The human platelet antigen-1b (Pro33) variant of αIIbβ3 allosterically shifts the dynamic conformational equilibrium of this integrin toward the active state

Abstract

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

Platelet Signaling in Primary Haemostasis and Arterial Thrombus Formation: Part 2

“…Thus, the fully active state of αIIbβ3 is promoted, thereby fostering the prothrombotic phenotype of Pro33 (HPA-1b) platelets. 50 kinases (SFK), entirely blocks activation of ERK2 (data not shown), providing indirect evidence that the ERK2 Tyr/Thr phosphorylation is mediated through a c-Src tyrosine kinases pathway. 51 Moreover, specific Rho inhibitors abrogate ERK2 activation in both αIIbβ3 isoforms (►Fig.…”

“…43 These results suggest that the Leu!Pro exchange may modulate functional properties of αIIbβ3, resulting in a prothrombotic integrin variant. 44 Indeed, in subsequent experiments under flow-dynamic conditions, it was documented that Pro33 (HPA-1b) platelets and αIIbβ3 (Pro33)-transfected cells display a prothrombotic phenotype, as characterized by increased adhesion activity, 45,46 increased resistance of adherent transfectants to high shear, 47,48 increased aggregate/thrombus formation 49,50 and increased outside-in signaling. 51,52 Recently, the molecular nature of this prothrombotic integrin variant was elucidated.…”

“…This effect alters the structural dynamics of αIIbβ3 to a more bent and splayed state, which results in a conformation that is closer to the active one, promoting the fully active state and fostering the prothrombotic phenotype of Pro33 platelets. 50…”

“…Metal ions are required for the binding of fibrinogen and VWF to β3 integrins, 12 and using Mn 2þ is an established experimental tool to induce an active 'unbent' conformation of αIIbβ3 by acting via three metal ion binding sites in the β3subunit, notably MIDAS (metal ion-dependent adhesion site). 53 Based on the molecular dynamics analysis, demonstrating an allosteric shift of the Pro33 variant toward an active integrin, 50 it is now possible to provide a concise explanation for the observed differences in c-Src activation between the two isoforms of αIIbβ3.…”

Genetic polymorphisms in early-onset myocardial infarction in a sample of Iraqi patients: a pilot study

“…Plasminogen activator inhibitor type 1 (PAI-1) is important in the regulation of endogenous fibrinolysis, and an increase in its circulating levels may be associated with the progression of atheromas and thrombosis [ 13 ]. A single nucleotide polymorphism of the GPIIIa gene causes Leu33Pro changing the human platelets antigen 1(HPA1) from 1a to 1b, with altered antigenic properties and an associated increase in platelet reactivity, thus fostering a prothrombotic state [ 14 ]. On the other hand, a polymorphism in the angiotensin-converting enzyme (ACE) gene characterized by insertion (I) or deletion (D) within its intron 16 has been described, with the D allele being associated with higher ACE levels and possible increased risk of coronary artery disease [ 15 ].…”