Cellular Biology Laboratory, Nutrition and Food Technology Institute, INTA, Universidad de Chile, Chile.
Int J Cancer. 2010 Jul 1;127(1):77-85. doi: 10.1002/ijc.25045.
TGF-beta1 is a potent inductor of malignance in cancer cells. TGF-beta1 stimulates the expression of extracellular matrix degrading proteases, cell migration and it is also involved in the epithelial-mesenchymal transition (EMT). In the present work, we analyzed the role of Spred2 in the urokinase-type plasminogen activator (uPA) stimulation, EMT and cell migration by TGF-beta1. We found that both the expression of mRNA and the protein level of Spred2 were lower in transformed keratinocytes PDV compared with immortalized keratinocytes MCA-3D. The transient ectopic expression of Spred2 in PDV cells inhibited the TGF-beta1-transactivated SRE-Luc reporter which is related with the ERK1,2 signal. The stable ectopic expression of Spred2 in PDV cells (SP cells) led to the loss of ERK 1,2 activation by TGF-beta1, although Smad2 activation was not affected, and the knockdown of Spred2 enhanced the activation of ERK1,2 signal by TGF-beta1. The increment of uPA expression induced by TGF-beta1 was suppressed in SP cells. In contrast, the stimulus on PAI-1 expression was not affected and comparable to parental PDV cells. SP cells under TGF-beta1 treatment were unable to display the EMT, since the overexpression of Spred2 abolished the TGF-beta1-induced disruption of the E-cadherin cell to cell interactions, reorganization of the actin cytoskeleton and upregulation of the mesenchymal marker vimentin. Finally, SP cells could not respond to the TGF-beta1 stimulus on cell migration. Taken together, the data in the present study suggests that Spred2 is a regulator of TGF-beta1-induced malignance in transformed keratinocytes.
TGF-β1 是癌细胞恶性转化的有效诱导剂。TGF-β1 可刺激细胞外基质降解蛋白酶的表达、细胞迁移，并且参与上皮-间充质转化（EMT）。在本研究中，我们分析了 Spred2 在尿激酶型纤溶酶原激活物（uPA）刺激、EMT 和 TGF-β1 诱导的细胞迁移中的作用。我们发现，与永生化角质细胞 MCA-3D 相比，转化角质细胞 PDV 中 Spred2 的 mRNA 表达和蛋白水平均降低。瞬时过表达 Spred2 可抑制与 ERK1、2 信号相关的 TGF-β1 转激活 SRE-Luc 报告基因。Spred2 在 PDV 细胞中的稳定过表达（SP 细胞）导致 TGF-β1 引起的 ERK1、2 激活丧失，尽管 Smad2 激活不受影响，而 Spred2 的敲低增强了 TGF-β1 对 ERK1、2 信号的激活。TGF-β1 诱导的 uPA 表达增加在 SP 细胞中受到抑制。相比之下，PAI-1 表达的刺激不受影响，与亲本 PDV 细胞相当。在 TGF-β1 处理下，SP 细胞无法显示 EMT，因为 Spred2 的过表达消除了 TGF-β1 诱导的 E-钙粘蛋白细胞间相互作用的破坏、肌动蛋白细胞骨架的重排和间充质标志物波形蛋白的上调。最后，SP 细胞无法对 TGF-β1 刺激的细胞迁移作出反应。总之，本研究的数据表明，Spred2 是转化角质细胞中 TGF-β1 诱导恶性转化的调节剂。
