Abstract
AbstractAbstract 161Platelets play an essential role in hemostasis, but excessive platelet responses to vascular injury or disease can be catastrophic. We have recently reported that members of the RGS protein family can modulate platelet responses to injury by limiting the duration of G protein-dependent signaling initiated by platelet agonists. Here we show that platelets contain a previously-unrecognized regulatory complex comprised of the 130 kDa scaffold protein, spinophilin (SPL) with at least two RGS proteins (RGS10 and RGS18) and the protein tyrosine phosphatase, SHP-1. In resting platelets, this complex is phosphorylated on spinophilin residues Y398 and Y483. Mutating both tyrosines to phenylalanine inhibits the binding of SHP-1 to spinophilin. Platelet activation by thrombin or thromboxane A2, but not ADP or collagen, stimulates a transient increase in spinophilin-associated SHP-1 tyrosine phosphatase activity and causes dephosphorylation and decay of the SPL/RGS/SHP-1 complex. Conversely, blocking SHP-1 phosphatase activity in platelets or omitting SHP-1 in transfected CHO cells inhibits dephosphorylation of spinophilin and prevents dissociation of the SPL/RGS/SHP-1 complex. While we have shown previously that inhibiting interactions between G proteins and RGS proteins produces a gain of function in platelets, knocking out spinophilin in mice inhibits platelet aggregation. This aggregation defect does not occur with all agonists, but is selective for those that are able to trigger decay and dephosphorylation of the SPL/RGS/SHP-1 complex. In addition to inhibiting platelet aggregation in vitro, the spinophilin knockout delays carotid artery occlusion in vivo following application of FeCl3 and reduces platelet accumulation following laser injury in cremaster muscle arterioles. Underlying these effects of the knockout is a decrease in Rap1 activation, an event that supports integrin activation, and attenuation of the cAMP increase otherwise caused by endothelial PGI2. Collectively, these observations show for the first time that a regulatory complex based on spinophilin helps to regulate platelet responses to injury and suggest that it does this by sequestering RGS proteins in resting platelets and releasing them after activation begins. Dissociation of the SPL/RGS complex is regulated by an agonist-induced increase in the activity of SHP-1 associated with spinophilin. Disclosures:No relevant conflicts of interest to declare.
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Disclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.