Skip to main content
Biochemical Journal logoLink to Biochemical Journal
. 1998 Jul 15;333(Pt 2):373–379. doi: 10.1042/bj3330373

Active Lyn protein tyrosine kinase is selectively enriched within membrane microdomains of resting platelets.

D J Dorahy 1, G F Burns 1
PMCID: PMC1219595  PMID: 9657978

Abstract

Circulating platelets are primed to respond very rapidly to thrombogenic stimuli, but most platelets complete their lifespan without ever becoming activated. Platelet activation is accompanied by waves of sequential tyrosine phosphorylation thought to involve members of the Src family of protein tyrosine kinases (PTKs). We show here that resting platelets contain highly active pp53/56(Lyn) PTK within membrane microdomains (rafts) isolated biochemically with or without the use of detergent. This fraction is also greatly enriched in the transmembrane glycoprotein CD36, known to associate with Lyn PTK, but in transfection studies we could find no evidence to suggest that CD36 affects the distribution or function of Lyn. Upon platelet activation Lyn activity remains constant or diminishes and pp60(c-src) PTK within this fraction becomes highly activated, indicating the dynamic nature of the membrane microdomains. It is suggested that the function of active Lyn PTK in the resting platelet is to allow prolonged survival of this anucleate cell.

Full Text

The Full Text of this article is available as a PDF (388.2 KB).

Selected References

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

  1. Aiken M. L., Ginsberg M. H., Byers-Ward V., Plow E. F. Effects of OKM5, a monoclonal antibody to glycoprotein IV, on platelet aggregation and thrombospondin surface expression. Blood. 1990 Dec 15;76(12):2501–2509. [PubMed] [Google Scholar]
  2. Asch A. S., Barnwell J., Silverstein R. L., Nachman R. L. Isolation of the thrombospondin membrane receptor. J Clin Invest. 1987 Apr;79(4):1054–1061. doi: 10.1172/JCI112918. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Barber A. J., Jamieson G. A. Isolation and characterization of plasma membranes from human blood platelets. J Biol Chem. 1970 Dec 10;245(23):6357–6365. [PubMed] [Google Scholar]
  4. 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]
  5. Brown D. The tyrosine kinase connection: how GPI-anchored proteins activate T cells. Curr Opin Immunol. 1993 Jun;5(3):349–354. doi: 10.1016/0952-7915(93)90052-t. [DOI] [PubMed] [Google Scholar]
  6. Dorahy D. J., Berndt M. C., Burns G. F. Capture by chemical crosslinkers provides evidence that integrin alpha IIb beta 3 forms a complex with protein tyrosine kinases in intact platelets. Biochem J. 1995 Jul 15;309(Pt 2):481–490. doi: 10.1042/bj3090481. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Dorahy D. J., Lincz L. F., Meldrum C. J., Burns G. F. Biochemical isolation of a membrane microdomain from resting platelets highly enriched in the plasma membrane glycoprotein CD36. Biochem J. 1996 Oct 1;319(Pt 1):67–72. doi: 10.1042/bj3190067. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Earp H. S., Austin K. S., Buessow S. C., Dy R., Gillespie G. Y. Membranes from T and B lymphocytes have different patterns of tyrosine phosphorylation. Proc Natl Acad Sci U S A. 1984 Apr;81(8):2347–2351. doi: 10.1073/pnas.81.8.2347. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Ferrell J. E., Jr, Martin G. S. Tyrosine-specific protein phosphorylation is regulated by glycoprotein IIb-IIIa in platelets. Proc Natl Acad Sci U S A. 1989 Apr;86(7):2234–2238. doi: 10.1073/pnas.86.7.2234. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Field K. A., Holowka D., Baird B. Compartmentalized activation of the high affinity immunoglobulin E receptor within membrane domains. J Biol Chem. 1997 Feb 14;272(7):4276–4280. doi: 10.1074/jbc.272.7.4276. [DOI] [PubMed] [Google Scholar]
  11. Fra A. M., Williamson E., Simons K., Parton R. G. De novo formation of caveolae in lymphocytes by expression of VIP21-caveolin. Proc Natl Acad Sci U S A. 1995 Sep 12;92(19):8655–8659. doi: 10.1073/pnas.92.19.8655. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Fra A. M., Williamson E., Simons K., Parton R. G. Detergent-insoluble glycolipid microdomains in lymphocytes in the absence of caveolae. J Biol Chem. 1994 Dec 9;269(49):30745–30748. [PubMed] [Google Scholar]
  13. Golden A., Brugge J. S. Thrombin treatment induces rapid changes in tyrosine phosphorylation in platelets. Proc Natl Acad Sci U S A. 1989 Feb;86(3):901–905. doi: 10.1073/pnas.86.3.901. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Gorodinsky A., Harris D. A. Glycolipid-anchored proteins in neuroblastoma cells form detergent-resistant complexes without caveolin. J Cell Biol. 1995 May;129(3):619–627. doi: 10.1083/jcb.129.3.619. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Hibbs M. L., Tarlinton D. M., Armes J., Grail D., Hodgson G., Maglitto R., Stacker S. A., Dunn A. R. Multiple defects in the immune system of Lyn-deficient mice, culminating in autoimmune disease. Cell. 1995 Oct 20;83(2):301–311. doi: 10.1016/0092-8674(95)90171-x. [DOI] [PubMed] [Google Scholar]
  16. Hirao A., Hamaguchi I., Suda T., Yamaguchi N. Translocation of the Csk homologous kinase (Chk/Hyl) controls activity of CD36-anchored Lyn tyrosine kinase in thrombin-stimulated platelets. EMBO J. 1997 May 1;16(9):2342–2351. doi: 10.1093/emboj/16.9.2342. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Huang M. M., Bolen J. B., Barnwell J. W., Shattil S. J., Brugge J. S. Membrane glycoprotein IV (CD36) is physically associated with the Fyn, Lyn, and Yes protein-tyrosine kinases in human platelets. Proc Natl Acad Sci U S A. 1991 Sep 1;88(17):7844–7848. doi: 10.1073/pnas.88.17.7844. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Jacobson M. D., Burne J. F., Raff M. C. Programmed cell death and Bcl-2 protection in the absence of a nucleus. EMBO J. 1994 Apr 15;13(8):1899–1910. doi: 10.1002/j.1460-2075.1994.tb06459.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Krissansen G. W., Lucas C. M., Stomski F. C., Elliott M. J., Berndt M. C., Boyd A. W., Horton M. A., Cheresh D. A., Vadas M. A., Burns G. F. Blood leukocytes bind platelet glycoprotein (IIb-IIIa)' but do not express the vitronectin receptor. Int Immunol. 1990;2(3):267–277. doi: 10.1093/intimm/2.3.267. [DOI] [PubMed] [Google Scholar]
  20. Lisanti M. P., Scherer P. E., Vidugiriene J., Tang Z., Hermanowski-Vosatka A., Tu Y. H., Cook R. F., Sargiacomo M. Characterization of caveolin-rich membrane domains isolated from an endothelial-rich source: implications for human disease. J Cell Biol. 1994 Jul;126(1):111–126. doi: 10.1083/jcb.126.1.111. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Lisanti M. P., Tang Z. L., Sargiacomo M. Caveolin forms a hetero-oligomeric protein complex that interacts with an apical GPI-linked protein: implications for the biogenesis of caveolae. J Cell Biol. 1993 Nov;123(3):595–604. doi: 10.1083/jcb.123.3.595. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Mayor S., Maxfield F. R. Insolubility and redistribution of GPI-anchored proteins at the cell surface after detergent treatment. Mol Biol Cell. 1995 Jul;6(7):929–944. doi: 10.1091/mbc.6.7.929. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Nada S., Yagi T., Takeda H., Tokunaga T., Nakagawa H., Ikawa Y., Okada M., Aizawa S. Constitutive activation of Src family kinases in mouse embryos that lack Csk. Cell. 1993 Jun 18;73(6):1125–1135. doi: 10.1016/0092-8674(93)90642-4. [DOI] [PubMed] [Google Scholar]
  24. Ockenhouse C. F., Magowan C., Chulay J. D. Activation of monocytes and platelets by monoclonal antibodies or malaria-infected erythrocytes binding to the CD36 surface receptor in vitro. J Clin Invest. 1989 Aug;84(2):468–475. doi: 10.1172/JCI114188. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Okada M., Nada S., Yamanashi Y., Yamamoto T., Nakagawa H. CSK: a protein-tyrosine kinase involved in regulation of src family kinases. J Biol Chem. 1991 Dec 25;266(36):24249–24252. [PubMed] [Google Scholar]
  26. Resh M. D. Myristylation and palmitylation of Src family members: the fats of the matter. Cell. 1994 Feb 11;76(3):411–413. doi: 10.1016/0092-8674(94)90104-x. [DOI] [PubMed] [Google Scholar]
  27. Rodgers W., Rose J. K. Exclusion of CD45 inhibits activity of p56lck associated with glycolipid-enriched membrane domains. J Cell Biol. 1996 Dec;135(6 Pt 1):1515–1523. doi: 10.1083/jcb.135.6.1515. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Rothberg K. G., Heuser J. E., Donzell W. C., Ying Y. S., Glenney J. R., Anderson R. G. Caveolin, a protein component of caveolae membrane coats. Cell. 1992 Feb 21;68(4):673–682. doi: 10.1016/0092-8674(92)90143-z. [DOI] [PubMed] [Google Scholar]
  29. Schnitzer J. E., McIntosh D. P., Dvorak A. M., Liu J., Oh P. Separation of caveolae from associated microdomains of GPI-anchored proteins. Science. 1995 Sep 8;269(5229):1435–1439. doi: 10.1126/science.7660128. [DOI] [PubMed] [Google Scholar]
  30. Schoenwaelder S. M., Jackson S. P., Yuan Y., Teasdale M. S., Salem H. H., Mitchell C. A. Tyrosine kinases regulate the cytoskeletal attachment of integrin alpha IIb beta 3 (platelet glycoprotein IIb/IIIa) and the cellular retraction of fibrin polymers. J Biol Chem. 1994 Dec 23;269(51):32479–32487. [PubMed] [Google Scholar]
  31. Simons K., Ikonen E. Functional rafts in cell membranes. Nature. 1997 Jun 5;387(6633):569–572. doi: 10.1038/42408. [DOI] [PubMed] [Google Scholar]
  32. Song K. S., Li Shengwen, Okamoto T., Quilliam L. A., Sargiacomo M., Lisanti M. P. Co-purification and direct interaction of Ras with caveolin, an integral membrane protein of caveolae microdomains. Detergent-free purification of caveolae microdomains. J Biol Chem. 1996 Apr 19;271(16):9690–9697. doi: 10.1074/jbc.271.16.9690. [DOI] [PubMed] [Google Scholar]
  33. Timmons S., Hawiger J. Isolation of human platelets by albumin gradient and gel filtration. Methods Enzymol. 1989;169:11–21. doi: 10.1016/0076-6879(89)69046-5. [DOI] [PubMed] [Google Scholar]
  34. Wei S., Liu J. H., Epling-Burnette P. K., Gamero A. M., Ussery D., Pearson E. W., Elkabani M. E., Diaz J. I., Djeu J. Y. Critical role of Lyn kinase in inhibition of neutrophil apoptosis by granulocyte-macrophage colony-stimulating factor. J Immunol. 1996 Dec 1;157(11):5155–5162. [PubMed] [Google Scholar]
  35. Wu C., Butz S., Ying Y., Anderson R. G. Tyrosine kinase receptors concentrated in caveolae-like domains from neuronal plasma membrane. J Biol Chem. 1997 Feb 7;272(6):3554–3559. doi: 10.1074/jbc.272.6.3554. [DOI] [PubMed] [Google Scholar]
  36. Yan S. R., Fumagalli L., Berton G. Activation of SRC family kinases in human neutrophils. Evidence that p58C-FGR and p53/56LYN redistributed to a Triton X-100-insoluble cytoskeletal fraction, also enriched in the caveolar protein caveolin, display an enhanced kinase activity. FEBS Lett. 1996 Feb 12;380(1-2):198–203. doi: 10.1016/0014-5793(96)00029-4. [DOI] [PubMed] [Google Scholar]
  37. Yousefi S., Hoessli D. C., Blaser K., Mills G. B., Simon H. U. Requirement of Lyn and Syk tyrosine kinases for the prevention of apoptosis by cytokines in human eosinophils. J Exp Med. 1996 Apr 1;183(4):1407–1414. doi: 10.1084/jem.183.4.1407. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES