MicroRNA-148a-3p and TULA-2 regulate platelet FcγRIIA signaling and heparininduced thrombocytopenia
FcγRIIA-mediated platelet activation is essential in heparin-induced thrombocytopenia (HIT) and other immune-mediated thrombocytopenia and thrombosis disorders. There is considerable inter-individual variation in platelet FcγRIIA activation, the reasons for which remain unclear. We hypothesized that genetic variations between FcγRIIA hyper- and hypo-responders regulate FcγRIIA-mediated platelet reactivity and influence HIT susceptibility. Using unbiased genome-wide expression profiling, we observed that human hypo-responders to FcγRIIA activation showed higher platelet T-cell ubiquitin ligand-2 (TULA-2), (encoded by UBASH3B gene), mRNA expression than hyper-responders. To investigate the role of TULA-2 in the FcγRIIA pathway and HIT pathogenesis, we crossed TULA-2-/- mice with transgenic FcγRIIA +/+ mice. Ablation of TULA-2 resulted in hyperphosphorylation of Syk, LAT, and PLCγ2 in platelets via FcγRIIA activation. Platelet integrin activation was also enhanced in TULA-2-/- murine platelets compared to TULA-2+/+ murine platelets in response to FcγRIIA as well as GPVI activation. Compared to wild-type mice, TULA-2-/- mice showed aggravated antibody-mediated thrombocytopenia, augmented thrombin generation. Tail bleeding time, a readout for in vivo hemostasis, was shortened by TULA-2 downregulation. In contrast, there was no significant difference between TULA-2-/- and TULA-2 +/+ platelets in platelet spreading and clot retraction. Of note, heterozygous TULA-2+/- mice, whose platelets contained 50% as much protein as TULA-2+/+ platelets, showed significantly increased platelet reactivity and more severe thrombocytopenia in vivo compared with TULA-2+/+ mice. We also found that miR-148a-3p targeted and inhibited both human and mouse TULA-2 mRNA. Inhibition of miR-148a in FcγRIIA+/+ transgenic mice up-regulated the TULA-2 level and reduced FcγRIIA- and GPVI-mediated platelet αIIbβ3 activation and calcium mobilization. Anti-miR-148a also reduced thrombus formation following intravascular platelet activation via FcγRIIA. These results show for the first time that TULA-2, a target of miR-148a-3p, serves as a negative regulator of FcγRIIA-mediated platelet activation. This study is also the first to show the effects of in vivo miRNA inhibition on platelet reactivity. Our work suggests that modulating miR-148a expression is a potential therapeutic approach for thrombosis.
Zhou, Yuhang, "MicroRNA-148a-3p and TULA-2 regulate platelet FcγRIIA signaling and heparininduced thrombocytopenia" (2016). ETD Collection for Thomas Jefferson University. AAI10170535.