WAVE2 Regulates Epithelial Morphology and Cadherin Isoform Switching through Regulation of Twist and Abl

BackgroundEpithelial morphogenesis is a dynamic process that involves coordination of signaling and actin cytoskeletal rearrangements.Principal FindingsWe analyzed the contribution of the branched actin regulator WAVE2 in the development of 3-dimensional (3D) epithelial structures. WAVE2-knockdown (WAVE2-KD) cells formed large multi-lobular acini that continued to proliferate at an abnormally late stage compared to control acini. Immunostaining of the cell-cell junctions of WAVE2-KD acini revealed weak and heterogeneous E-cadherin staining despite little change in actin filament localization to the same junctions. Analysis of cadherin expression demonstrated a decrease in E-cadherin and an increase in N-cadherin protein and mRNA abundance in total cell lysates. In addition, WAVE2-KD cells exhibited an increase in the mRNA levels of the epithelial-mesenchymal transition (EMT)-associated transcription factor Twist1. KD of Twist1 expression in WAVE2-KD cells reversed the cadherin switching and completely rescued the aberrant 3D morphological phenotype. Activity of the WAVE2 complex binding partner Abl kinase was also increased in WAVE2-KD cells, as assessed by tyrosine phosphorylation of the Abl substrate CrkL. Inhibition of Abl with STI571 rescued the multi-lobular WAVE2-KD 3D phenotype whereas overexpression of Abl kinase phenocopied the WAVE2-KD phenotype.ConclusionsThe WAVE2 complex regulates breast epithelial morphology by a complex mechanism involving repression of Twist1 expression and Abl kinase activity. These data reveal a critical role for WAVE2 complex in regulation of cellular signaling and epithelial morphogenesis.

背景
上皮形态发生是一个动态过程，涉及信号转导与肌动蛋白细胞骨架重排的协同调控。
主要研究结果
我们分析了分支状肌动蛋白调控因子WAVE2在三维（3D）上皮结构发育中的作用。与对照腺泡相比，WAVE2敲低（WAVE2-KD）细胞形成了大型多叶状腺泡，且在异常晚的阶段仍持续增殖。对WAVE2-KD腺泡的细胞间连接进行免疫染色发现，尽管肌动蛋白丝在相同连接处的定位几乎未发生变化，但E-钙粘蛋白的染色信号较弱且呈异质性。对钙粘蛋白表达的分析显示，在总细胞裂解物中，E-钙粘蛋白的丰度降低，而N-钙粘蛋白的蛋白与mRNA水平均升高。此外，WAVE2-KD细胞中上皮间质转化（epithelial-mesenchymal transition, EMT）相关转录因子Twist1的mRNA水平显著升高。在WAVE2-KD细胞中敲低Twist1的表达，可逆转钙粘蛋白的表达转换，并完全挽救异常的3D形态表型。通过检测Abl底物CrkL的酪氨酸磷酸化水平，我们发现WAVE2复合物的结合伴侣Abl激酶的活性在WAVE2-KD细胞中也有所升高。使用STI571抑制Abl激酶可挽救多叶状的WAVE2-KD 3D表型，而过表达Abl激酶则可重现WAVE2-KD的表型。
结论
WAVE2复合物通过涉及抑制Twist1表达与调控Abl激酶活性的复杂机制调控乳腺上皮形态。上述研究结果揭示了WAVE2复合物在细胞信号转导与上皮形态发生调控中的关键作用。