Platelet integrins exhibit anisotropic mechanosensing and harness piconewton forces to mediate platelet aggregation

Abstract

Platelet aggregation at the site of vascular injury is essential in clotting. During this process, platelets are bridged by soluble fibrinogen that binds surface integrin receptors. One mystery in the mechanism of platelet aggregation pertains to how resting platelets ignore soluble fibrinogen, the third most abundant protein in the bloodstream, and yet avidly bind immobile fibrinogen on the surface of other platelets at the primary injury site. We speculate that platelet integrins are mechanosensors that test their ligands across the platelet-platelet synapse. To investigate this model, we interrogate human platelets using approaches that include the supported lipid bilayer platform as well as DNA tension sensor technologies. Experiments suggest that platelet integrins require lateral forces to mediate platelet-platelet interactions. Mechanically labile ligands dampen platelet activation, and the onset of piconewton integrin tension coincides with calcium flux. Activated platelets display immobilized fibrinogen on their surface, thus mediating further recruitment of resting platelets. The distribution of integrin tension was shown to be spatially regulated through two myosin-signaling pathways, myosin light chain kinase and Rho-associated kinase. Finally, we discovered that the termination of integrin tension is coupled with the exposure of phosphatidylserine. Our work reveals the highest spatial and temporal resolution maps of platelet integrin mechanics and its role in platelet aggregation, suggesting that platelets are physical substrates for one another that establish mechanical feedback loops of activation. The results are reminiscent of mechanical regulation of the T-cell receptor, E-cadherin, and Notch pathways, suggesting a common feature for signaling at cell junctions.

Platelets exploit fibrillar adhesions to assemble fibronectin matrix revealing new force-regulated thrombus remodeling mechanisms

“…These three contractile phenotypes are not directly congruent with the well-known platelet phenotypes defined by α-/δ-granule secretion or PS exposure (van der Meijden and Heemskerk 2019). On one hand, more than one biochemical phenotype has low contractility, such as resting but also PS positive platelets (Zhang et al 2018;Nechipurenko et al 2019;Agbani et al 2015). On the other hand, little is known about the contractility of platelets that have or have not undergone degranulation.…”

“…The platelet kindlin 3 equally promotes integrin β1 and β3 clustering (Malinin et al 2009) and talin binds either integrin, ruling out a deficiency for a specific adaptor protein for α5β1 in the myeloid lineage (Humphries et al 2009). With ~1'500 copies per cell (Zeiler, Moser, and Mann 2014) and a typical average force of ~5 pN per integrin (Zhang et al 2018), all α5β1 integrins together could only sustain 7.5 nN over prolonged time. This suggests that α5β1 integrins on platelets are merely outcompeted by the abundant αIIbβ3 during Fn fibrillogenesis.…”

“…Platelets contract rapidly when they get in contact with immobilized fibrinogen and reach maximal forces within 15 minutes (Hansen et al 2018). Actomyosin-based contractility mediates the probing and sensing of substrate stiffness, which modulates platelet activation (Hansen et al 2018;Zhang et al 2018) and drives the bundling and alignment of fibrin fibers (Ono et al 2008) and thus, clot compaction.…”

DNA Nanotechnology as an Emerging Tool to Study Mechanotransduction in Living Systems

“…C) Time lapse images of a spreading human platelet and associated integrin tension signal. Reproduced with permission . Copyright 2018, National Academy of Sciences, USA.…”

“…This hairpin force probe technology was recently adapted to investigate the spatiotemporal distribution of integrin forces during platelet activation, which is a key initial event in blood clotting. We found that platelets used their α II b β3 ‐integrins to generate two populations of force patterns: a central zone with integrin forces >19 pN and a ring of peripheral forces along the lamellipodial edge with the magnitudes between 4.7 and 13.1 pN (Figure C).…”