摘要
抄写因素 Oct4 在维持 pluripotency 并且控制胚胎的干细胞(转换字符) 的系承诺起关键作用。我们的以前的学习显示那 Wwp2，鼠标 HECT 类型 E3 ubiquitin ligase， ubiquitinates Oct4 并且在一个 heterologous 系统支持它的降级。然而，在象 Wwp2 的功能的分子的特征一样调整内长的 Oct4 蛋白质层次的 Wwp2 的角色一直不是坚定的。这里，我们报导 Wwp2 在 embryonal 癌房间(ECC ) 的区别期间在 Oct4 ubiquitination 和降级起一个重要作用，尽管它不看起来在无差别的 ECC 和转换字符影响 Oct4 蛋白质层次。重要地，由特定的 RNA 干扰的 Wwp2 表示的抑制提高 Oct4 蛋白质水平，导致在 retinoid 的变细区别相关的标记基因的导致酸的激活。机械学地， Wwp2 经由离氨酸催化 Oct4 poly-ubiquitination 以一种剂量依赖者方式的 63 连接。有趣地， Wwp2 也以一种类似的方式调整它的自己的 ligase 活动。而且， Wwp2 的 auto-ubiquitination 通过 intra 分子的机制发生。一起拿，这些结果在在 ECC 的区别过程期间控制内长的 Oct4 蛋白质层次表明 Wwp2 的一个关键角色并且为调整 E3 ubiquitin ligase 的催化活动建议有趣的剂量依赖者机制， Wwp2。
Transcription factor Oct4 plays critical roles in maintaining pluripotency and controlling lineage commitment of embryonic stem cells （ESCs）. Our previous study indicates that Wwp2, a mouse HECT-type E3 ubiquitin ligase, ubiquitinates Oct4 and promotes its degradation in a heterologous system. However, roles of Wwp2 in regulating en- dogenous Oct4 protein levels as well as molecular characteristics of the function of Wwp2 have not been determined. Here, we report that Wwp2 plays an important role in Oct4 ubiquitination and degradation during differentiation of embryonal carcinoma cells （ECCs）, although it does not appear to affect Oct4 protein levels in the undifferentiated ECCs and ESCs. Importantly, inhibition of Wwp2 expression by specific RNA interference elevates the Oct4 protein level, leading to attenuation in retinoid acid-induced activation of differentiation-related marker genes. Mechanisti- cally, Wwp2 catalyzes Oct4 poly-ubiquitination via the lysine 63 linkage in a dosage-dependent manner. Interest- ingly, Wwp2 also regulates its own ligase activity in a similar manner. Moreover, auto-ubiquitination of Wwp2 occurs through an intra-molecular mechanism. Taken together, these results demonstrate a crucial role of Wwp2 in con- trolling endogenous Oct4 protein levels during differentiation processes of ECCs and suggest an interesting dosage- dependent mechanism for regulating the catalytic activity of the E3 ubiquitin ligase, Wwp2.
基金
We thank Dr Richard Baer （Pathology, Columbia University, New York, USA） for generously providing various Ub mutant plasmids. This study was supported by Grants from the National Natural Science Foundation of China （30871257, 30730051） and the National High Technology Research, Development Program of China （2006CB943901 and 2007CB947904）, the Shanghai Sci- ence and Technology Developmental Foundation （08JC1413100） and the Shanghai Leading Academic Discipline Project （$30201）.
