Background: Platelets have the hemostatic function, and their aberrant activation is associated with occlusive thrombus formation. Plasma exosomes are rich in platelets containing ubiquitin-specific peptidase 15 (USP15). Herein, we aim to explore the effect of USP15 on thrombosis, as well as expounding whether USP15 acts as an upstream target of FK506 binding protein 5 (FKBP5) to regulate occlusive thrombus formation. Methods: Washed human platelets were treated with thrombin for measurement of USP15 and FKBP5 expressions. USP15 loss/gain-of-function variant in HEK293 cells was performed by cell transfection, and the interaction between USP15 and FKBP5 was examined using immunoprecipitation and ubiquitination assays. Mice with USP15-knockout platelets (Plt USP15-/-) were modeled, and subjected to calculation of bleeding time, artery thrombosis imaging and clot retraction assay. FKBP5 expression and the inhibitor of nuclear factor kappa B kinase subunit epsilon (IKBKE)/phosphatidylinositol 3-kinase (PI3K)/Rap1 pathway in wild-type and Plt USP15-/- mice-derived platelets were detected using Western blot. The activation of αIIbβ3 in washed platelets was analyzed using flow cytometry. Results: USP15 and FKBP5 expressions were upregulated in platelets after thrombin treatment. Following transfection of USP15 knockdown and USP15 overexpression plasmids into HEK293 cells, FKBP5 protein expression was downregulated by USP15 knockdown while being upregulated by USP15 overexpression. USP15 bound to FKBP5 and protected FKBP5 against ubiquitination. Knockdown of platelet USP15 prolonged bleeding time, inhibited arterial thrombosis and delayed clot retraction in mice. Knockdown of platelet USP15 also decreased protein expressions of FKBP5, IKBKE and Rap1, p-PI3K/PI3K ratio, and activation of αIIbβ3 in mice. Conclusion: USP15 knockdown in platelets affects thrombosis in mice by promoting the instability of FKBP5 to repress the activation of IKBKE/PI3K/Rap1 pathway-mediated αIIbβ3.
Pulmonary hypertension (PH) is a life-threatening cardiopulmonary disease characterized by pulmonary vascular remodeling. Excessive growth and migration of pulmonary artery smooth muscle cells (PASMCs) are believed to be major contributors to pulmonary vascular remodeling. Ubiquitin-specific protease 15 (USP15) is a vital deubiquitinase that has been shown to be critically involved in many pathologies. However, the effect of USP15 on PH has not yet been explored. In this study, the upregulation of USP15 was identified in the lungs of PH patients, mice with SU5416/hypoxia (SuHx)-induced PH and rats with monocrotaline (MCT)-induced PH. Moreover, adeno-associated virus-mediated functional loss of USP15 markedly alleviated PH exacerbation in SuHx-induced mice and MCT-induced rats. In addition, the abnormal upregulation and nuclear translocation of YAP1/TAZ was validated after PH modeling. Human pulmonary artery smooth muscle cells (hPASMCs) were exposed to hypoxia to mimic PH in vitro, and USP15 knockdown significantly inhibited cell proliferation, migration, and YAP1/TAZ signaling in hypoxic hPASMCs. Rescue assays further suggested that USP15 promoted hPASMC proliferation and migration in a YAP1/TAZ-dependent manner. Coimmunoprecipitation assays indicated that USP15 could interact with YAP1, while TAZ bound to USP15 after hypoxia treatment. We further determined that USP15 stabilized YAP1 by inhibiting the K48-linked ubiquitination of YAP1. In summary, our findings reveal the regulatory role of USP15 in PH progression and provide novel insights into the pathogenesis of PH.
Clear cell renal cell carcinoma (ccRCC) is the most common malignant cancer in kidney. Exploring new therapeutic targets is crucial to improving the ccRCC patients’ overall survival. Kinesin family member 15 (KIF15), a member of the kinesin superfamily proteins, participates in various biological processes. However, the role and action mechanisms of KIF15 in ccRCC remain unknown. We observed that KIF15 was highly expressed in ccRCC tissues and its expression was negatively correlated with the prognosis of ccRCC patients. In vitro experiments showed that KIF15 promoted the proliferation of ccRCC cells. Consistently, in vivo assays showed that KIF15 accelerated tumor growth in ccRCC-bearing nude mice. Further investigation revealed that zinc finger protein 750 (ZNF750) repressed the transcription of KIF15 by directly binding its promoter region, simultaneously repressing ccRCC cell growth. Mechanistically, KIF15 stabilized MDM2 via enhancing USP15-mediated deubiquitylation, thus suppressing the p53 pathway. Our study indicates the oncogenic role and molecular mechanisms of KIF15 in ccRCC progression and provides valuable insights into potential therapeutic strategies for ccRCC.
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