The theme of this project is to understand how complexes of proteins within the blood clotting cascade assemble on membrane surfaces and how this assembly process regulates enzymatic activity. The proposed studies focus on assembly of the cell-surface complex that triggers blood clotting, consisting of the regulatory protein, tissue factor, and its cognate serine protease, factor Vila;and investigating the mechanism of action of novel regulators of blood clotting that we have recently discovered. Specifically, we will investigate the following research questions: (1) How does membrane topography contribute to tissue factor function? These studies will examine the consequences of altering the membrane topography of tissue factor on the proteolytic activity of the tissue factor-factor Vila complex. Included is an investigation of a novel way of attaching the extracellular domain of tissue factor (sTF) to membrane surfaces employing oligohistidine-tagged sTF and metal ion-chelating lipids. (2) How does polyphosphate modulate the blood clotting cascade? We discovered that polyphosphate, an ancient molecule that is abundant in many microorganisms, and newly discovered in dense granules of human platelets, is a potent modulator of blood clotting and fibrinolysis. We will examine: (a) platelet secretion of polyphosphate, including studies comparing normal platelets and platelets from patients with Hermansky-Pudlak syndrome;(b) the ability of polyphosphate to stimulate the contact pathway of blood clotting;(c) the mechanism by which polyphosphate completely abrogates the anticoagulant function of tissue factor pathway inhibitor (TFPI);(d) how polyphosphate modulates thrombin function;and (e) how expression of polyphosphate in bacteria (Escherichia coli and Salmonella typhimurium) modulates the blood clotting and fibrinolytic cascades. These studies are aimed at understanding how newly discovered modulators of the blood clotting system contribute to normal hemostasis and to the activation of the blood clotting system in human disease.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL047014-17
Application #
7751229
Study Section
Hemostasis and Thrombosis Study Section (HT)
Program Officer
Link, Rebecca P
Project Start
1992-08-01
Project End
2011-06-30
Budget Start
2010-01-01
Budget End
2011-06-30
Support Year
17
Fiscal Year
2010
Total Cost
$344,250
Indirect Cost
Name
University of Illinois Urbana-Champaign
Department
Biochemistry
Type
Schools of Arts and Sciences
DUNS #
041544081
City
Champaign
State
IL
Country
United States
Zip Code
61820
Gajsiewicz, Joshua M; Smith, Stephanie A; Morrissey, James H (2017) Polyphosphate and RNA Differentially Modulate the Contact Pathway of Blood Clotting. J Biol Chem 292:1808-1814
Sylman, Joanna L; Daalkhaijav, Uranbileg; Zhang, Ying et al. (2017) Differential Roles for the Coagulation Factors XI and XII in Regulating the Physical Biology of Fibrin. Ann Biomed Eng 45:1328-1340
Smith, Stephanie A; Baker, Catherine J; Gajsiewicz, Joshua M et al. (2017) Silica particles contribute to the procoagulant activity of DNA and polyphosphate isolated using commercial kits. Blood 130:88-91
Kalathottukaren, Manu Thomas; Abraham, Libin; Kapopara, Piyushkumar R et al. (2017) Alteration of blood clotting and lung damage by protamine are avoided using the heparin and polyphosphate inhibitor UHRA. Blood 129:1368-1379
Puy, Cristina; Tucker, Erik I; Ivanov, Ivan S et al. (2016) Platelet-Derived Short-Chain Polyphosphates Enhance the Inactivation of Tissue Factor Pathway Inhibitor by Activated Coagulation Factor XI. PLoS One 11:e0165172
Wijeyewickrema, Lakshmi C; Lameignere, Emilie; Hor, Lilian et al. (2016) Polyphosphate is a novel cofactor for regulation of complement by a serpin, C1 inhibitor. Blood 128:1766-76
Zilberman-Rudenko, Jevgenia; Itakura, Asako; Wiesenekker, Chantal P et al. (2016) Coagulation Factor XI Promotes Distal Platelet Activation and Single Platelet Consumption in the Bloodstream Under Shear Flow. Arterioscler Thromb Vasc Biol 36:510-7
Hassanian, S M; Dinarvand, P; Smith, S A et al. (2015) Inorganic polyphosphate elicits pro-inflammatory responses through activation of the mammalian target of rapamycin complexes 1 and 2 in vascular endothelial cells. J Thromb Haemost 13:860-71
Travers, R J; Smith, S A; Morrissey, J H (2015) Polyphosphate, platelets, and coagulation. Int J Lab Hematol 37 Suppl 1:31-5
Smith, Stephanie A; Morrissey, James H (2015) 2013 scientific sessions Sol Sherry distinguished lecture in thrombosis: polyphosphate: a novel modulator of hemostasis and thrombosis. Arterioscler Thromb Vasc Biol 35:1298-305

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