Modulating platelet reactivity through control of RGS18 availability

Abstract

Key Points• RGS18 acts as a brake on persistent or inappropriate platelet activation after it is released from binding sites in resting platelets.• Control of free RGS18 levels provides a mechanism for coordinating signaling networks in platelets.Most platelet agonists activate platelets by binding to G-protein-coupled receptors. We have shown previously that a critical node in the G-protein signaling network in platelets is formed by a scaffold protein, spinophilin (SPL), the tyrosine phosphatase, Src homology region 2 domain-containing phosphatase-1 (SHP-1), and the regulator of G-protein signaling family member, RGS18. Here, we asked whether SPL and other RGS18 binding proteins such as 14-3-3g regulate platelet reactivity by sequestering RGS18 and, if so, how this is accomplished. The results show that, in resting platelets, free RGS18 levels are relatively low, increasing when platelets are activated by thrombin. Free RGS18 levels also rise when platelets are rendered resistant to activation by exposure to prostaglandin I 2 (PGI 2 ) or forskolin, both of which increase platelet cyclic adenosine monophosphate (cAMP) levels. However, the mechanism for raising free RGS18 is different in these 2 settings. Whereas thrombin activates SHP-1 and causes dephosphorylation of SPL tyrosine residues, PGI 2 and forskolin cause phosphorylation of SPL Ser94 without reducing tyrosine phosphorylation. Substituting alanine for Ser94 blocks cAMP-induced dissociation of the SPL/RGS/SHP-1 complex. Replacing Ser94 with aspartate prevents formation of the complex and produces a loss-of-function phenotype when expressed in mouse platelets. Together with the defect in platelet function we previously observed in SPL 2/2 mice, these data show that (1) regulated sequestration and release of RGS18 by intracellular binding proteins provides a mechanism for coordinating activating and inhibitory signaling networks in platelets, and (2) differential phosphorylation of SPL tyrosine and serine residues provides a key to understanding both. (Blood. 2015;126(24):2611-2620

Platelets at the vascular interface

“…Consistently, it was previously shown that knockdown of RGS18[24,25], RGS10[26] or mutation of the RGS binding site on Gαi[27] results in increased platelet reactivity. At this ISTH conference, Gupta et al confirmed that knockdown of RGS10 results in a gain of platelet function, in particular in response to stimulation by the second wave mediators, ADP and TxA 2 [28].…”

Platelets and hemostasis: a new perspective on an old subject

“…26 As an example of pathway convergence, dissociation of the SPL/RGS complex also occurs when endothelium-derived PGI 2 suppresses platelet activation by raising platelet cAMP levels ( Figure 2). 31 For Rap1b, regulation occurs at the level of Rasa3, whose ability to act as a GAP is inhibited by signaling downstream of G i2 . 25,32 Rap1b 33 and CalDAG-GEF1, 34,35 like spinophilin, are targets for cAMP-dependent phosphorylation.…”

Differential Expression of Ccn4 and Other Genes Between Metastatic and Non-metastatic EL4 Mouse Lymphoma Cells

“…RGS18 negatively modulates the function of G-protein-coupled receptors (GPCRs). RGS18 makes platelets less sensitive to activation (30,31). Since PLD2 can be activated downstream of GPCRs (32), it is conceivable that the increase in RGS18 is a compensatory response to some of the effects of PLD2 overexpression.…”