Sialylated glycoproteins bind to Siglec-9 in a cis manner on platelets to suppress platelet activation.

Background: The endogenous negative regulation of platelets is important in preventing spontaneous thrombosis, while the mechanism of homeostasis is incompletely understood.

Objectives: In this study, we aimed to explore whether Siglec-9 plays a negative regulative role and identify the ligand of Siglec-9 on platelets.

Methods: To determine the role of Siglec-9 on platelets, platelet factor 4-cre:Siglec-E knockout mouse model and human platelet in vitro culture system were used. Furthermore, recombinant glycoprotein (GP) of Siglec-9 ligand on platelets was expressed and used.

Results: We found that Siglec-E conditional knockout can lead to significant increase in platelet coagulation activities both in vivo and in vitro, which strongly suggests that Siglec-9/E plays an inhibitory physiological role in platelet activation. Siglec-9 ligand is an O-link GP with an α2,3-linked sialic acid terminal structure, and the protein carrier of the ligand is mucin-like region of GPIbα. Our data further showed that the ligands on platelets could not engage Siglec-9 on other cells via trans-binding, which indicates that the ligands on platelets play a self-modulation role. Furthermore, we provided evidence that the activation of Siglec-9 pathway with exogenous specific ligands could inhibit the activity of platelets. These data demonstrate a previously unanticipated role for GPIbα in inhibiting platelet activation and provide a novel mechanism for the homeostasis of platelets.

Conclusions: We conclude that the cis-binding between mucin-like region of GPIbα and Siglec-9 acts as a "parking brake" on platelet activation. This finding provides a potential druggable target for novel antiplatelet medicine.