KW - Platelet protein synthesis

AB - Myocardial infarction and stroke are caused by blood clots forming over a ruptured or denuded atherosclerotic plaque (atherothrombosis). Production of prostaglandin E2 (PGE2) by an inflamed plaque exacerbates atherothrombosis and may limit the effectiveness of current therapeutics. Platelets express multiple G-protein coupled receptors, including receptors for ADP and PGE2. ADP can mobilize Ca2+ and through the P 2Y12 receptor can inhibit cAMP production, causing platelet activation and aggregation. Clopidogrel (Plavix), a selective P 2Y12 antagonist, prevents platelets from clotting but thereby increases the risk of severe or fatal bleeding. The platelet EP 3 receptor for PGE2, like the P2Y12 receptor, also inhibits cAMP synthesis. However, unlike ADP, facilitation of platelet aggregation via the PGE2/EP3 pathway is dependent on co-agonists that can mobilize Ca2+. We used a ligand-based design strategy to develop peri-substituted bicylic acylsulfon- amides as potent and selective EP3 antagonists. We show that DG-041, a selective EP 3 antagonist, inhibits PGE2 facilitation of platelet aggregation in vitro and ex vivo. PGE2 can resensitize platelets to agonist even when the P2Y12 receptor has been blocked by clopidogrel, and this can be inhibited by DG-041. Unlike clopidogrel, DG-041 does not affect bleeding time in rats, nor is bleeding time further increased when DG-041 is co-administered with clopidogrel. This indicates that EP 3 antagonists potentially have a superior safety profile compared to P2Y12 antagonists and represent a novel class of antiplatelet agents.

T1 - Protein synthesis by platelets

Protein synthetic activity of various platelet populations

Synthesis of platelet protein in a cell‐free system

Discovery of synthesis of Bcl-3 by activated platelets sparked a search for the identities of other protein products, yielding IL-1β and TF. It also led to the unexpected discovery that their synthesis is preceded by signal-dependent cytoplasmic splicing of IL-1β and TF pre-mRNAs, yielding mature transcripts that are translated into precursor (IL-1β) and active (TF) proteins.,, This identified a novel mechanism not previously recognized in activated mammalian cells. The splicing capacities of activated platelets are intricate and will be reviewed separately. Signal-dependent splicing, together with the mTOR-dependent translational control mechanism and other regulatory pathways discussed here, indicate that platelets have unexpected diversity in posttranscriptional control. Previous and ongoing studies add to this conclusion and suggest that platelets may also use ribosomal “stalling” or polypeptide termination, participation of micro RNAs (Denis MM, Trask B, Schwertz H, Weyrich AS, Zimmerman GA, 2004) and, potentially, other modes of control. Diverse mechanisms of this sort are now emerging as important regulators of cell- and locale-selective mRNA translation and protein expression., Thus the findings to date indicate that platelets have a rich and varied repertoire of posttranscriptional pathways that are only now being revealed.

Protein synthesis in maturing human platelets

Other proteins are also reported to be synthesized by activated human platelets, although in some cases the control mechanisms have not been explored. These include plasminogen activator inhibitor (PAI) 1, tetrahydrobiopterin, cyclo-oxygenase-1, and the SVCT2 ascorbate transporter. There is also evidence that the inducible isoform of nitric oxide synthase is produced in activated platelets, and that apoptotic proteins may be synthesized as platelets age (reviewed in reference). As noted, experiments using radiolabeled amino acid incorporation and electrophoretic separation of newly synthesized proteins indicate that there are many other protein products, with preliminary identification of some of these in progress.

Stable messenger RNA in the synthesis of contractile protein in human platelets.
Biosynthesis of major platelet proteins in human blood platelets.

Protein synthesis in aging blood platelets.

As in other cell types, rapamycin inhibits synthesis of a subset of protein products—not all—in activated human platelets, indicating that mTOR is a specialized, rather than a general, translational control pathway. Discrimination and differential translation of these mTOR-regulated mRNAs lie in sequence features of the 5′ UTR of the transcripts, which include complex secondary structure that requires significant free energy to unwind, as well as other sequence variables.,

Extracellular factors influencing the in vitro protein synthesis of platelets.

(2005) Platelet Signal-Dependent Protein Synthesis.

Platelets are anucleate cytoplasts that circulate in the bloodstream for approximately 9–11 days. Because they lack nuclei, platelets were considered incapable of protein synthesis. However, studies over the last decade have revealed that platelets use a variety of translational control pathways to synthesize proteins.

Studies on the protein and nucleic acid synthesis of normal human blood platelets.

Platelet Signal-Dependent Protein Synthesis | …

Here, we chronologically review our increased understanding of protein synthetic responses in platelets and discuss how the field may evolve over the next decade.