Cerebral amyloid angiopathy (CAA) is a vascular dysfunction disorder characterized by deposits of amyloid-β (Aβ) in the walls of cerebral vessels. CAA and Aβ deposition in the brain parenchyma contribute to dementia and Alzheimer's disease (AD). We investigated the contribution of platelets, which accumulate at vascular Aβ deposits, to CAA. We found that synthetic monomeric Aβ40 bound through its RHDS (Arg-His-Asp-Ser) sequence to integrin αIIbβ3, which is the receptor for the extracellular matrix protein fibrinogen, and stimulated the secretion of adenosine diphosphate (ADP) and the chaperone protein clusterin from platelets. Clusterin promoted the formation of fibrillar Aβ aggregates, and ADP acted through its receptors P2Y1 and P2Y12 on platelets to enhance integrin αIIbβ3 activation, further increasing the secretion of clusterin and Aβ40 binding to platelets. Platelets from patients with Glanzmann's thrombasthenia, a bleeding disorder in which platelets have little or dysfunctional αIIbβ3, indicated that the abundance of this integrin dictated Aβ-induced clusterin release and platelet-induced Aβ aggregation. The antiplatelet agent clopidogrel, which irreversibly inhibits P2Y12, inhibited Aβ aggregation in platelet cultures; in transgenic AD model mice, this drug reduced the amount of clusterin in the circulation and the incidence of CAA. Our findings indicate that activated platelets directly contribute to CAA by promoting the formation of Aβ aggregates and that Aβ, in turn, activates platelets, creating a feed-forward loop. Thus, antiplatelet therapy may alleviate fibril formation in cerebral vessels of AD patients.

Integrins are α/β heterodimeric transmembrane adhesion receptors. Evidence exists that their transmembrane domain (TMD) separates upon activation. Subunit-specific differences in activation sensitivity of integrins were reported. However, whether sequence variations in the TMD lead to differential TMD association has remained elusive. Here, we show by molecular dynamics simulations and association free energy calculations on TMDs of integrin αIIbβ3, αvβ3, and α5β1 that αIIbβ3 TMD is most stably associated; this difference is related to interaction differences across the TMDs. The order of TMD association stability is paralleled by the basal activity of these integrins, which suggests that TMD differences can have a decisive effect on integrin conformational free energies. We also identified a specific order of clasp disintegration upon TMD dissociation, which suggests that the closed state of integrins may comprise several microstates. Our results provide unprecedented insights into a possibly contributing role of TMD towards subunit-specific sensitivity of integrin activation.

Integrins are heterodimeric cell-adhesion receptors comprising α and β subunits that transmit signals allosterically in both directions across the membrane by binding to intra-and extracellular components. The human platelet antigen-1 (HPA-1) polymorphism in α IIb β 3 arises from a Leu→Pro exchange at residue 33 in the genu of the β3 subunit, resulting in or Pro-33 (HPA1b) isoforms. Although clinical investigations have provided conflicting results, some studies have suggested that Pro-33 platelets exhibit increased thrombogenicity. Under flow-dynamic conditions, the Pro-33 variant displays prothrombotic properties, characterized by increased platelet adhesion, aggregate/thrombus formation, and outside-in signaling. However, the molecular events underlying this prothrombotic phenotype have remained elusive. As residue 33 is located > 80 Å away from extracellular binding sites or transmembrane domains, we hypothesized that the Leu→Pro exchange allosterically shifts the dynamic conformational equilibrium of αIIbβ3 toward an active state. Multiple microsecond-long, allatom molecular dynamics simulations of the ectodomain of the Leu-33 and Pro-33 isoforms provided evidence that the Leu→Pro exchange weakens interdomain interactions at the genu and alters the structural dynamics of the integrin to a more unbent and splayed state. Using FRET analysis of fluorescent proteins fused with αIIbβ3 in transfected HEK293 cells, we found that the Pro-33 variant in its resting state displays a lower energy transfer http://www.jbc.org/cgi