Skip to main content
PMC home page
The Journal of Clinical Investigation logoLink to The Journal of Clinical Investigation
. 1997 May 1;99(9):2232–2238. doi: 10.1172/JCI119397

Scott syndrome erythrocytes contain a membrane protein capable of mediating Ca2+-dependent transbilayer migration of membrane phospholipids.

J G Stout 1, F Bassé 1, R A Luhm 1, H J Weiss 1, T Wiedmer 1, P J Sims 1
PMCID: PMC508054  PMID: 9151796

Abstract

Phospholipid (PL) scramblase is a plasma membrane protein that mediates accelerated transbilayer migration of PLs upon binding Ca2+, facilitating rapid mobilization of phosphatidylserine to the cell surface upon elevation of internal Ca2+. In patients with Scott syndrome, a congenital bleeding disorder related to defective expression of membrane coagulant activity, circulating blood cells show decreased cell surface exposure of phosphatidylserine at elevated cytosolic [Ca2+], implying an underlying defect or deficiency of PL scramblase. To gain insight into the molecular basis of this disorder, we compared PL scramblase in Scott erythrocyte membranes to those of normal controls. Whereas membranes of Scott cells were unresponsive to Ca2+-induced activation of PL scramblase at neutral pH, apparently normal PL scramblase activity was induced at pH < 6.0. After extraction with octylglucoside, a membrane protein was isolated from the Scott cells which exhibited normal PL scramblase activity when reconstituted in vesicles with exogenous PLs. Like PL scramblase from normal erythrocytes, PL scramblase from Scott erythrocytes was maximally activated either by addition of Ca2+ (at pH 7.4) or by acidification to pH < 6.0, and similar apparent affinities for Ca2+ and rates of transbilayer transfer of PLs were observed. This suggests that the defect in Scott syndrome is related to an altered interaction of Ca2+ with PL scramblase on the endofacial surface of the cell membrane, due either to an intrinsic constraint upon the protein preventing interaction with Ca2+ in situ, or due to an unidentified inhibitor or cofactor in the Scott cell that is dissociated by detergent.

Full Text

The Full Text of this article is available as a PDF (222.0 KB).

Selected References

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

  1. Ahmad S. S., Rawala-Sheikh R., Ashby B., Walsh P. N. Platelet receptor-mediated factor X activation by factor IXa. High-affinity factor IXa receptors induced by factor VIII are deficient on platelets in Scott syndrome. J Clin Invest. 1989 Sep;84(3):824–828. doi: 10.1172/JCI114242. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Auland M. E., Roufogalis B. D., Devaux P. F., Zachowski A. Reconstitution of ATP-dependent aminophospholipid translocation in proteoliposomes. Proc Natl Acad Sci U S A. 1994 Nov 8;91(23):10938–10942. doi: 10.1073/pnas.91.23.10938. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bassé F., Gaffet P., Rendu F., Bienvenüe A. Translocation of spin-labeled phospholipids through plasma membrane during thrombin- and ionophore A23187-induced platelet activation. Biochemistry. 1993 Mar 9;32(9):2337–2344. doi: 10.1021/bi00060a027. [DOI] [PubMed] [Google Scholar]
  4. Bassé F., Stout J. G., Sims P. J., Wiedmer T. Isolation of an erythrocyte membrane protein that mediates Ca2+-dependent transbilayer movement of phospholipid. J Biol Chem. 1996 Jul 19;271(29):17205–17210. doi: 10.1074/jbc.271.29.17205. [DOI] [PubMed] [Google Scholar]
  5. Bevers E. M., Comfurius P., Zwaal R. F. Platelet procoagulant activity: physiological significance and mechanisms of exposure. Blood Rev. 1991 Sep;5(3):146–154. doi: 10.1016/0268-960x(91)90031-7. [DOI] [PubMed] [Google Scholar]
  6. Bevers E. M., Wiedmer T., Comfurius P., Shattil S. J., Weiss H. J., Zwaal R. F., Sims P. J. Defective Ca(2+)-induced microvesiculation and deficient expression of procoagulant activity in erythrocytes from a patient with a bleeding disorder: a study of the red blood cells of Scott syndrome. Blood. 1992 Jan 15;79(2):380–388. [PubMed] [Google Scholar]
  7. Bevers E. M., Wiedmer T., Comfurius P., Zhao J., Smeets E. F., Schlegel R. A., Schroit A. J., Weiss H. J., Williamson P., Zwaal R. F. The complex of phosphatidylinositol 4,5-bisphosphate and calcium ions is not responsible for Ca2+-induced loss of phospholipid asymmetry in the human erythrocyte: a study in Scott syndrome, a disorder of calcium-induced phospholipid scrambling. Blood. 1995 Sep 1;86(5):1983–1991. [PubMed] [Google Scholar]
  8. Bratton D. L. Polyamine inhibition of transbilayer movement of plasma membrane phospholipids in the erythrocyte ghost. J Biol Chem. 1994 Sep 9;269(36):22517–22523. [PubMed] [Google Scholar]
  9. Chang C. P., Zhao J., Wiedmer T., Sims P. J. Contribution of platelet microparticle formation and granule secretion to the transmembrane migration of phosphatidylserine. J Biol Chem. 1993 Apr 5;268(10):7171–7178. [PubMed] [Google Scholar]
  10. Comfurius P., Senden J. M., Tilly R. H., Schroit A. J., Bevers E. M., Zwaal R. F. Loss of membrane phospholipid asymmetry in platelets and red cells may be associated with calcium-induced shedding of plasma membrane and inhibition of aminophospholipid translocase. Biochim Biophys Acta. 1990 Jul 24;1026(2):153–160. doi: 10.1016/0005-2736(90)90058-v. [DOI] [PubMed] [Google Scholar]
  11. Comfurius P., Williamson P., Smeets E. F., Schlegel R. A., Bevers E. M., Zwaal R. F. Reconstitution of phospholipid scramblase activity from human blood platelets. Biochemistry. 1996 Jun 18;35(24):7631–7634. doi: 10.1021/bi9606859. [DOI] [PubMed] [Google Scholar]
  12. Connor J., Schroit A. J. Determination of lipid asymmetry in human red cells by resonance energy transfer. Biochemistry. 1987 Aug 11;26(16):5099–5105. doi: 10.1021/bi00390a031. [DOI] [PubMed] [Google Scholar]
  13. Dachary-Prigent J., Pasquet J. M., Freyssinet J. M., Nurden A. T. Calcium involvement in aminophospholipid exposure and microparticle formation during platelet activation: a study using Ca2+-ATPase inhibitors. Biochemistry. 1995 Sep 12;34(36):11625–11634. doi: 10.1021/bi00036a039. [DOI] [PubMed] [Google Scholar]
  14. Devaux P. F. Static and dynamic lipid asymmetry in cell membranes. Biochemistry. 1991 Feb 5;30(5):1163–1173. doi: 10.1021/bi00219a001. [DOI] [PubMed] [Google Scholar]
  15. Fadok V. A., Voelker D. R., Campbell P. A., Cohen J. J., Bratton D. L., Henson P. M. Exposure of phosphatidylserine on the surface of apoptotic lymphocytes triggers specific recognition and removal by macrophages. J Immunol. 1992 Apr 1;148(7):2207–2216. [PubMed] [Google Scholar]
  16. Fox J. E., Reynolds C. C., Austin C. D. The role of calpain in stimulus-response coupling: evidence that calpain mediates agonist-induced expression of procoagulant activity in platelets. Blood. 1990 Dec 15;76(12):2510–2519. [PubMed] [Google Scholar]
  17. Gaffet P., Bettache N., Bienvenüe A. Transverse redistribution of phospholipids during human platelet activation: evidence for a vectorial outflux specific to aminophospholipids. Biochemistry. 1995 May 23;34(20):6762–6769. doi: 10.1021/bi00020a022. [DOI] [PubMed] [Google Scholar]
  18. Gorren A. C., Duine J. A. The effects of pH and cations on the spectral and kinetic properties of methylamine dehydrogenase from Thiobacillus versutus. Biochemistry. 1994 Oct 11;33(40):12202–12209. doi: 10.1021/bi00206a024. [DOI] [PubMed] [Google Scholar]
  19. Kojima H., Newton-Nash D., Weiss H. J., Zhao J., Sims P. J., Wiedmer T. Production and characterization of transformed B-lymphocytes expressing the membrane defect of Scott syndrome. J Clin Invest. 1994 Dec;94(6):2237–2244. doi: 10.1172/JCI117586. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Lamb J. A., Allen P. G., Tuan B. Y., Janmey P. A. Modulation of gelsolin function. Activation at low pH overrides Ca2+ requirement. J Biol Chem. 1993 Apr 25;268(12):8999–9004. [PubMed] [Google Scholar]
  21. McIntyre J. C., Sleight R. G. Fluorescence assay for phospholipid membrane asymmetry. Biochemistry. 1991 Dec 24;30(51):11819–11827. doi: 10.1021/bi00115a012. [DOI] [PubMed] [Google Scholar]
  22. Miletich J. P., Kane W. H., Hofmann S. L., Stanford N., Majerus P. W. Deficiency of factor Xa-factor Va binding sites on the platelets of a patient with a bleeding disorder. Blood. 1979 Nov;54(5):1015–1022. [PubMed] [Google Scholar]
  23. Pomorski T., Herrmann A., Zachowski A., Devaux P. F., Müller P. Rapid determination of the transbilayer distribution of NBD-phospholipids in erythrocyte membranes with dithionite. Mol Membr Biol. 1994 Jan-Mar;11(1):39–44. doi: 10.3109/09687689409161028. [DOI] [PubMed] [Google Scholar]
  24. Schroit A. J., Zwaal R. F. Transbilayer movement of phospholipids in red cell and platelet membranes. Biochim Biophys Acta. 1991 Nov 13;1071(3):313–329. doi: 10.1016/0304-4157(91)90019-s. [DOI] [PubMed] [Google Scholar]
  25. Sims P. J., Faioni E. M., Wiedmer T., Shattil S. J. Complement proteins C5b-9 cause release of membrane vesicles from the platelet surface that are enriched in the membrane receptor for coagulation factor Va and express prothrombinase activity. J Biol Chem. 1988 Dec 5;263(34):18205–18212. [PubMed] [Google Scholar]
  26. Sims P. J., Wiedmer T., Esmon C. T., Weiss H. J., Shattil S. J. Assembly of the platelet prothrombinase complex is linked to vesiculation of the platelet plasma membrane. Studies in Scott syndrome: an isolated defect in platelet procoagulant activity. J Biol Chem. 1989 Oct 15;264(29):17049–17057. [PubMed] [Google Scholar]
  27. Smeets E. F., Comfurius P., Bevers E. M., Zwaal R. F. Calcium-induced transbilayer scrambling of fluorescent phospholipid analogs in platelets and erythrocytes. Biochim Biophys Acta. 1994 Nov 2;1195(2):281–286. doi: 10.1016/0005-2736(94)90268-2. [DOI] [PubMed] [Google Scholar]
  28. Steck T. L., Kant J. A. Preparation of impermeable ghosts and inside-out vesicles from human erythrocyte membranes. Methods Enzymol. 1974;31:172–180. doi: 10.1016/0076-6879(74)31019-1. [DOI] [PubMed] [Google Scholar]
  29. Sulpice J. C., Moreau C., Devaux P. F., Zachowski A., Giraud F. Antagonist effects of Ca2+ and spermine on phosphatidylinositol 4,5-bisphosphate-mediated transmembrane redistribution of phospholipids in large unilamellar vesicles and in erythrocytes. Biochemistry. 1996 Oct 15;35(41):13345–13352. doi: 10.1021/bi960624a. [DOI] [PubMed] [Google Scholar]
  30. Sulpice J. C., Zachowski A., Devaux P. F., Giraud F. Requirement for phosphatidylinositol 4,5-bisphosphate in the Ca(2+)-induced phospholipid redistribution in the human erythrocyte membrane. J Biol Chem. 1994 Mar 4;269(9):6347–6354. [PubMed] [Google Scholar]
  31. Tang X., Halleck M. S., Schlegel R. A., Williamson P. A subfamily of P-type ATPases with aminophospholipid transporting activity. Science. 1996 Jun 7;272(5267):1495–1497. doi: 10.1126/science.272.5267.1495. [DOI] [PubMed] [Google Scholar]
  32. Toti F., Satta N., Fressinaud E., Meyer D., Freyssinet J. M. Scott syndrome, characterized by impaired transmembrane migration of procoagulant phosphatidylserine and hemorrhagic complications, is an inherited disorder. Blood. 1996 Feb 15;87(4):1409–1415. [PubMed] [Google Scholar]
  33. Wang R. H., Phillips G., Jr, Medof M. E., Mold C. Activation of the alternative complement pathway by exposure of phosphatidylethanolamine and phosphatidylserine on erythrocytes from sickle cell disease patients. J Clin Invest. 1993 Sep;92(3):1326–1335. doi: 10.1172/JCI116706. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Weiss H. J. Scott syndrome: a disorder of platelet coagulant activity. Semin Hematol. 1994 Oct;31(4):312–319. [PubMed] [Google Scholar]
  35. Weiss H. J., Vicic W. J., Lages B. A., Rogers J. Isolated deficiency of platelet procoagulant activity. Am J Med. 1979 Aug;67(2):206–213. doi: 10.1016/0002-9343(79)90392-9. [DOI] [PubMed] [Google Scholar]
  36. Williamson P., Bevers E. M., Smeets E. F., Comfurius P., Schlegel R. A., Zwaal R. F. Continuous analysis of the mechanism of activated transbilayer lipid movement in platelets. Biochemistry. 1995 Aug 22;34(33):10448–10455. doi: 10.1021/bi00033a017. [DOI] [PubMed] [Google Scholar]
  37. Williamson P., Kulick A., Zachowski A., Schlegel R. A., Devaux P. F. Ca2+ induces transbilayer redistribution of all major phospholipids in human erythrocytes. Biochemistry. 1992 Jul 14;31(27):6355–6360. doi: 10.1021/bi00142a027. [DOI] [PubMed] [Google Scholar]

Articles from The Journal of Clinical Investigation are provided here courtesy of American Society for Clinical Investigation

RESOURCES