Molecular Basis of Platelet Function

The last 30 years have seen an explosion in our understanding of the intricate molecular mechanisms that operate in platelets during hemostasis and of the aberrations that lead to platelet diseases. This advance in our knowledge has occurred by integrated application of a wide range of techniques, including classical cell biology and biochemistry, molecular biology, and gene knockouts. A small fraction of this impressive expansion of our platelet knowledge is captured in this series of reviews, written by individuals who have made major contributions to the field. This issue focuses on the molecular mechanisms operating in platelets in hemostasis. The next issue deals with the aberrant mechanisms that lead to platelet diseases.

It all begins with the bone marrow megakaryocytes and the events that lead to the formation of circulating anucleate fragments, the platelets. Megakaryocytes extend elongated cytoplasmic projections called proplatelets that are eventually released to form platelets. In the first review, Schulze and Shivdasani provide an elegant review of the molecular and cellular mechanisms involved in megakaryocyte differentiation and platelet formation. This intricate sequence of coordinated processes is regulated by specific transcription factors, which modulate distinct events such as lineage commitment, megakaryocyte differentiation, proplatelet formation, and platelet release. The authors provide an overview of these mechanisms as well as of the human and murine models of thrombocytopenia resulting from altered transcriptional regulation of events of platelet formation.

Formation of the platelet plug is fundamental to the restoration of hemostasis following a vessel-wall breach. The events involved encompass the initiation of the process by an interaction between platelets, von Willebrand factor (vWF), and collagen, leading to the capture of the moving platelet, recruitment of additional platelets through mediation of agonists such as thrombin, adenosine diphosphate (ADP), and thromboxane A2 to extend the platelet plug and, finally, the stabilization of the plug until wound healing occurs. In the second review, Brass and colleagues focus on the events surrounding platelet-platelet interactions, and discuss the role of critical molecules, including the integrin α_IIbβ₃, cell adhesion molecules (CAMs), ligands shed from the surface of activated platelets (CD40L and Sema 4D/CD100), and receptor tyrosine kinases (EphA4 and EphB1) and ligands that bind them, called ephrins. What emerges is an exciting picture of complex events that result from or modulate platelet-platelet interactions, and continue well after the initial platelet aggregation.

Platelets possess several receptors on their surface, the activation of which results in the initiation of the signaling events. ADP was the first platelet agonist to be recognized. In the next review, Murugappan, Shankar, and Kunapuli provide an overview of the platelet adenine nucleotide receptors, their distinct features as well as interactions in mediating the various platelet responses to activation. Recent evidence from multiple approaches, including the knockout models and patients deficient in specific ADP receptors, has extended our knowledge of the physiologic importance of these receptors. They also discuss the role of the two thromboxane receptors on platelets in signaling and as potential targets for antithrombotic agents.

Platelet-collagen interactions are crucial for hemostasis and thrombosis. What has been challenging to define with clarity is the issue of the specific platelet receptors that modulate the multiple and complex interactions between platelets and collagen. Current models encompass a role for at least two, if not more, receptors (GPVI and integrin α₂β₁) that regulate one or more aspects of these interactions. In the next review, Kahn presents the molecular aspects of platelet-collagen interactions and the associated signaling events. He provides a cogent argument for exploring inhibition of platelet-collagen interactions as a therapeutic goal in cardiovascular diseases.

In the next review, Buensuceso, Arias-Salgado, and Shattil provide a detailed discussion of the impressive number of proteins that interact with the cytoplasmic tails of α_2bβ₃ in the context of signaling that occurs through this integrin. Platelet activation induces binding of ligands (fibrinogen, vWF) to α_IIbβ₃, which is highly regulated by inside-out signals generated by the interaction of an agonist with the platelet surface receptor. This ligand binding initiates outside-in signaling that modulates other processes, such as cytoskeletal reorganization and shape change. A unifying theme that addresses the mechanisms governing the α_2bβ₃-related bidirectional signaling invokes a major role of specific proteins that interact with cytoplasmic tails of either the α_IIb or β₃. These proteins and the available evidence are presented in this review.

Activated platelets secrete their granule contents, which are impressive in their number, diversity, and functional capabilities. This exocytosis is essential to the role of platelets in hemostasis and thrombosis. In the next review, Reed presents the rapidly emerging molecular mechanisms regulating platelet secretion, focusing on discrete events such as granule formation, membrane fusion, and release of contents, and on the specific proteins involved, such as soluble N-ethylmaleimide-sensitive factor associated protein receptor (SNARE) proteins. It is fascinating that platelet secretory mechanisms are homologous to those in other cells, notably neurons.

Increased intracellular levels of cyclic adenosine monophosphate (cAMP) inhibit almost all platelet responses to platelet activation. Phosphodiesterases hydrolyze cAMP, regulate cellular cAMP levels, and are thus major regulators of platelet responses. In the next review, Colman presents the state of the field with respect to the platelet cAMP phosphodiesterases and regulation of cAMP activity in platelets. This information forms a basis for modulating platelet function for therapeutic purposes by focusing on inhibition of the phosphodiesterases.

In the next review, Walsh delves in depth into the intimate role of platelets in the enzymatic reactions of blood coagulation-events in which these anucleate cells dramatically amplify the initial stimulus through diverse mechanisms, including localizing the coagulation enzymes, cofactors, and substrates, and protecting them from inactivation by inhibitors. That this role of platelets is not a passive phenomenon is evidenced by the essential requirement of platelet activation in the assembly of coagulation factors on the membrane and amplification of the enzymatic reactions.

Three reviews in this issue tackle topics that signal a paradigm shift in the way we view platelets. Previous investigations have revealed that circulating platelets retain mRNA derived from the parent cell. The relevance or the potential information from this mRNA has not been adequately explored until recently. Bahou and Gnatenko describe the application of microarray analysis to platelets to demonstrate a molecular expression signature unique to platelets. This review on the feasibility of delineating the platelet transcriptome reflects the promise of expression profiling and its application to human disease. In the next review, Garcia, Zitzmann, and Watson extend this into the realm of defining the platelet proteome-the whole set of proteins present in the cell. The era of platelet proteomics and genomics has been launched, and these approaches are undoubtedly applicable to platelets, and in the future will constitute major tools to understand platelet biology and pathology. The third review in this set challenges the widely held belief that platelets, because of their anucleate status, are incapable of protein synthesis. Weyrich and colleagues review the evidence that platelets are endowed with the ability to translate mRNA into proteins with significant biological roles in inflammation and thrombosis. It is remarkable that the platelet can respond to activation with protein synthesis as well. Together, these three reviews may be construed as a promissory note of new platelet information to be garnered through application of these approaches.

Lastly, I thank the authors who have contributed to this issue of Seminars in Thrombosis and Hemostasis. Their contributions have advanced not only this issue but the field as well.