Galectin-9 activates platelet ITAM receptors glycoprotein VI and C-type lectin-like receptor-2

Z.G. Zhi, N.J. Jooss, Y. Sun, M. Colicchia, A. Slater, L.A. Moran, H.Y.F. Cheung, Y. Di, J. Rayes, N.S. Poulter, S.P. Watson*, A.J. Iqbal*

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review


Background Platelets are multifunctional cellular mediators in many physiological and pathophysiological processes such as thrombosis, angiogenesis, and inflammation. Several members of galectins, a family of carbohydrate-binding proteins with a broad range of immunomodulatory actions, have been reported to activate platelets. Objective In this study, we investigated the role of galectin-9 (Gal-9) as a novel ligand for platelet glycoprotein VI (GPVI) and C-type lectin-like receptor 2 (CLEC-2). Methods Platelet spreading, aggregation, and P-selectin expression in response to Gal-9 were measured in washed platelet suspensions via static adhesion assay, light transmission aggregometry, and flow cytometry, respectively. Solid-phase binding assay and protein phosphorylation studies were utilized to validate the interaction between Gal-9 and GPVI, and immunoprecipitation for detecting CLEC-2 phosphorylation. Wild-type (WT), GPVI-knockout (Gp6(-/-)), and GPVI and CLEC-2-double knockout (Gp6(-/-)/Gp1ba-Cre-Clec1b(fl)(/)(fl)) mice were used. Results We have shown that recombinant Gal-9 stimulates aggregation in human and mouse washed platelets dose-dependently. Platelets from both species adhere and spread on immobilized Gal-9 and express P-selectin. Gal-9 competitively inhibited the binding of human recombinant D1 and D2 domains of GPVI to collagen. Gal-9 stimulated tyrosine phosphorylation of CLEC-2 and proteins known to lie downstream of GPVI and CLEC-2 including spleen tyrosine kinase and linker of activated T cells in human platelets. GPVI-deficient murine platelets exhibited significantly impaired aggregation in response to Gal-9, which was further abrogated in GPVI and CLEC-2-double-deficient platelets. Conclusions We have identified Gal-9 as a novel platelet agonist that induces activation through interaction with GPVI and CLEC-2.
Original languageEnglish
Pages (from-to)936-950
Number of pages15
JournalJournal of Thrombosis and Haemostasis
Issue number4
Early online date6 Jan 2022
Publication statusPublished - Apr 2022


  • C-type lectin-like receptor 2
  • galectin-9
  • glycoprotein VI
  • immunoreceptor tyrosine-based activation motif
  • platelet
  • GPVI


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