Olig1 is a Smad cofactor involved in cell motility induced by transforming growth factor-b. Mus musculus

Transforming growth factor (TGF)-β plays crucial roles in embryonic development and adult tissue homeostasis by eliciting various cellular responses in target cells. TGF-β signaling is principally mediated through receptor-activated Smad proteins, which regulate expression of target genes in cooperation with other DNA-binding transcriptionfactors (Smad cofactors). In this study, we found that the basic helix-loop-helix transcription factor Olig1 is a Smad cofactor involved in TGF-b-induced cell motility. Knockdown of Olig1 attenuated TGF-β-induced cell motility in chamber migration and wound healing assays. In contrast, Olig1 knockdown had no effect on bone morphogenetic protein-induced cell motility, TGF-β-induced cytostasis or epithelial-mesenchymal transition. Furthermore, we observed that cooperation of Smad2/3 with Olig1 is regulated by a peptidyl-prolyl cis/trans isomerase, Pin1. TGF-b-induced cell motility, induction of Olig1-regulated genes, and physical interaction between Smad2/3 and Olig1 were all inhibited after knockdown of Pin1, indicating a novel mode of regulation of Smad signaling. We also found that Olig1 interacts with the L3 loop of Smad3. Using a synthetic peptide corresponding to the L3 loop of Smad3, we succeeded in selectively inhibiting TGF-b-induced cell motility. These findings may lead to a new strategy for selective regulation of TGF-b-induced cellular responses. Overall design: NMuMG cells were transfected with siRNAs (siControl, siOlig1 or siPin1) and treated with or without TGF-b for 1h. We compared genes affected by knockdown of Olig1 and that of Pin1.

转化生长因子β（Transforming growth factor-β, TGF-β）可在靶细胞中引发多种细胞应答，在胚胎发育与成年组织稳态维持中发挥关键作用。TGF-β信号通路主要通过受体激活型Smad蛋白（receptor-activated Smad proteins）介导，这类蛋白可与其他DNA结合转录因子（DNA-binding transcription factors）协同调控靶基因的表达。本研究发现，碱性螺旋-环-螺旋转录因子Olig1（basic helix-loop-helix transcription factor Olig1）是参与TGF-β诱导的细胞运动的Smad辅因子（Smad cofactor）。在小室迁移与划痕愈合实验中，敲低Olig1可削弱TGF-β诱导的细胞运动能力。与之相反，敲低Olig1对骨形态发生蛋白（bone morphogenetic protein, BMP）诱导的细胞运动、TGF-β诱导的细胞静止或上皮间质转化（epithelial-mesenchymal transition, EMT）均无影响。进一步研究发现，Smad2/3（Smad2/3）与Olig1的协同作用受到肽酰脯氨酰顺反异构酶（peptidyl-prolyl cis/trans isomerase）Pin1的调控。敲低Pin1后，TGF-β诱导的细胞运动、Olig1调控基因的表达，以及Smad2/3与Olig1之间的物理相互作用均受到抑制，这揭示了一种全新的Smad信号通路调控模式。此外，我们发现Olig1可与Smad3的L3环（L3 loop）相互作用。利用对应Smad3 L3环的合成肽（synthetic peptide），我们成功实现了对TGF-β诱导的细胞运动的选择性抑制。上述研究结果有望为选择性调控TGF-β诱导的细胞应答提供新策略。
实验设计概述：将NMuMG细胞（NMuMG cells）用小干扰RNA（small interfering RNA, siRNA）（siControl、siOlig1或siPin1）进行转染，随后用或不用TGF-β处理1小时。我们对敲低Olig1与敲低Pin1所影响的基因进行了比较分析。