State Key Laboratory of Molecular Biology.
Mol Cell Biol. 2013 Oct;33(20):4095-105. doi: 10.1128/MCB.00418-13. Epub 2013 Aug 19.
Ubiquitination plays important and diverse roles in modulating protein functions. As a C2-WW-HECT-type ubiquitin ligase, Smad ubiquitination regulatory factor 1 (Smurf1) commonly serves to regulate ubiquitin-dependent protein degradation in a number of signaling pathways. Here, we report a novel function of Smurf1 in regulating Wnt/β-catenin signaling through targeting axin for nonproteolytic ubiquitination. Our data unambiguously demonstrate that Smurf1 ubiquitinates axin through Lys 29 (K29)-linked polyubiquitin chains. Unexpectedly, Smurf1-mediated axin ubiquitination does not lead to its degradation but instead disrupts its interaction with the Wnt coreceptors LRP5/6, which subsequently attenuates Wnt-stimulated LRP6 phosphorylation and represses Wnt/β-catenin signaling. The inhibitory function of Smurf1 on Wnt/β-catenin signaling is further evidenced by analysis with Smurf1 knockout murine embryonic fibroblasts. We next identified K789 and K821 in axin as the ubiquitination sites by Smurf1. Consistently, Smurf1 could neither disrupt the interaction of an axin(K789/821R) double mutant with LRP5/6 nor attenuate the phosphorylation of LRP6 in axin(K789/821R)-expressing cells. Collectively, our studies uncover Smurf1 as a new regulator for the Wnt/β-catenin signaling pathway via modulating the activity of axin.
泛素化在调节蛋白质功能方面发挥着重要而多样的作用。Smad 泛素化调节因子 1（Smurf1）作为一种 C2-WW-HECT 型泛素连接酶，通常在许多信号通路中调节泛素依赖性蛋白质降解。在这里，我们报告了 Smurf1 通过靶向轴蛋白进行非蛋白水解泛素化来调节 Wnt/β-catenin 信号的新功能。我们的数据明确表明 Smurf1 通过赖氨酸 29（K29）连接的多泛素链泛素化轴蛋白。出乎意料的是，Smurf1 介导的轴蛋白泛素化不会导致其降解，而是破坏其与 Wnt 核心受体 LRP5/6 的相互作用，从而减弱 Wnt 刺激的 LRP6 磷酸化并抑制 Wnt/β-catenin 信号。Smurf1 对 Wnt/β-catenin 信号的抑制功能进一步通过 Smurf1 敲除鼠胚胎成纤维细胞的分析得到证实。我们接下来通过 Smurf1 鉴定了轴蛋白中的 K789 和 K821 作为泛素化位点。一致地，Smurf1 既不能破坏 axin(K789/821R)双突变体与 LRP5/6 的相互作用，也不能减弱 axin(K789/821R)表达细胞中 LRP6 的磷酸化。总之，我们的研究揭示了 Smurf1 通过调节轴蛋白的活性成为 Wnt/β-catenin 信号通路的新调节剂。
