Roles of platelet STIM1 and Orai1 in glycoprotein VI- and thrombin-dependent procoagulant activity and thrombus formation.
In platelets, STIM1 has been recognized as the key regulatory protein in store-operated Ca(2+) entry (SOCE) with Orai1 as principal Ca(2+) entry channel. Both proteins contribute to collagen-dependent arterial thrombosis in mice in vivo. It is unclear whether STIM2 is involved. A key platelet response relying on Ca(2+) entry is the surface exposure of phosphatidylserine (PS), which accomplishes platelet procoagulant activity. We studied this response in mouse platelets deficient in STIM1, STIM2, or Orai1. Upon high shear flow of blood over collagen, Stim1(-/-) and Orai1(-/-) platelets had greatly impaired glycoprotein (GP) VI-dependent Ca(2+) signals, and they were deficient in PS exposure and thrombus formation. In contrast, Stim2(-/-) platelets reacted normally. Upon blood flow in the presence of thrombin generation and coagulation, Ca(2+) signals of Stim1(-/-) and Orai1(-/-) platelets were partly reduced, whereas the PS exposure and formation of fibrin-rich thrombi were normalized. Washed Stim1(-/-) and Orai1(-/-) platelets were deficient in GPVI-induced PS exposure and prothrombinase activity, but not when thrombin was present as co-agonist. Markedly, SKF96365, a blocker of (receptor-operated) Ca(2+) entry, inhibited Ca(2+) and procoagulant responses even in Stim1(-/-) and Orai1(-/-) platelets. These data show for the first time that: (i) STIM1 and Orai1 jointly contribute to GPVI-induced SOCE, procoagulant activity, and thrombus formation; (ii) a compensating Ca(2+) entry pathway is effective in the additional presence of thrombin; (iii) platelets contain two mechanisms of Ca(2+) entry and PS exposure, only one relying on STIM1-Orai1 interaction.
Gilio
Karen
K
Department of Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), Maastricht University, 6200 MD Maastricht, The Netherlands.
van Kruchten
Roger
R
Braun
Attila
A
Berna-Erro
Alejandro
A
Feijge
Marion A H
MA
Stegner
David
D
van der Meijden
Paola E J
PE
Kuijpers
Marijke J E
MJ
Varga-Szabo
David
D
Heemskerk
Johan W M
JW
Nieswandt
Bernhard
B
D000818:Animals; D001792:Blood Platelets / Q000378:metabolism; D002118:Calcium / Q000737:chemistry / Q000378:metabolism; D015220:Calcium Channels / Q000378:metabolism*; D003029:Coagulants / Q000378:metabolism*; D005260:Female; D008562:Membrane Glycoproteins / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D008954:Models, Biological; D000071740:ORAI1 Protein; D010980:Platelet Membrane Glycoproteins / Q000378:metabolism*; D000071737:Stromal Interaction Molecule 1; D013917:Thrombin / Q000378:metabolism*; D013927:Thrombosis / Q000378:metabolism*
D015220:Calcium Channels; D003029:Coagulants; D008562:Membrane Glycoproteins; D000071740:ORAI1 Protein; C520430:Orai1 protein, mouse; D010980:Platelet Membrane Glycoproteins; C100757:Stim1 protein, mouse; D000071737:Stromal Interaction Molecule 1; C098341:platelet membrane glycoprotein VI; D013917:Thrombin; D002118:Calcium