Jamieson Cara, Lui Christina, Brocardo Mariana G, Martino-Echarri Estefania, Henderson Beric R
Center for Cancer Research, The Westmead Millennium Institute for Medical Research, The University of Sydney, Westmead, New South Wales 2145, Australia.
Center for Cancer Research, The Westmead Millennium Institute for Medical Research, The University of Sydney, Westmead, New South Wales 2145, Australia
J Cell Sci. 2015 Nov 1;128(21):3933-46. doi: 10.1242/jcs.167742. Epub 2015 Sep 24.
β-Catenin transduces the Wnt signaling pathway and its nuclear accumulation leads to gene transactivation and cancer. Rac1 GTPase is known to stimulate β-catenin-dependent transcription of Wnt target genes and we confirmed this activity. Here we tested the recent hypothesis that Rac1 augments Wnt signaling by enhancing β-catenin nuclear import; however, we found that silencing/inhibition or up-regulation of Rac1 had no influence on nuclear accumulation of β-catenin. To better define the role of Rac1, we employed proximity ligation assays (PLA) and discovered that a significant pool of Rac1-β-catenin protein complexes redistribute from the plasma membrane to the nucleus upon Wnt or Rac1 activation. More importantly, active Rac1 was shown to stimulate the formation of nuclear β-catenin-lymphoid enhancer factor 1 (LEF-1) complexes. This regulation required Rac1-dependent phosphorylation of β-catenin at specific serines, which when mutated (S191A and S605A) reduced β-catenin binding to LEF-1 by up to 50%, as revealed by PLA and immunoprecipitation experiments. We propose that Rac1-mediated phosphorylation of β-catenin stimulates Wnt-dependent gene transactivation by enhancing β-catenin-LEF-1 complex assembly, providing new insight into the mechanism of cross-talk between Rac1 and canonical Wnt/β-catenin signaling.
β-连环蛋白转导Wnt信号通路，其在细胞核内的积累会导致基因转录激活及引发癌症。已知Rac1 GTP酶可刺激Wnt靶基因的β-连环蛋白依赖性转录，我们也证实了这种活性。在此，我们检验了最近的一种假说，即Rac1通过增强β-连环蛋白的核输入来增强Wnt信号；然而，我们发现沉默/抑制或上调Rac1对β-连环蛋白的核积累并无影响。为了更好地界定Rac1的作用，我们采用了邻近连接分析（PLA），并发现大量的Rac1-β-连环蛋白蛋白复合物在Wnt或Rac1激活后从质膜重新分布至细胞核。更重要的是，活性Rac1被证明可刺激核内β-连环蛋白-淋巴样增强因子1（LEF-1）复合物的形成。这种调控需要Rac1依赖的β-连环蛋白在特定丝氨酸位点的磷酸化，如PLA和免疫沉淀实验所示，当这些位点发生突变（S191A和S605A）时，β-连环蛋白与LEF-1的结合减少了多达50%。我们提出，Rac1介导的β-连环蛋白磷酸化通过增强β-连环蛋白-LEF-1复合物组装来刺激Wnt依赖性基因转录激活，这为Rac1与经典Wnt/β-连环蛋白信号通路之间的相互作用机制提供了新的见解。
