional regulation of the mouse IL-4 gene. Int Immunol. 1994 Feb;6(2):179–188. doi: 10.1093/intimm/6.2.179. [DOI] [PubMed] [Google Scholar]
  27. Kuwabara I., Ohno H., Punt J. A., Hashimoto Y., Saito T. Transition from TCR-beta dimer to TCR-alpha beta-expressing cells by introduction of an alpha-chain in an immature thymocyte cell line. J Immunol. 1994 Mar 1;152(5):2148–2156. [PubMed] [Google Scholar]
  28. Levin S. D., Anderson S. J., Forbush K. A., Perlmutter R. M. A dominant-negative transgene defines a role for p56lck in thymopoiesis. EMBO J. 1993 Apr;12(4):1671–1680. doi: 10.1002/j.1460-2075.1993.tb05812.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Li N., Batzer A., Daly R., Yajnik V., Skolnik E., Chardin P., Bar-Sagi D., Margolis B., Schlessinger J. Guanine-nucleotide-releasing factor hSos1 binds to Grb2 and links receptor tyrosine kinases to Ras signalling. Nature. 1993 May 6;363(6424):85–88. doi: 10.1038/363085a0. [DOI] [PubMed] [Google Scholar]
  30. Lowenstein E. J., Daly R. J., Batzer A. G., Li W., Margolis B., Lammers R., Ullrich A., Skolnik E. Y., Bar-Sagi D., Schlessinger J. The SH2 and SH3 domain-containing protein GRB2 links receptor tyrosine kinases to ras signaling. Cell. 1992 Aug 7;70(3):431–442. doi: 10.1016/0092-8674(92)90167-b. [DOI] [PubMed] [Google Scholar]
  31. Macgregor P. F., Abate C., Curran T. Direct cloning of leucine zipper proteins: Jun binds cooperatively to the CRE with CRE-BP1. Oncogene. 1990 Apr;5(4):451–458. [PubMed] [Google Scholar]
  32. McMahon A. P., Giebelhaus D. H., Champion J. E., Bailes J. A., Lacey S., Carritt B., Henchman S. K., Moon R. T. cDNA cloning, sequencing and chromosome mapping of a non-erythroid spectrin, human alpha-fodrin. Differentiation. 1987;34(1):68–78. doi: 10.1111/j.1432-0436.1987.tb00052.x. [DOI] [PubMed] [Google Scholar]
  33. Molina T. J., Kishihara K., Siderovski D. P., van Ewijk W., Narendran A., Timms E., Wakeham A., Paige C. J., Hartmann K. U., Veillette A. Profound block in thymocyte development in mice lacking p56lck. Nature. 1992 May 14;357(6374):161–164. doi: 10.1038/357161a0. [DOI] [PubMed] [Google Scholar]
  34. Moran E., Mathews M. B. Multiple functional domains in the adenovirus E1A gene. Cell. 1987 Jan 30;48(2):177–178. doi: 10.1016/0092-8674(87)90418-1. [DOI] [PubMed] [Google Scholar]
  35. Mustelin T., Altman A. Dephosphorylation and activation of the T cell tyrosine kinase pp56lck by the leukocyte common antigen (CD45). Oncogene. 1990 Jun;5(6):809–813. [PubMed] [Google Scholar]
  36. Mustelin T., Burn P. Regulation of src family tyrosine kinases in lymphocytes. Trends Biochem Sci. 1993 Jun;18(6):215–220. doi: 10.1016/0968-0004(93)90192-p. [DOI] [PubMed] [Google Scholar]
  37. Mustelin T., Coggeshall K. M., Altman A. Rapid activation of the T-cell tyrosine protein kinase pp56lck by the CD45 phosphotyrosine phosphatase. Proc Natl Acad Sci U S A. 1989 Aug;86(16):6302–6306. doi: 10.1073/pnas.86.16.6302. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Olivier J. P., Raabe T., Henkemeyer M., Dickson B., Mbamalu G., Margolis B., Schlessinger J., Hafen E., Pawson T. A Drosophila SH2-SH3 adaptor protein implicated in coupling the sevenless tyrosine kinase to an activator of Ras guanine nucleotide exchange, Sos. Cell. 1993 Apr 9;73(1):179–191. doi: 10.1016/0092-8674(93)90170-u. [DOI] [PubMed] [Google Scholar]
  39. Ostergaard H. L., Shackelford D. A., Hurley T. R., Johnson P., Hyman R., Sefton B. M., Trowbridge I. S. Expression of CD45 alters phosphorylation of the lck-encoded tyrosine protein kinase in murine lymphoma T-cell lines. Proc Natl Acad Sci U S A. 1989 Nov;86(22):8959–8963. doi: 10.1073/pnas.86.22.8959. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Pawson T. Non-catalytic domains of cytoplasmic protein-tyrosine kinases: regulatory elements in signal transduction. Oncogene. 1988 Nov;3(5):491–495. [PubMed] [Google Scholar]
  41. Potts W. M., Reynolds A. B., Lansing T. J., Parsons J. T. Activation of pp60c-src transforming potential by mutations altering the structure of an amino terminal domain containing residues 90-95. Oncogene Res. 1988;3(4):343–355. [PubMed] [Google Scholar]
  42. Prasad K. V., Kapeller R., Janssen O., Repke H., Duke-Cohan J. S., Cantley L. C., Rudd C. E. Phosphatidylinositol (PI) 3-kinase and PI 4-kinase binding to the CD4-p56lck complex: the p56lck SH3 domain binds to PI 3-kinase but not PI 4-kinase. Mol Cell Biol. 1993 Dec;13(12):7708–7717. doi: 10.1128/mcb.13.12.7708. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Punt J. A., Kubo R. T., Saito T., Finkel T. H., Kathiresan S., Blank K. J., Hashimoto Y. Surface expression of a T cell receptor beta (TCR-beta) chain in the absence of TCR-alpha, -delta, and -gamma proteins. J Exp Med. 1991 Oct 1;174(4):775–783. doi: 10.1084/jem.174.4.775. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Reedquist K. A., Fukazawa T., Druker B., Panchamoorthy G., Shoelson S. E., Band H. Rapid T-cell receptor-mediated tyrosine phosphorylation of p120, an Fyn/Lck Src homology 3 domain-binding protein. Proc Natl Acad Sci U S A. 1994 May 10;91(10):4135–4139. doi: 10.1073/pnas.91.10.4135. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Ren R., Mayer B. J., Cicchetti P., Baltimore D. Identification of a ten-amino acid proline-rich SH3 binding site. Science. 1993 Feb 19;259(5098):1157–1161. doi: 10.1126/science.8438166. [DOI] [PubMed] [Google Scholar]
  46. Ren R., Ye Z. S., Baltimore D. Abl protein-tyrosine kinase selects the Crk adapter as a substrate using SH3-binding sites. Genes Dev. 1994 Apr 1;8(7):783–795. doi: 10.1101/gad.8.7.783. [DOI] [PubMed] [Google Scholar]
  47. Rozakis-Adcock M., Fernley R., Wade J., Pawson T., Bowtell D. The SH2 and SH3 domains of mammalian Grb2 couple the EGF receptor to the Ras activator mSos1. Nature. 1993 May 6;363(6424):83–85. doi: 10.1038/363083a0. [DOI] [PubMed] [Google Scholar]
  48. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Seidel-Dugan C., Meyer B. E., Thomas S. M., Brugge J. S. Effects of SH2 and SH3 deletions on the functional activities of wild-type and transforming variants of c-Src. Mol Cell Biol. 1992 Apr;12(4):1835–1845. doi: 10.1128/mcb.12.4.1835. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Shaw A. S., Amrein K. E., Hammond C., Stern D. F., Sefton B. M., Rose J. K. The lck tyrosine protein kinase interacts with the cytoplasmic tail of the CD4 glycoprotein through its unique amino-terminal domain. Cell. 1989 Nov 17;59(4):627–636. doi: 10.1016/0092-8674(89)90008-1. [DOI] [PubMed] [Google Scholar]
  51. Simon M. A., Dodson G. S., Rubin G. M. An SH3-SH2-SH3 protein is required for p21Ras1 activation and binds to sevenless and Sos proteins in vitro. Cell. 1993 Apr 9;73(1):169–177. doi: 10.1016/0092-8674(93)90169-q. [DOI] [PubMed] [Google Scholar]
  52. Skolnik E. Y., Margolis B., Mohammadi M., Lowenstein E., Fischer R., Drepps A., Ullrich A., Schlessinger J. Cloning of PI3 kinase-associated p85 utilizing a novel method for expression/cloning of target proteins for receptor tyrosine kinases. Cell. 1991 Apr 5;65(1):83–90. doi: 10.1016/0092-8674(91)90410-z. [DOI] [PubMed] [Google Scholar]
  53. Smith D. B., Johnson K. S. Single-step purification of polypeptides expressed in Escherichia coli as fusions with glutathione S-transferase. Gene. 1988 Jul 15;67(1):31–40. doi: 10.1016/0378-1119(88)90005-4. [DOI] [PubMed] [Google Scholar]
  54. Straus D. B., Weiss A. Genetic evidence for the involvement of the lck tyrosine kinase in signal transduction through the T cell antigen receptor. Cell. 1992 Aug 21;70(4):585–593. doi: 10.1016/0092-8674(92)90428-f. [DOI] [PubMed] [Google Scholar]
  55. Veillette A., Bookman M. A., Horak E. M., Bolen J. B. The CD4 and CD8 T cell surface antigens are associated with the internal membrane tyrosine-protein kinase p56lck. Cell. 1988 Oct 21;55(2):301–308. doi: 10.1016/0092-8674(88)90053-0. [DOI] [PubMed] [Google Scholar]
  56. Vogel L. B., Fujita D. J. The SH3 domain of p56lck is involved in binding to phosphatidylinositol 3'-kinase from T lymphocytes. Mol Cell Biol. 1993 Dec;13(12):7408–7417. doi: 10.1128/mcb.13.12.7408. [DOI] [PMC free article] [PubMed] [Google Scholar]
  57. Wu H., Reynolds A. B., Kanner S. B., Vines R. R., Parsons J. T. Identification and characterization of a novel cytoskeleton-associated pp60src substrate. Mol Cell Biol. 1991 Oct;11(10):5113–5124. doi: 10.1128/mcb.11.10.5113. [DOI] [PMC free article] [PubMed] [Google Scholar]
  58. Yamanashi Y., Okada M., Semba T., Yamori T., Umemori H., Tsunasawa S., Toyoshima K., Kitamura D., Watanabe T., Yamamoto T. Identification of HS1 protein as a major substrate of protein-tyrosine kinase(s) upon B-cell antigen receptor-mediated signaling. Proc Natl Acad Sci U S A. 1993 Apr 15;90(8):3631–3635. doi: 10.1073/pnas.90.8.3631. [DOI] [PMC free article] [PubMed] [Google Scholar]

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