Contribution of PAR-1, PAR-4 and GPIbalpha in intracellular signaling leading to the cleavage of the beta3 cytoplasmic domain during thrombin-induced platelet aggregation.
Integrin alphaIIbbeta3 plays a pivotal role in platelet aggregation by binding to fibrinogen. The beta3 cytoplasmic domain of alphaIIbbeta3 interacts with cytoskeletal and signaling proteins and is cleaved by micro -calpain, a calcium regulated cysteine protease. In the present study, we have investigated in more detail the cleavage of the beta3 cytoplasmic domain during platelet aggregation induced by thrombin, TRAP-1 and TRAP-4. Our data show that beta3 is cleaved in all three cases. The time course of beta3 cleavage and the amount of cleaved beta3 depends on the way platelets are activated and on the complete activation of micro -calpain, with a maximum of 90% of cleaved beta3 obtained when thrombin is used. Furthermore, our results also show that the cleaved alphaIIbbeta3 is mainly distributed in the Triton soluble fraction, indicating its inability to bind to the cytoskeleton. Interestingly, in the absence of GPIbalpha or following inhibition of thrombin binding to GPIbalpha, there is a reduction in the thrombin-induced calcium flux, beta3 cleavage and micro -calpain activation. These results suggest that cleavage of the beta3 cytoplasmic domain by micro -calpain might be an important step regulating the link between the cytoskeleton and alphaIIbbeta3 during platelet aggregation, and that GPIbalpha could function as a cofactor for the complete activation of platelets by thrombin.
Dubois
Christophe
C
Sylvie Meyer Reigner, PRBD-M, 68/217b, Hoffmann-La Roche, 4070 Basel, Switzerland.
Steiner
Beat
B
Meyer Reigner
Sylvie C
SC
D002154:Calpain / Q000378:metabolism; D003599:Cytoskeleton / Q000378:metabolism; D006801:Humans; D039661:Integrin beta3 / Q000378:metabolism*; D010974:Platelet Aggregation; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000378:metabolism; D019038:Platelet Glycoprotein GPIb-IX Complex / Q000502:physiology; D010980:Platelet Membrane Glycoproteins / Q000502:physiology*; D044463:Receptor, PAR-1 / Q000502:physiology; D018179:Receptors, Thrombin / Q000502:physiology; D015398:Signal Transduction; D013917:Thrombin / Q000502:physiology*
D039661:Integrin beta3; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D019038:Platelet Glycoprotein GPIb-IX Complex; D010980:Platelet Membrane Glycoproteins; D044463:Receptor, PAR-1; D018179:Receptors, Thrombin; D013917:Thrombin; D002154:Calpain; C449002:mu-calpain; C112908:protease-activated receptor 4