Drahos, Karen E.Welsh, John D.Finkielstein, Carla V.Capelluto, Daniel G. S.2018-11-142018-11-142009-11-24e8007http://hdl.handle.net/10919/85840Background Platelets contact each other at the site of vascular injury to stop bleeding. One negative regulator of platelet aggregation is Disabled-2 (Dab2), which is released to the extracellular surface upon platelet activation. Dab2 inhibits platelet aggregation through its phosphotyrosine-binding (PTB) domain by competing with fibrinogen for αIIbβ3 integrin receptor binding by an unknown mechanism. Methodology/Principal Findings Using protein-lipid overlay and liposome-binding assays, we identified that the N-terminal region of Dab2, including its PTB domain (N-PTB), specifically interacts with sulfatides. Moreover, we determined that such interaction is mediated by two conserved basic motifs with a dissociation constant (Kd) of 0.6 µM as estimated by surface plasmon resonance (SPR) analysis. In addition, liposome-binding assays combined with mass spectroscopy studies revealed that thrombin, a strong platelet agonist, cleaved N-PTB at a site located between the basic motifs, a region that becomes protected from thrombin cleavage when bound to sulfatides. Sulfatides on the platelet surface interact with coagulation proteins, playing a major role in haemostasis. Our results show that sulfatides recruit N-PTB to the platelet surface, sequestering it from integrin receptor binding during platelet activation. This is a transient recruitment that follows N-PTB internalization by an actin-dependent process. Conclusions/Significance Our experimental data support a model where two pools of Dab2 co-exist at the platelet surface, in both sulfatide- and integrin receptor-bound states, and their balance controls the extent of the clotting response.application/pdfenCreative Commons Attribution 4.0 InternationalArticle - Refereedhttps://doi.org/10.1371/journal.pone.0008007411199566251932-6203