Download PDF
Thromb Haemost 2004; 92(05): 1040-1051
DOI: 10.1160/TH04-03-0170
Platelets and Blood Cells
Schattauer GmbH

Glanzmann thrombasthenia Frankfurt I is associated with a point mutation Thr176Ile in the N-terminal region of αIIb subunit integrin

Dagmar Westrup
1   Deutsche Klinik für Diagnostik, Wiesbaden, Germany
,
Sentot Santoso
2   Institute for Clinical Immunology and Transfusion Medicine, Justus Liebig University, Giessen, Germany
,
Katja Follert-Hagendorff
1   Deutsche Klinik für Diagnostik, Wiesbaden, Germany
,
Steffen Bassus
1   Deutsche Klinik für Diagnostik, Wiesbaden, Germany
,
Melitta Just
3   Aventis, Frankfurt-Hoechst, Germany
,
Bernd Jablonka
3   Aventis, Frankfurt-Hoechst, Germany
,
Carl M. Kirchmaier
1   Deutsche Klinik für Diagnostik, Wiesbaden, Germany
› Author AffiliationsFinancial support: This work has been supported, in part, by a grant from the Deutsche Forschungsgemeinschaft, Bonn (DFG Ki 1-1 and SFB547 (SS)).
Further Information

Publication History

Received 18 March 2004

Accepted after revision 25 August 2004

Publication Date:
04 December 2017 (online)

    Permissions and Reprints

Summary

In this study, we report on the characterization of a patient with Glanzmann thrombasthenia (GT). Immunochemical analysis on platelets from the patient showed that the expression of αIIbβ3 was only 25% of that in normal healthy controls, suggesting a case of GT. Functional analysis revealed a total lack of fibrinogen binding capacity. Molecular genetic analysis of the full-length cDNA sequences of αIIb and β3 subunits showed a novel point mutation C621T in αIIb cDNA, leading to a missense substitution of threonine for isoleucine at position 176. Coexpression of normal β3 and mutant αIIbI176 isoform in mammalian cells showed a marked reduction in the expression of αIIbβ3 heterodimer when compared to the wild-type and a decreased intracellular level of αIIb. The T176 I mutation is located in the N-terminal region in the W3:1-2 connecting strand of the β-propeller. These data suggest that the N-terminal αIIb domain plays an important structural role in the formation of heterodimer and that it is also involved in fibrinogen binding.

Keywords

Bleeding disorder - platelet - integrin - β-propeller
 

  • References

  • 1 Phillips DR, Charo IF, Parise LV. et al. The platelet membrane glycoprotein IIb-IIIa complex. Blood 1988; 71: 831-43.
    PubMedGoogle Scholar
  • 2 Ginsberg MH, Xiaoping D, Otool TE. et al. Platelet integrins. Thromb Haemost 1993; 70: 87-93.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 3 Calvete JJ. Clues for understanding the structure and function of a prototypic human integrin.: the platelet glycoprotein IIb/IIIa complex. Thromb Haemost 1994; 72: 1-15.
    PubMedGoogle Scholar
  • 4 Wager CL, Mascelli MA, Neblock DS. et al. Analysis of GPIIb-IIIa receptor number by quantification of 7E3 binding to human platelets. Blood 1996; 88: 907.
    PubMedGoogle Scholar
  • 5 Georg JN, Caen JP, Nurden AT. Glanzmann´s thrombasthenia: the spectrum of clinical disease. Blood 1990; 75: 1383-95.
    PubMedGoogle Scholar
  • 6 Coller BS, Seligsohn U, Peretz H. et al. Glanzmann thrombasthenia: new insights from an historical perspective. Sem Hematol 1994; 31: 301-11.
    PubMedGoogle Scholar
  • 7 Shattil SJ, Kashiwagi H, Pampori N. Integrin signaling: The platelet paradigm. Blood 1998; 91: 2645-57.
    PubMedGoogle Scholar
  • 8 D´Souza SE, Ginsberg MH, Burke TA. et al. Localization of an Arg-GlyAsp recognition site within an integrin adhesion receptor. Science 1988; 242: 91-3.
    CrossrefPubMedGoogle Scholar
  • 9 Farrell DH, Thiagarajan P, Chung DW. et al. Role of fibrinogen α an γ chain sites in platelet aggregation. Proc Natl Acad Sci USA 1992; 89: 10729-32.
    CrossrefPubMedGoogle Scholar
  • 10 D´Souza SE, Ginsberg MH, Burke TA. et al. Localization of an Arg-Gly-Asp recognition site within an integrin adhesion receptor. Science 1988; 242: 91-3.
    CrossrefPubMedGoogle Scholar
  • 11 Loftus JC, Smith JW, Ginsberg MH. Integrinmediated cell adhesion: the extracellular face. J Biol Chem 1994; 269: 25235-8.
    PubMedGoogle Scholar
  • 12 Loftus JC, O´Toole TE, Plow EF. et al. A β3 integrin mutation abolishes ligand binding and alters divalent cation-dependent conformation. Science 1990; 249: 915-18.
    CrossrefPubMedGoogle Scholar
  • 13 D´Souza SE, Ginsberg MH, Burke TA. et al. The ligand binding site of the platelet integrin receptor GPIIb-IIIa is proximal to the second calcium binding domain of ist α subunit. J Biol Chem 1990; 265: 3440-6.
    PubMedGoogle Scholar
  • 14 Kamata T, Irie A, Tokuhira M. et al. Critical residues of integrin αIIbβ3 (glycoprotein IIb-IIIa) to fibrinogen binding and ligand-mimetic anti- bodies (PAC-1, OP-G2 and LJ-CP3). J Biol Chem 1996; 271: 18610-15.
    CrossrefPubMedGoogle Scholar
  • 15 Calvete JJ. Platelet integrin GPIIb/IIIa: structure-function correlations. An update and lessons from other integrins. Proc Soc Exp Biol Med 1999; 222: 29-38.
    CrossrefPubMedGoogle Scholar
  • 16 Adair BD, Yeager M. Three-dimensional model of the human platelet integrin αIIbß3 based on electron cryomicroscopy and x-ray crystallography. PNAS 2002; 99: 14059-64.
    CrossrefPubMedGoogle Scholar
  • 17 Feuston BP, Culberson JC, Hartman GD. Molecular model of the αIIbß3 integrin. J Med Chem 2003; 46: 5316-25.
    CrossrefPubMedGoogle Scholar
  • 18 French DL, Coller BS. Hematologically important mutations: Glanzmann thrombasthenia. blood cells, molecules and diseases. 1997 23. 39-51 GT Database: ttp://med.mssm.edu/glanzmanndb
    PubMedGoogle Scholar
  • 19 Puzon-McLaughlin W, Kamata T, Takada Y. Multiple discontinous ligand-mimetic antibody binding sites define a ligand binding pockelt in integrin αIIbß3. J Biol Chem 2000; 275: 7795-802.
    CrossrefPubMedGoogle Scholar
  • 20 French DL, Seligsohn U. Platelet glycoprotein IIb/IIIa receptors and Glannzmann´s thrombasthenia. Arterioscler Thromb Vasc Biol 2000; 20: 607-10.
    CrossrefPubMedGoogle Scholar
  • 21 Springer TA. Folding of the N-terminal, ligandbinding region of integrin α-subunits into a ßpropeller domain. Proc Natl Acad Sci USA 1997; 94: 65-72.
    CrossrefPubMedGoogle Scholar
  • 22 Basani BB, French DL, Vilaire G. et al. A naturally occurring mutation near the amino terminus of αIIb defines a new region involved in ligand binding to αIIbß3. Blood 2000; 95: 180-8.
    PubMedGoogle Scholar
  • 23 Santoso S, Kalb R, Kroll H. et al. A point mutation leads to an unpaired cysteine residue and a molecular weight polymorphism of a functional platelet beta 3 integrin subunit. The Sra alloantigen system of GPIIIa. J Biol Chem 1994; 269: 8439-44.
    PubMedGoogle Scholar
  • 24 Breddin HK. Zur Messung der Thrombozytenadhäsivität. Thromb Diath Haemorrh 1964; 12: 269-80.
    PubMedGoogle Scholar
  • 25 Jaffe EA, Nachman RC, Becker CG. et al. Culture of human endothelial cells derived from umbilical veins. J Clin Invest 1973; 52: 2745-56.
    CrossrefPubMedGoogle Scholar
  • 26 Marguerie GA, Plow EF, Edgington TS. Human platelets possess an inducible and saturable receptor specific for fibrinogen. J Biol Chem 1979; 254: 5357-63.
    PubMedGoogle Scholar
  • 27 Djaffar I, Vilette D, Bray PF. et al. Quantitative isolation of RNA from human platelets. Thromb Res 1991; 62: 127-35.
    CrossrefPubMedGoogle Scholar
  • 28 Lanza F, Stierle A, Fournier D. et al. A new variant of Glanzmann´s thrombasthenia (Strasbourg I). Platelets with functionally defective glycoprotein IIb-IIIa complexes and a glycoprotein IIIa 214Arg→ 214 Trp mutation, J Clin Invest 1992; 89: 1995-2004.
    PubMedGoogle Scholar
  • 29 Santoso S, Amrhein J, Hofmann HA. et al. Point mutation Thr(799)Met on the alpha(2) integrin leads to the formation of new human platelet alloantigen Sit(a) and affects collageninduced aggregation. Blood 1999; 94: 4103-11.
    PubMedGoogle Scholar
  • 30 Harrison P, Wilbourn BR, Saundry RH. et al. Absence of the gamma-Leu 427 (gamma’) variant in the platelet alpha-granular fibrinogen pool supports the role of glycoprotein IIb/IIIa in mediating fibrinogen uptake into platelets/megakaryocytes. Blood 1992; 79: 3394-5.
    PubMedGoogle Scholar
  • 31 Kahn MJ, Kieber-Emmons T, Vilaire G. et al. Effect of mutagenesis of GPIIb amino acid 273 on the expression and conformation of the platelet integrin GPIIb-IIIa. Biochemistry 1996; 35: 14304-11.
    CrossrefPubMedGoogle Scholar
  • 32 Grimaldi CM, Chen FP, Wu CH. et al. Glycoprotein IIb Leu214Pro mutation produces Glanzmann thrombasthenia with both quantitative and qualitative abnormalities in GPIIb/IIIa. Blood 1998; 91: 1562-71.
    PubMedGoogle Scholar
  • 33 Kiyoi T, Tomoyama Y, Honda S. et al. A naturally occurring Tyr143HisαIIb mutation abolishes αIIbβ3 function for soluble ligands but retains its ability for mediating cell adhesion and clot retraction: comparison with other mutations causing ligand-binding defects. Blood 2003; 101: 3485-91.
    CrossrefPubMedGoogle Scholar
  • 34 Vinciguerra C, Khelif A, Alemany M. et al. A nonsense mutation in the GPIIb heavy chain (Ser 870 stop) impairs platelet GP IIb-IIIa expression. Br J Hematol 1996; 95: 399-407.
    CrossrefPubMedGoogle Scholar
  • 35 Mitchell WB, Li JH, Singh F. et al. Two novel mutations in the αIIb calcium-binding domains identify hydrophobic regions essential for αIIbβ3 biogenesis. Blood 2003; 101: 2268-76.
    CrossrefPubMedGoogle Scholar
  • 36 Springer TA. Folding of the N-terminal, ligandbinding region of integrin α-subunits into a ßpropeller domain. Proc Natl Acad Sci USA 1997; 94: 65-72.
    CrossrefPubMedGoogle Scholar
  • 37 Oxvig C, Springer TA. Experimental support for a ß-propeller domain in integrin alpha-subunits and a calcium binding site on its lower surface. Proc Natl Acad Sci USA 1998; 95: 4870-5.
    CrossrefPubMedGoogle Scholar
  • 38 Lu C, Oxvig C, Springer TA. The structure of the beta-propeller domain and C-terminal region of the integrin alphaM subunit: dependence on beta subunit association and prediction of domains. J Biol Chem 1998; 273: 15138-47.
    CrossrefPubMedGoogle Scholar
  • 39 Xiong JP, Stehle T, Diefenbach B. et al. Crystal structure of the extracellular segment of integrin alpha V beta3. Science 2001; 294: 339-45.
    CrossrefPubMedGoogle Scholar
  • 40 Xiong JP, Stehle T, Zhang R. et al. Crystal structure of the extracellular segment of integrin alpha Vbeta3 in complex with an Arg-Gly-Asp ligand. Science 2002; 296: 151-5.
    CrossrefPubMedGoogle Scholar
  • 41 Honda S, Tomiyama Y, Pampori N. et al. Ligand binding to integrin alpha(v)beta(3) requires tyrosine 178 in the alpha(v) subunit. Blood 2001; 97: 175-82.
    CrossrefPubMedGoogle Scholar
  • 42 Kamata T, Tieu KK, Irie A. et al. Amino acid residues in the alpha IIb subunit that are critical for ligand binding to integrin alpha IIbbeta 3 are clustered in the beta-propeller model. J Biol Chem 2001; 276: 44275-83.
    CrossrefPubMedGoogle Scholar
  • 43 Poncz M, Rifat S, Coller BS. et al. Glanzmann thrombasthenia secondary to a Gly273Asp mutation adjacent to the first calcium-binding domain of platelet glycoprotein IIb. J Clin Invest 1994; 93: 172-9.
    CrossrefPubMedGoogle Scholar
  • 44 Wilcox DA, Wautier JL, Pidard D. et al. A single amino acid substitution flanking the fourth calcium binding domain of alpha IIb prevents maturation of the integrin alpha IIb beta 3 complex. J Biol Chem 1994; 269: 4450-7.
    PubMedGoogle Scholar
  • 45 Basani RB, Vilaire G, Shattil SJ. et al. Glanzmann thrombasthenia due to a two amino acid deletion in the fourth calcium-binding domain of alpha IIb: Demonstration of the importance of calciumbinding domains in the conformation of alpha IIb. Blood 1996; 88: 167-73.
    PubMedGoogle Scholar
  • 46 Tozer EC, Baker EK, Ginsberg MH. et al. A mutation in the α subunit of the platelet αIIbß3 identifies a novel region important for ligand binding. Blood 1999; 93: 918-24.
    PubMedGoogle Scholar
  • 47 D´Souza SE, Ginsberg MH, Burke TA. et al. The ligand binding site of the platelet integrin receptor GPIIb-IIIa is proximal to the second calcium binding domain of its alpha subunit. J Biol Chem 1990; 265: 3440-6.
    PubMedGoogle Scholar
  • 48 Loftus JC, Halloran CE, Ginsberg MH. et al. The amino-terminal one-third of alpha IIb defines the ligand recognition specifity of integrin alpha IIb beta 3. J Biol Chem 1996; 271: 2033-9.
    CrossrefPubMedGoogle Scholar
  • 49 Honda S, Tomiyama Y, Shiraga M. et al. A twoamino acid insertion in the Cys146-Cys167 loop of the alphaIIb subunit is associated with a variant of Glanzmann thrombasthenia. Critical role of Asp163 in ligand binding. J Clin Invest 1998; 102: 1183-92.
    CrossrefPubMedGoogle Scholar
  • 50 Tamura T, Hato T, Yamanouchi J. et al. Critical residues for ligand binding in blade 2 of the propeller domain of the integrin alphaIIb subunit. Thromb Haemost 2004; 09: 111-8.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 51 Basani RB, D´Andrea G, Mitra N. et al. RGDcontaining peptides inhibit fibrinogen binding to platelet αIIbβ3 by inducing an allosteric change in the amino-terminal portion of αIIb. J Biol Chem 2001; 276: 13975-81.
    CrossrefPubMedGoogle Scholar
  • 52 Biris N, Abatzis M, Mitsios JV. et al. Mapping the binding domains of the αIIb subunit. Eur J Biochem (FEBS) 20: 3760-7.
    PubMedGoogle Scholar