Mechanisms regulating factor V endocytosis by megakaryocytes
I received my Ph.D. in the Cell & Molecular Biology Program at the University of Vermont in 1996. My dissertation research focused on the regulation of procoagulant enzyme complex assembly and function on cells. During a Postdoctoral Fellowship with Barbara Furie, Ph.D. at Harvard Medical School, I continued with my research interests in coagulation studying the vitamin K-dependent gamma-glutamyl carboxylase, the enzyme responsible for catalyzing an important post-translational modification of the blood clotting proteins. In 2000, I returned to UVM and am currently an Assistant Professor of Biochemistry.
Platelet-derived factor Va is critical for blood clot formation by serving as the essential cofactor for thrombin generation via the prothrombinase complex assembled on activated platelets. The entire platelet-derived factor Va pool originates via endocytosis of the procofactor, factor V, from plasma by megakaryocytes, the platelet precursor cells. Factor V endocytosis by megakaryocytes is specific, clathrin-dependent, and appears to be mediated by a two receptor system. In this model, factor V initially binds to a specific factor V receptor expressed only on megakaryocytes able to endocytose factor V. This binding event facilitates an interaction between a second factor V molecule and LDL receptor-related protein-1 (LRP-1), an endocytic receptor, which subsequently mediates the endocytosis of bound factor V. These combined observations represent a unique role for LRP-1 in endocytosis of a coagulation protein not destined for lysosomal degradation. Rather, subsequent to its endocytosis, factor V is functionally modified, trafficked to, and stored in alpha-granules.
The overall goal of our laboratory is to identify and characterize the proteins that form the megakaryocyte two receptor system involved in the binding and endocytosis of plasma-derived factor V.
Some specific projects include: (1) Identification of the specific, factor V receptor expressed on human megakaryocytes; (2) Determination of the fates of the factor V receptor and LRP-1 subsequent to factor V endocytosis, and during megakaryocyte development and platelet formation; (3) Identification of the minimum factor V amino acid sequence(s) involved in its interactions with the factor V receptor, as well as LRP-1 expressed on megakaryocytes; and (4) Definition of the intracellular signaling events that regulate factor V binding and endocytosis by megakaryocytes.
Our laboratory is also involved in large population-based studies of thrombophilic and hemophilic individuals. These studies, being performed in collaboration with Dr. Kathleen Brummel-Ziedins, use a flow cytometry-based assay of prothrombinase complex assembly on activated platelets in whole blood developed by our laboratory to predict an individual's propensity to bleed or clot.
Model of factor V endocytosis by megakaryocytes
Endocytosis of factor V by megakaryocytes
Bouchard, B.A., Meisler, N.T., Nesheim, M.E., Liu, C.X., Strickland, D.K., and Tracy, P.B. A unique function for LRP-1: a component of a two-receptor system mediating specific endocytosis of plasma-derived factor V by megakaryocytes. J Thromb Haemosta 6(4):638-44, 2008.
Bouchard, B.A., Butenas, S., Mann K.G. and Tracy, P.B. Interactions between platelets and the coagulation system. In: Platelets, Michelson AD (2nd Ed), Academic Press, San Diego, CA, pp. 377-402, 2007.
Bouchard, B.A., Williams, J.L., Meisler, N.T., Long, M.W. and Tracy, P.B. Endocytosis of plasma factor V by megakaryocytes occurs via a clathrin-dependent, specific receptor-mediated event. J Thromb Haemosta 3:541-551, 2005
Damron, D.P., Bouchard, B.A., Shapiro, R.E., Schonberg, A.L. and Bernstein, I.M. Platelet activation, sympathetic tone, and plasma volume in nulligravid women of reproductive age. Obstet Gynecol 103:931-936, 2004
Bouchard, B., Paradis, A., and Brummel-Ziedins, K. Measurement of procoagulant platelet subpopulations in whole blood: Development of an assay for population-based studies. Thromb Res. 2011 Jan;127(1):62-4. Epub 2010 Jun 11.
* indicates equal contribution
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