Pivotal Role of the C2 Domain of the Smurf1 Ubiquitin Ligase in Substrate Selection
Pivotal Role of the C2 Domain of the Smurf1 Ubiquitin Ligase in Substrate Selection
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Smurf1 泛素连接酶的 C2 结构域在底物选择中的关键作用
DOI：
10.1074/jbc.m110.211979
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发表时间：
2011-05-13
影响因子：
4.8
通讯作者：
He, Fuchu
中科院分区：
文献类型：
--
作者：
Lu, Kefeng;Li, Ping;He, Fuchu
C2-WW-HECT型泛素连接酶Smurf 1和Smurf 2通过调节骨形态发生蛋白、Wnt和RhoA信号通路在胚胎发生和成人骨稳态中发挥关键作用。C2和HECT结构域之间的分子内相互作用自动抑制Smurf 2的连接酶活性。然而，Smurf 1 C2结构域的作用仍然难以捉摸。在这里，我们表明，C2-HECT的自抑制机制是没有观察到Smurf 1，而是其C2结构域的功能在底物选择。Smurf 1 C2结构域在质膜定位中发挥关键作用，并赋予Smurf 1对RhoA与Smad 5和Runx 2的差异活性。晶体结构分析表明，Smurf 1 C2结构域具有典型的反平行β-夹心折叠。硫酸盐结合位点分析的检查揭示了C2结构域内的两个关键赖氨酸残基，Lys-28和Lys-85，这对于Smurf 1在质膜上的定位、对细胞迁移的调节和对RhoA的稳健连接酶活性是重要的，这进一步支持了Smurf 1的Ca 2+非依赖性定位机制。这些发现证明了Smurf 1 C2结构域在底物选择和细胞定位中先前未识别的作用。
The C2-WW-HECT-type ubiquitin ligases Smurf1 and Smurf2 play a critical role in embryogenesis and adult bone homeostasis via regulation of bone morphogenetic protein, Wnt, and RhoA signaling pathways. The intramolecular interaction between C2 and HECT domains autoinhibits the ligase activity of Smurf2. However, the role of the Smurf1 C2 domain remains elusive. Here, we show that the C2-HECT autoinhibition mechanism is not observed in Smurf1, and instead its C2 domain functions in substrate selection. The Smurf1 C2 domain exerts a key role in localization to the plasma membrane and endows Smurf1 with differential activity toward RhoA versus Smad5 and Runx2. Crystal structure analysis reveals that the Smurf1 C2 domain possesses a typical anti-parallel beta-sandwich fold. Examination of the sulfate-binding site analysis reveals two key lysine residues, Lys-28 and Lys-85, within the C2 domain that are important for Smurf1 localization at the plasma membrane, regulation on cell migration, and robust ligase activity toward RhoA, which further supports a Ca2+-independent localization mechanism for Smurf1. These findings demonstrate a previously unidentified role of the Smurf1 C2 domain in substrate selection and cellular localization.