Snake venom metalloproteinases, crotarhagin and alborhagin, induce ectodomain shedding of the platelet collagen receptor, glycoprotein VI

Lakshmi C. Wijeyewickrema; Elizabeth E. Gardiner; Masaaki Moroi; Michael C. Berndt; Robert K. Andrews

doi:10.1160/TH07-06-0402

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 2007; 98(06): 1285-1290
DOI: 10.1160/TH07-06-0402

Platelets and Blood Cells

Schattauer GmbH

Snake venom metalloproteinases, crotarhagin and alborhagin, induce ectodomain shedding of the platelet collagen receptor, glycoprotein VI

Lakshmi C. Wijeyewickrema

¹Department of Biochemistry and Molecular Biology

,

Elizabeth E. Gardiner

²Department of Immunology, Monash University, Alfred Medical Research & Education Precinct, Melbourne, Victoria, Australia

,

Masaaki Moroi

³Department of Protein Biochemistry, Institute of Life Science, Kurume University, Kurume, Fukuoka, Japan

,

Michael C. Berndt

²Department of Immunology, Monash University, Alfred Medical Research & Education Precinct, Melbourne, Victoria, Australia

,

Robert K. Andrews

²Department of Immunology, Monash University, Alfred Medical Research & Education Precinct, Melbourne, Victoria, Australia

› Author Affiliations

Abstract

Summary

Glycoprotein (GP)VI, that binds collagen, together with GPIb- IX-V which binds von Willebrand factor, forms an adheso-signalling complex on platelets that initiates thrombus formation in haemostasis and thrombosis. In this study, we show that two snake venom metalloproteinases, crotarhagin and alborhagin, induce ectodomain shedding of GPVI by a mechanism that involves activation of endogenous platelet metalloproteinases. Alborhagin is a viper venom metalloproteinase from Trimeresurus albolabris, while crotarhagin is a previously undescribed toxin from the rattlesnake Crotalus horridus horridus (~60-kDa non-reduced and reduced). Like alborhagin, crotarhagin induces aggregation in human platelet-rich plasma (maximal activity, ~0.3 μg/ ml). Aggregation of washed platelets was inhibited by soluble GPVI ectodomain expressed as an Fc-fusion protein, confirming crotarhagin targeted GPVI. Treating washed platelets with crotarhagin or alborhagin resulted in time-dependent loss of surface GPVI and the appearance of an ~55-kDa soluble GPVI fragment in supernatants. Crotarhagin also induced shedding in GPVItransfected RBL-2H3 cells. Crotarhagin-induced shedding was metalloproteinase-dependent (inhibited by EDTA), but also blocked by inhibitors of GPVI signalling (Src kinase inhibitors, PP1 or PP2, or Syk inhibitor, piceatannol), indicating shedding required GPVI-dependent platelet activation. Together, the data suggest that the rattlesnake metalloproteinase, crotarhagin, and the viper toxin alborhagin, induce GPVI shedding by a mechanism involving activation of endogenous platelet metalloproteinases rather than direct cleavage of GPVI.

Keywords

ADAMS - adhesion receptors - snake venoms - platelet glycoproteins

Full Text

References

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