Tao Hong, Li Haiying, Su Yanhuang, Feng Danni, Wang Xilong, Zhang Chun, Ma Hui, Hu Qikuan
Department of Physiology, Ningxia Medical University, 1160 Shengli Street, Yinchuan, 750004, Ningxia, China.
Mol Cell Biochem. 2014 Sep;394(1-2):23-30. doi: 10.1007/s11010-014-2077-4. Epub 2014 May 16.
Epigenetic modification is crucial to keep the self-renewal and the "stemness" states of stem cells, not letting them to differentiate. The actual roles of Histone 3 Lysine 9 dimethylation (H3K9me2) and its methyltransferase G9a in this process are still unclear, especially in cancer stem cells. In our study, we found an interesting observation that most CD133-positive cells were H3K9me2 negative, both in glioma tissues and in cultured cells, although most cancer cells were detected to be H3K9me2 immunopositive. This implied that the G9a-dependent H3K9me2 was one of the crucial barriers of cancer stem cell self-renewal. To test the hypothesis, we examined the loss-of-function and gain-of-function of G9a. We found that bix01294, the selective inhibitor of G9a, can stimulate the sphere formation rate of glioma cancer stem cells, together with increasing Sox2 and CD133 expressions. The increase of CD133-active stem cells was confirmed by flow cytometry. On the other aspect, overexpression of G9a increased the H3K9me2 and decreased the sphere formation rate as well as the CD133 and Sox2 expressions. Since H3K9me2 modification is the major repressive switch, we predict that the repressive H3K9me2 modification may happen at the CD133 promoter regions. By chromatin precipitation assay, we confirmed that the CD133 and Sox2 promoter regions were modified by the H3K9me2. Therefore, we concluded that the G9a-dependent H3K9me2 repression on CD133 and Sox2 was one of the main switches of the self-renewal in glioma cancer stem cells.
表观遗传修饰对于维持干细胞的自我更新和“干性”状态至关重要，可防止其分化。组蛋白3赖氨酸9二甲基化（H3K9me2）及其甲基转移酶G9a在此过程中的实际作用仍不清楚，尤其是在癌症干细胞中。在我们的研究中，我们发现了一个有趣的现象：在胶质瘤组织和培养细胞中，大多数CD133阳性细胞的H3K9me2呈阴性，尽管大多数癌细胞的H3K9me2免疫呈阳性。这表明G9a依赖性H3K9me2是癌症干细胞自我更新的关键障碍之一。为了验证这一假设，我们检测了G9a的功能丧失和功能获得情况。我们发现，G9a的选择性抑制剂bix01294可以刺激胶质瘤癌症干细胞的球形成率，同时增加Sox2和CD133的表达。流式细胞术证实了CD133活性干细胞的增加。另一方面，G9a的过表达增加了H3K9me2，降低了球形成率以及CD133和Sox2的表达。由于H3K9me2修饰是主要的抑制开关，我们预测抑制性H3K9me2修饰可能发生在CD133启动子区域。通过染色质沉淀试验，我们证实CD133和Sox2启动子区域被H3K9me2修饰。因此，我们得出结论，G9a依赖性H3K9me2对CD133和Sox2的抑制是胶质瘤癌症干细胞自我更新的主要开关之一。
