Environmental Health Science Laboratory, Sumitomo Chemical Company Limited, Osaka 554-8558, Japan.
J Biol Chem. 2011 Feb 11;286(6):4485-92. doi: 10.1074/jbc.M110.151936. Epub 2010 Nov 28.
Transforming growth factor-β (TGF-β) is considered to be a major factor contributing to liver fibrosis. We have previously shown that nuclear translocation of YB-1 antagonizes the TGF-β/Smad3 signaling in regulating collagen gene expression. More recently, we have demonstrated that the novel small compound HSc025 promotes nuclear translocation of YB-1, resulting in the improvement of skin and pulmonary fibrosis. Here, we presented evidence as to the mechanism by which HSc025 stimulates nuclear translocation of YB-1 and the pharmacological effects of HSc025 on a murine model of hepatic fibrosis. A proteomics approach and binding assays using HSc025-immobilized resin showed that HSc025 binds to the amino acid sequence within the C-tail region of YB-1. In addition, immunoprecipitation experiments and glutathione S-transferase pulldown assays identified poly(A)-binding protein (PABP) as one of the cytoplasmic anchor proteins of YB-1. HSc025 directly binds to YB-1 and interrupts its interaction with PABP, resulting in accelerated nuclear translocation of YB-1. Transfection of cells with PABP siRNA promoted nuclear translocation of YB-1 and subsequently inhibited basal and TGF-β-stimulated collagen gene expression. Moreover, HSc025 significantly suppressed collagen gene expression in cultured activated hepatic stellate cells. Oral administration of HSc025 to mice with carbon tetrachloride-induced hepatic fibrosis improved liver injury as well as the degree of hepatic fibrosis. Altogether, the results provide a novel insight into therapy for organ fibrosis using YB-1 modulators.
转化生长因子-β(TGF-β)被认为是导致肝纤维化的主要因素。我们之前已经表明,YB-1 的核转位拮抗 TGF-β/Smad3 信号通路调节胶原基因表达。最近,我们已经证明,新型小分子化合物 HSc025 促进 YB-1 的核转位,从而改善皮肤和肺纤维化。在这里,我们提出了 HSc025 刺激 YB-1 核转位的机制以及 HSc025 对肝纤维化小鼠模型的药理作用的证据。一种蛋白质组学方法和使用 HSc025 固定树脂的结合测定表明,HSc025 与 YB-1 的 C 尾区域内的氨基酸序列结合。此外,免疫沉淀实验和谷胱甘肽 S-转移酶 pulldown 测定鉴定了多聚(A)结合蛋白(PABP)为 YB-1 的细胞质锚定蛋白之一。HSc025 直接与 YB-1 结合并中断其与 PABP 的相互作用,导致 YB-1 的核转位加速。用 PABP siRNA 转染细胞可促进 YB-1 的核转位,并随后抑制基础和 TGF-β 刺激的胶原基因表达。此外,HSc025 显著抑制培养的活化肝星状细胞中的胶原基因表达。HSc025 对四氯化碳诱导的肝纤维化小鼠的口服给药可改善肝损伤和肝纤维化程度。总之,这些结果为使用 YB-1 调节剂治疗器官纤维化提供了新的见解。