Seo Seung Un, Woo Seon Min, Lee Seul Gi, Kim Min Yeong, Lee Hyun Shik, Choi Yung Hyun, Kim Sang Hyun, Chang Young-Chae, Min Kyoung-Jin, Kwon Taeg Kyu
Department of Immunology, School of Medicine, Keimyung University, Daegu, 42601, South Korea.
Department of Biochemistry, College of Korean Medicine, Dong-Eui University, Busan, 47227, South Korea.
Redox Biol. 2022 Jul;53:102336. doi: 10.1016/j.redox.2022.102336. Epub 2022 May 13.
Cathepsin K inhibitor (odanacatib; ODN) and cathepsin K knockdown (siRNA) enhance oxaliplatin-induced apoptosis through p53-dependent Bax upregulation. However, its underlying mechanisms remain unclear. In this study, we elucidated the mechanism behind enhancement of oxaliplatin-induced apoptosis by ODN. We also investigated the molecular mechanisms of ODN-induced Bax upregulation. Here, we demonstrated that ODN-induced Bax upregulation required p53, but it was independent of p53 transcriptional activity. Various mutants of the DNA-binding domain of p53 induced Bax upregulation in ODN-treated cells. p53 functional domain analysis showed that the C-terminal domain of p53 participates in the physical interaction and stabilization of Sp1, a major transcription factor of Bax. We screened a specific siRNA encoding 50 deubiquitinases and identified that BAP1 stabilizes Sp1. The knockdown or catalytic mutant form of BAP1 abolished the ODN-induced upregulation of Sp1 and Bax expression. Mechanistically, ODN induced BAP1 phosphorylation and enhanced Sp1-BAP1 interaction, resulting in Sp1 ubiquitination and degradation. Interestingly, ODN-induced BAP1 phosphorylation and DNA damage were modulated by the production of mitochondrial reactive oxygen species (ROS). Mitochondrial ROS scavengers prevented DNA damage, BAP1-mediated Sp1 stabilization, and Bax upregulation by ODN. BAP1 downregulation by siRNA inhibited apoptosis induced by the combined treatment of ODN and oxaliplatin/etoposide. Therefore, Sp1 is a crucial transcription factor for ODN-induced Bax upregulation, and Sp1 stabilization is regulated by BAP1.
组织蛋白酶K抑制剂（奥达卡替；ODN）和组织蛋白酶K基因敲低（小干扰RNA）通过p53依赖的Bax上调增强奥沙利铂诱导的细胞凋亡。然而，其潜在机制仍不清楚。在本研究中，我们阐明了ODN增强奥沙利铂诱导的细胞凋亡背后的机制。我们还研究了ODN诱导Bax上调的分子机制。在此，我们证明ODN诱导的Bax上调需要p53，但它独立于p53转录活性。p53 DNA结合结构域的各种突变体在ODN处理的细胞中诱导Bax上调。p53功能结构域分析表明，p53的C末端结构域参与了Bax主要转录因子Sp1的物理相互作用和稳定。我们筛选了一种编码50种去泛素化酶的特异性小干扰RNA，并确定BAP1可稳定Sp1。BAP1的基因敲低或催化突变形式消除了ODN诱导的Sp1上调和Bax表达。机制上，ODN诱导BAP1磷酸化并增强Sp1-BAP1相互作用，导致Sp1泛素化和降解。有趣的是，ODN诱导的BAP1磷酸化和DNA损伤受到线粒体活性氧（ROS）产生的调节。线粒体ROS清除剂可防止DNA损伤、BAP1介导的Sp1稳定以及ODN诱导的Bax上调。小干扰RNA介导的BAP1下调抑制了ODN与奥沙利铂/依托泊苷联合治疗诱导的细胞凋亡。因此，Sp1是ODN诱导Bax上调的关键转录因子，Sp1的稳定受BAP1调节。
