TULA-2 Deficiency Enhances Platelet Functional Responses to CLEC-2 Agonists

• Published in: TH open : companion journal to thrombosis and haemostasis 2018-10, Vol.2 (4), p.e411-e419
• Main Authors: Kostyak, John C., Mauri, Benjamin R., Dangelmaier, Carol, Patel, Akruti, Zhou, Yuhang, Eble, Johannes A., Tsygankov, Alexander Y., McKenzie, Steven E., Kunapuli, Satya P.
• Format: Article
• Language: English
• Subjects: Original; Original Article; platelets; thrombosis; thromboxane a2
• ISSN: 2512-9465; 2567-3459; 2512-9465
• DOI: 10.1055/s-0038-1676358

Abstract Platelet activation is essential for hemostasis. Central to platelet activation are the signals transmitted through surface receptors such as glycoprotein VI, the protease-activated receptors, and C-type lectin-like receptor 2 (CLEC-2). CLEC-2 is a HemITAM (hem-immunoreceptor tyrosine activation motif)-bearing receptor that binds podoplanin and signals through spleen tyrosine kinase (Syk). T-cell ubiquitin ligand-2 (TULA-2) is a protein tyrosine phosphatase that is highly expressed in platelets and targets phosphorylated Y352 of Syk. We wanted to determine whether TULA-2 regulates Syk phosphorylation and activity downstream of CLEC-2. To that end, we used TULA-2 knockout mice and wild-type (WT) littermate controls. We found that TULA-2 deficiency enhances the aggregation and secretion response following stimulation with an excitatory CLEC-2 antibody or the CLEC-2 agonist rhodocytin. Consistently, Syk phosphorylation of Y346 is enhanced, as well as phosphorylation of the downstream signaling molecule PLCγ2, in TULA-2 knockout platelets treated with either CLEC-2 antibody or rhodocytin, compared with WT control platelets. Furthermore, the kinetics of Syk phosphorylation, as well as that of PLCγ2 and SLP-76, is enhanced in TULA-2 knockout platelets treated with 2.5-μg/mL CLEC-2 antibody compared with WT platelets. Similarly, thromboxane production was enhanced, in both amount and kinetics, in TULA-2 −/− platelets treated with 2.5-μg/mL CLEC-2 antibody. TULA-2 acts as a negative regulator of CLEC-2 signaling by dephosphorylating Syk on Y346 and restraining subsequent Syk-mediated signaling.