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. 1984 Jan 1;98(1):8–15. doi: 10.1083/jcb.98.1.8

Recycling of platelet phosphorylation and cytoskeletal assembly

PMCID: PMC2112981  PMID: 6423649

Abstract

The shape change and aggregation of washed platelets induced by 10 microM arachidonic acid (AA) can be reversed by 20 ng/ml prostacyclin (PGI2), but these platelets can be reactivated by treatment with 30 microM epinephrine and subsequent addition of 10 microM AA mixture. These events may be modulated by cAMP since 2 mM dibutyryl cAMP also reversed activation without reactivation by epinephrine and AA. We examined protein phosphorylation and formation of cytoskeletal cores resistant to 1% Triton X-100 extraction of these platelets and correlated these processes with aggregation, fibrinogen binding, and changes in ultrastructure. Unactivated platelet cores contained less than 15% of the total actin and no detectable myosin or actin-binding protein. AA-induced cytoskeletal cores, which contained 60-80% of the total actin, myosin, and actin-binding protein as the major components, were disassembled back to unactivated levels by PGI2 and then fully reassembled by epinephrine and AA. Phosphorylation of myosin light chain and a 40,000-dalton protein triggered by AA (two- to fivefold) was reversed to basal levels by PGI2 but was completely restored to peak levels upon addition of the epinephrine and AA mixture. The reversibility of actin-binding protein phosphorylation could not be established clearly because both PGI2 and dibutyryl cAMP caused its phosphorylation independent of activation. With this possible exception, cytoskeletal assembly with associated protein phosphorylation, aggregation, fibrinogen binding, and changes in ultrastructure triggered by activation are readily and concertedly recyclable.

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Selected References

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

  1. Bennett J. S., Vilaire G., Cines D. B. Identification of the fibrinogen receptor on human platelets by photoaffinity labeling. J Biol Chem. 1982 Jul 25;257(14):8049–8054. [PubMed] [Google Scholar]
  2. Blikstad I., Markey F., Carlsson L., Persson T., Lindberg U. Selective assay of monomeric and filamentous actin in cell extracts, using inhibition of deoxyribonuclease I. Cell. 1978 Nov;15(3):935–943. doi: 10.1016/0092-8674(78)90277-5. [DOI] [PubMed] [Google Scholar]
  3. Bolton A. E., Hunter W. M. The labelling of proteins to high specific radioactivities by conjugation to a 125I-containing acylating agent. Biochem J. 1973 Jul;133(3):529–539. doi: 10.1042/bj1330529. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Carlsson L., Markey F., Blikstad I., Persson T., Lindberg U. Reorganization of actin in platelets stimulated by thrombin as measured by the DNase I inhibition assay. Proc Natl Acad Sci U S A. 1979 Dec;76(12):6376–6380. doi: 10.1073/pnas.76.12.6376. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Carroll R. C., Butler R. G., Morris P. A., Gerrard J. M. Separable assembly of platelet pseudopodal and contractile cytoskeletons. Cell. 1982 Sep;30(2):385–393. doi: 10.1016/0092-8674(82)90236-7. [DOI] [PubMed] [Google Scholar]
  6. Carroll R. C., Gerrard J. M. Phosphorylation of platelet actin-binding protein during platelet activation. Blood. 1982 Mar;59(3):466–471. [PubMed] [Google Scholar]
  7. Conti M. A., Adelstein R. S. The relationship between calmodulin binding and phosphorylation of smooth muscle myosin kinase by the catalytic subunit of 3':5' cAMP-dependent protein kinase. J Biol Chem. 1981 Apr 10;256(7):3178–3181. [PubMed] [Google Scholar]
  8. Cox A. C., Inyangetor P., Esmon C. T., White B. N. The effect of aggregation and release on platelet prothrombin-converting activity. Blood. 1979 Sep;54(3):659–672. [PubMed] [Google Scholar]
  9. Daniel J. L., Molish I. R., Holmsen H. Myosin phosphorylation in intact platelets. J Biol Chem. 1981 Jul 25;256(14):7510–7514. [PubMed] [Google Scholar]
  10. Davies P. J., Wallach D., Willingham M. C., Pastan I., Yamaguchi M., Robson R. M. Filamin-actin interaction. Dissociation of binding from gelation by Ca2+-activated proteolysis. J Biol Chem. 1978 Jun 10;253(11):4036–4042. [PubMed] [Google Scholar]
  11. Feinstein M. B., Egan J. J., Opas E. E. Reversal of thrombin-induced myosin phosphorylation and the assembly of cytoskeletal structures in platelets by the adenylate cyclase stimulants prostaglandin D2 and forskolin. J Biol Chem. 1983 Jan 25;258(2):1260–1267. [PubMed] [Google Scholar]
  12. Fox J. E., Say A. K., Haslam R. J. Subcellular distribution of the different platelet proteins phosphorylated on exposure of intact platelets to ionophore A23187 or to prostaglandin E1. Possible role of a membrane phosphopolypeptide in the regulation of calcium-ion transport. Biochem J. 1979 Dec 15;184(3):651–661. doi: 10.1042/bj1840651. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Gerrard J. M., Carroll R. C. Stimulation of platelet protein phosphorylation by arachidonic acid and endoperoxide analogs. Prostaglandins. 1981 Jul;22(1):81–94. doi: 10.1016/0090-6980(81)90055-1. [DOI] [PubMed] [Google Scholar]
  14. Gerrard J. M., Phillips D. R., Rao G. H., Plow E. F., Walz D. A., Ross R., Harker L. A., White J. G. Biochemical studies of two patients with the gray platelet syndrome. Selective deficiency of platelet alpha granules. J Clin Invest. 1980 Jul;66(1):102–109. doi: 10.1172/JCI109823. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Hathaway D. R., Adelstein R. S. Human platelet myosin light chain kinase requires the calcium-binding protein calmodulin for activity. Proc Natl Acad Sci U S A. 1979 Apr;76(4):1653–1657. doi: 10.1073/pnas.76.4.1653. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hawiger J., Parkinson S., Timmons S. Prostacyclin inhibits mobilisation of fibrinogen-binding sites on human ADP- and thrombin-treated platelets. Nature. 1980 Jan 10;283(5743):195–197. doi: 10.1038/283195a0. [DOI] [PubMed] [Google Scholar]
  17. Lebowitz E. A., Cooke R. Contractile properties of actomyosin from human blood platelets. J Biol Chem. 1978 Aug 10;253(15):5443–5447. [PubMed] [Google Scholar]
  18. Lyons R. M., Atherton R. M. Characterization of a platelet protein phosphorylated during the thrombin-induced release reaction. Biochemistry. 1979 Feb 6;18(3):544–552. doi: 10.1021/bi00570a025. [DOI] [PubMed] [Google Scholar]
  19. Lyons R. M., Stanford N., Majerus P. W. Thrombin-induced protein phosphorylation in human platelets. J Clin Invest. 1975 Oct;56(4):924–936. doi: 10.1172/JCI108172. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Mihalyi E. Physicochemical studies of bovine fibrinogen. IV. Ultraviolet absorption and its relation to the structure of the molecule. Biochemistry. 1968 Jan;7(1):208–223. doi: 10.1021/bi00841a026. [DOI] [PubMed] [Google Scholar]
  21. Mustard J. F., Perry D. W., Kinlough-Rathbone R. L., Packham M. A. Factors responsible for ADP-induced release reaction of human platelets. Am J Physiol. 1975 Jun;228(6):1757–1765. doi: 10.1152/ajplegacy.1975.228.6.1757. [DOI] [PubMed] [Google Scholar]
  22. Peerschke E. I., Zucker M. B., Grant R. A., Egan J. J., Johnson M. M. Correlation between fibrinogen binding to human platelets and platelet aggregability. Blood. 1980 May;55(5):841–847. [PubMed] [Google Scholar]
  23. Phillips D. R., Jennings L. K., Edwards H. H. Identification of membrane proteins mediating the interaction of human platelets. J Cell Biol. 1980 Jul;86(1):77–86. doi: 10.1083/jcb.86.1.77. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Rao G. H., Reddy K. R., White J. G. The influence of epinephrine on prostacyclin (PGI2) induced dissociation of ADP aggregated plateletes. Prostaglandins Med. 1980 Jun;4(6):385–397. doi: 10.1016/0161-4630(80)90047-6. [DOI] [PubMed] [Google Scholar]
  25. Studier F. W. Analysis of bacteriophage T7 early RNAs and proteins on slab gels. J Mol Biol. 1973 Sep 15;79(2):237–248. doi: 10.1016/0022-2836(73)90003-x. [DOI] [PubMed] [Google Scholar]
  26. White J. G. Fine structural alterations induced in platelets by adenosine diphosphate. Blood. 1968 May;31(5):604–622. [PubMed] [Google Scholar]
  27. White J. G., Krivit W. An ultrastructural basis for the shape changes induced in platelets by chilling. Blood. 1967 Nov;30(5):625–635. [PubMed] [Google Scholar]

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