Proteolysis of plasma-derived factor V following its endocytosis by megakaryocytes forms the platelet-derived factor V/Va pool


  • Manuscript handled by: R. Medcalf
  • Final decision: D. Lane, 16 May 2013

Correspondence: Beth A. Bouchard, Department of Biochemistry, University of Vermont College of Medicine, 89 Beaumont Ave., Given C440, Burlington, VT 05405, USA.

Tel.: +1 802 656 6529; fax: +1 802 656 8220.




Central to appropriate thrombin formation at sites of vascular injury is the concerted assembly of plasma- and/or platelet-derived factor (F) Va and FXa on the activated platelet surface. While the plasma-derived procofactor, FV, must be proteolytically activated by α-thrombin to FVa to function in prothrombinase, the platelet molecule is released from α-granules in a partially activated state, obviating the need for proteolytic activation.


The current study was performed to test the hypothesis that subsequent to its endocytosis by megakaryocytes, plasma-derived FV is proteolytically processed to form the platelet-derived pool.

Methods & Results

Subsequent to FV endocytosis, a time-dependent increase in FV proteolytic products was observed in megakaryocyte lysates by SDS-PAGE followed by phosphorimaging or western blotting. This cleavage was specific and resulted in the formation of products similar in size to FV/Va present in a platelet lysate as well as to the α-thrombin-activated FVa heavy chain and light chain, and their respective precursors. Other proteolytic products were unique to endocytosed FV. The product/precursor relationships of these fragments were defined using anti-FV heavy and light chain antibodies with defined epitopes. Activity measurements indicated that megakaryocyte-derived FV fragments exhibited substantial FVa cofactor activity that was comparable to platelet-derived FV/Va.


Taken together, these observations suggest that prior to its packaging in α-granules endocytosed FV undergoes proteolysis by one or more specific megakaryocyte protease(s) to form the partially activated platelet-derived pool.