The serpin proteinase inhibitor 8: An endogenous furin inhibitor released from human platelets

Julie Leblond; Marie-Hélène Laprise; Simon Gaudreau; Francine Grondin; Walter Kisiel; Claire M. Dubois

doi:10.1160/TH05-08-0561

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 2006; 95(02): 243-252
DOI: 10.1160/TH05-08-0561

Platelets and Blood Cells

Schattauer GmbH

The serpin proteinase inhibitor 8: An endogenous furin inhibitor released from human platelets

Julie Leblond

¹Immunology Division, Faculty of Medicine, Université de Sherbrooke, Sherbrooke, Qc, Canada

,

Marie-Hélène Laprise

¹Immunology Division, Faculty of Medicine, Université de Sherbrooke, Sherbrooke, Qc, Canada

,

Simon Gaudreau

¹Immunology Division, Faculty of Medicine, Université de Sherbrooke, Sherbrooke, Qc, Canada

,

Francine Grondin

¹Immunology Division, Faculty of Medicine, Université de Sherbrooke, Sherbrooke, Qc, Canada

,

Walter Kisiel

²Department of Pathology, University of New Mexico School of Medicine, Albuquerque, New Mexico, USA

,

Claire M. Dubois

¹Immunology Division, Faculty of Medicine, Université de Sherbrooke, Sherbrooke, Qc, Canada

› Author Affiliations

Abstract

Summary

The cornerstone of hemostasis is the ability of the organism to limit the enzymatic processes involved, thereby avoiding thrombosis. For this, anticoagulant systems in place involve serpins, such as PAI-1 and antithrombin III, which bind to their targeted serine proteases and limit their period of activity. We have previously identified the serine protease furin as a platelet-derived enzyme with an intrinsic role in platelet functions. We now report that furin enzymatic activity decreased rapidly following platelet activation, corresponding with the increase in formation of a high 180 M _r SDS-stable complex composed of furin and the PI8 serpin. PI8 is shown to be a platelet-derived constituent, synthesized by megakaryocytes and stored in platelets prior to its release. Immunoprecipitation and purification of the PI8-furin complex confirmed their direct interaction and indicates that one of the roles of PI8 is to inhibit furin enzymatic activity. Furthermore, our findings demonstrate the inhibitory capacity of exogenous PI8 in platelet aggregation assays. The finding that PI8 is released by platelets and controls functional responses suggests a role for this serpin in platelet-regulated pathophysiological responses.

Keywords

Proteinase inhibitor 8 - serpin - furin - furin inhibitor - platelets

Full Text

References

References
1 Huber R, Carrell RW. Implications of the threedimensional structure of alpha 1-antitrypsin for structure and function of serpins. Biochemistry 1989; 28: 8951-66.
2 Lawrence DA, Olson ST, Palaniappan S. et al. Serpin reactive center loop mobility is required for inhibitor function but not for enzyme recognition. J Biol Chem 1994; 269: 27657-62.
3 Potempa J, Korzus E, Travis J. The serpin superfamily of proteinase inhibitors: structure, function, and regulation. J Biol Chem 1994; 269: 15957-60.
4 Patston PA, Gettins PG, Schapira M. The mechanism by which serpins inhibit thrombin and other serine proteinases. AnnN Y Acad Sci 1994; 714: 13-20.
5 Egeberg O. Proceedings: Inherited antithrombin III deficiency causing thrombophilia. Thromb Diath Haemorrh 1965; 13: 516.
6 Schleef RR, Higgins DL, Pillemer E. et al. Bleeding diathesis due to decreased functional activity of type 1 plasminogen activator inhibitor. J Clin Invest 1989; 83: 1747-52.
7 Hosaka M, Nagahama M, Kim WS. et al. ArgX-Lys/Arg-Arg motif asa signal for precursor cleavage catalyzed by furin within the constitutive secretory pathway. J Biol Chem 1991; 266: 12127-30.
8 Thomas G. Furin at the cutting edge: from protein traffic to embryogenesis and disease. Nat Rev Mol Cell Biol 2002; 03: 753-66.
9 Diuguid DL, Rabiet MJ, Furie BC. et al. Molecular basis of hemophilia B:a defective enzyme due to an unprocessed propeptide is caused by a point mutation in the factor IX precursor. Proc Natl Acad Sci USA 1986; 83: 5803-7.
10 Lind B, Johnsen AH, Thorsen S. Naturally occurring Arg(-1) to His mutation in human proteinC leads to aberrant propeptide processing and secretion of dysfunctional protein C. Blood 1997; 89: 2807-16.
11 Himmelspach M, Pfleiderer M, Fischer BE. et al. Recombinant human factor X: high yield expression and the role of furin in proteolytic maturation in vivo and in vitro . Thromb Res 2000; 97: 51-67.
12 Wise RJ, Barr PJ, Wong PA. et al. Expression of a human proprotein processing enzyme: correct cleavage of the von Willebrand factor precursor at a paired basic amino acid site. Proc Natl Acad Sci USA 1990; 87: 9378-82.
13 Rehemtulla A, Kaufman RJ. Preferred sequence requirements for cleavage of pro-von Willebrand factor by propeptide-processing enzymes. Blood 1992; 79: 2349-55.
14 Dubois CM, Laprise MH, Blanchette F. et al. Processing of transforming growth factor beta1 precursor by human furin convertase. J Biol Chem 1995; 270: 10618-24.
15 Fugere M, Day R. Inhibitors of the subtilase-like pro-protein convertases (SPCs). Curr Pharm Des 2002; 08: 549-62.
16 Jean F, Stella K, Thomas L. et al. alpha1-Antitrypsin Portland, a bioengineered serpin highly selective for furin: application as an antipathogenic agent. Proc Natl Acad Sci USA 1998; 95: 7293-8.
17 Dufour EK, Denault JB, Hopkins PC. et al. Serpinlike properties of alpha1-antitrypsin Portland towards furin convertase. FEBS Lett 1998; 426: 41-6.
18 Richer MJ, Keays CA, Waterhouse J. et al. The Spn4 gene of Drosophila encodes a potent furin-directed secretory pathway serpin. Proc Natl Acad Sci USA 2004; 101: 10560-5.
19 Dahlen JR, Jean F, Thomas G. et al. Inhibition of soluble recombinant furin by human proteinase inhibitor 8. J Biol Chem 1998; 273: 1851-4.
20 Silverman GA, Bird PI, Carrell RW. et al. The serpins are an expanding superfamily of structurally similar but functionally diverse proteins. Evolution, mechanism of inhibition, novel functions, anda revised nomenclature. J Biol Chem 2001; 276: 33293-6.
21 Strik MC, Bladergroen BA, Wouters D. et al. Distribution of the human intracellular serpin protease in-hibitor 8 in human tissues. J Histochem Cytochem 2002; 50: 1443-54.
22 Dahlen JR, Foster DC, Kisiel W. The inhibitory specificity of human proteinase inhibitor 8 is expanded through the use of multiple reactive site residues. Biochem Biophys Res Commun 1998; 244: 172-7.
23 Dahlen JR, Foster DC, Kisiel W. Expression, Purification, and Inhibitory Properties of Human Proteinase Inhibitor 8. Biochemistry 1997; 36: 14874-82.
24 Sprecher CA, Morgenstern KA, Mathewes S. et al. Molecular cloning, expression, and partial characterization of two novel members of the ovalbumin family of serine proteinase inhibitors. J Biol Chem 1995; 270: 29854-61.
25 van Duijnhoven HL, Creemers JW, Kranenborg MG. et al. Development and characterization of a panel of monoclonal antibodies against the novel subtilisinlike proprotein processing enzyme furin. Hybridoma 1992; 11: 71-86.
26 Laprise MH, Grondin F, Cayer P. et al. Furin gene (fur) regulation in differentiating human megakaryoblastic Dami cells: involvement of the proximal GATA recognition motif in the P1 promoter and impact on the maturation of furin substrates. Blood 2002; 100: 3578-87.
27 Vidricaire G, Denault JB, Leduc R. Characterization of a secreted form of human furin endoprotease. Biochem Biophys Res Commun 1993; 195: 1011-8.
28 Schleef RR, Chuang TL. Protease inhibitor 10 inhibits tumor necrosis factor alpha -induced cell death. Evidence for the formation of intracellular high M(r) protease inhibitor 10-containing complexes. J Biol Chem 2000; 275: 26385-9.
29 Laprise MH, Grondin F, Dubois CM. Enhanced TGFbeta1 maturation in high five cells coinfected with recombinant baculovirus encoding the convertase furin/pace: improved technology for the production of recombinant proproteins in insect cells. Biotechnol Bioeng 1998; 58: 85-91.
30 Vey M, Schafer W, Berghofer S. et al. Maturation of the trans-Golgi network protease furin: compartmentalization of propeptide removal, substrate cleavage, and COOH-terminal truncation. J Cell Biol 1994; 127: 1829-42.
31 Plaimauer B, Mohr G, Wernhart W. et al. ‘Shed’ furin: mapping of the cleavage determinants and identification of its C-terminus. Biochem J 2001; 354: 689-95.
32 Denault J, Bissonnette L, Longpre J. et al. Ectodomain shedding of furin: kinetics and role of the cysteine-rich region. FEBS Lett 2002; 527: 309-14.
33 Kestin AS, Ellis PA, Barnard MR. et al. Effect of strenuous exercise on platelet activation state and reactivity. Circulation 1993; 88: 1502-11.
34 Michelson AD, Barnard MR, Khuri SF. et al. The effects of aspirin and hypothermia on platelet function in vivo . Br J Haematol 1999; 104: 64-8.
35 Alessi MC, Chomiki N, Berthier R. et al. Detection of plasminogen activator inhibitor-1 (PAI-1) mRNA in human megakaryocytes by in situ hybridization. Thromb Haemost 1994; 72: 931-6.
36 Plow EF, Collen D. The presence and release of alpha 2-antiplasmin from human platelets. Blood 1981; 58: 1069-74.
37 Nachman RL, Harpel PC. Platelet alpha2-macroglobulin and alpha1-antitrypsin. J Biol Chem 1976; 251: 4512-21.
38 Booth NA. Fibrinolysis and thrombosis. Baillieres Best Pract Res Clin Haematol 1999; 12: 423-33.
39 Mann KG, Krishnaswamy S, Lawson JH. Surfacedependent hemostasis. Semin Hematol 1992; 29: 213-26.
40 Chang CP, Zhao J, Wiedmer T. et al. Contribution of platelet microparticle formation and granule secretion to the transmembrane migration of phosphatidylserine. J Biol Chem 1993; 268: 7171-8.
41 Belin D, Wohlwend A, Schleuning WD. et al. Facultative polypeptide translocation allows a single mRNA to encode the secreted and cytosolic forms of plasminogen activators inhibitor 2. Embo J 1989; 08: 3287-94.
42 Gombau L, Schleef RR. Processing of type 1 plasminogen activator inhibitor (PAI-1) into the regulated secretory pathway. J Biol Chem 1994; 269: 3875-80.
43 Lang IM, Chuang TL, Barbas 3rd CF. et al. Purification of storage granule protein-23. A novel protein identified by phage display technology and interaction with type I plasminogen activator inhibitor. J Biol Chem 1996; 271: 30126-35.
44 Norris LA. Blood coagulation. Best Pract Res Clin Obstet Gynaecol 2003; 17: 369-83.
45 Blakytny R, Ludlow A, Martin GE. et al. Latent TGF-beta1 activation by platelets. J Cell Physiol 2004; 199: 67-76.
46 Turecek PL, Pichler L, Auer W. et al. Evidence for extracellular processing of pro-von Willebrand factor after infusion in animals with and without severe von Willebrand disease. Blood 1999; 94: 1637-47.
47 Fair DS, Bahnak BR. Human hepatoma cells secrete single chain factor X, prothrombin, and antithrombin III. Blood 1984; 64: 194-204.
48 Graves CB, Munns TW, Willingham AK. et al. Rat factor X is synthesized as a single chain precursor inducible by prothrombin fragments. J Biol Chem 1982; 257: 13108-13.