Extracellular Matrix-Specific Platelet Activation Leads to a Differential Translational Response and Protein De Novo Synthesis in Human Platelets.

Bjoern F Kraemer,Hanna Mannell,Marc Geimer,Mirita Franz-Wachtel,Tobias Lamkemeyer,Stephan Lindemann

doi:10.3390/ijms21218155

Abstract

Platelets are exposed to extracellular matrix (ECM) proteins like collagen and laminin and to fibrinogen during acute vascular events. However, beyond hemostasis, platelets have the important capacity to migrate on ECM surfaces, but the translational response of platelets to different extracellular matrix stimuli is still not fully characterized. Using 2D-gel electrophoresis, confocal microscopy, polysome analysis and protein sequencing by mass spectrometry, we demonstrate that platelets show a differential expression profile of newly synthesized proteins on laminin, collagen or fibrinogen. In this context, we observed a characteristic, ECM-dependent translocation phenotype of translation initiation factor eIF4E to the ribosomal site. eIF4E accumulated in polysomes with increased binding of mRNA and co-localization with vinculin, leading to de novo synthesis of important cytoskeletal regulator proteins. As the first study, we included a proteome analysis of laminin-adherent platelets and interestingly identified upregulation of essentially important proteins that mediate cytoskeletal regulation and mobility in platelets, such as filamin A, talin, vinculin, gelsolin, coronin or kindlin-3. In summary, we demonstrate that platelet activation with extracellular matrix proteins results in a distinct stimulus-specific translational response of platelets that will help to improve our understanding of the regulation of platelet mobility and migration.

Highlights

Platelets are essential for cell-mediated hemostasis and activation of platelets results in a fast and effective formation of platelet aggregates and blood clots
We demonstrate that platelets that are exposed to different extracellular matrix components or fibrinogen show a distinct phenotype and pattern of cytoskeletal arrangement after activation and that the translation initiation factor eIF4E characteristically redistributes to the ribosomal sites inside the cytoskeleton. eIF4E shows markedly enhanced mRNA binding and translation initiation that leads to increased synthesis of important cytoskeletal regulators such as vinculin, gelsolin, talin 1, filamin A, coronin or kindlin-3 (FERMT3)
By using methionine incorporation for detection of newly synthesized proteins, we confirmed up-front that the extent of protein synthesis markedly differs between platelet activators

Summary

Introduction

Platelets are essential for cell-mediated hemostasis and activation of platelets results in a fast and effective formation of platelet aggregates and blood clots This is especially important in pathophysiological events like myocardial infarction and stroke, where platelets are exposed to laminin, a component of the basal lamina, and collagen, the predominant extracellular matrix protein. Besides classical functions of hemostasis, previous work from our group and others has discovered that platelets are able to perform a much more complex and dynamic cytoskeletal rearrangement that enables platelets to migrate [4,5,6,7,8,9,10,11] These studies demonstrated that platelets can migrate under flow conditions, towards a cytokine gradient or along bacterial trails, and that morphology and phenotype of migrating platelets differs depending on the underlying extracellular matrix surface [6,7,8]. Despite fast-growing information from proteomic analysis, cellular mechanisms for differential protein translation and cytoskeletal regulation are not fully understood

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