An MAPK-dependent pathway induces epithelial-rnesenchymal Twist activation in human breast cancer cell lines
An MAPK-dependent pathway induces epithelial-rnesenchymal Twist activation in human breast cancer cell lines
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DOI：
10.1016/j.surg.2013.05.012
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发表时间：
2013-08-01
期刊：
影响因子：
3.8
通讯作者：
Mi, Zhiyong
中科院分区：
文献类型：
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作者：
Li, Neill Y.;Weber, Cynthia E.;Mi, Zhiyong
背景Twist是一种上皮-间充质转化（EMT）转录因子，可促进细胞侵袭。我们的研究表明，骨桥蛋白（OPN）调节EMT因子Twist。潜在的信号通路是未知的。我们假设OPN激活Twist诱导人乳腺癌EMT。使用NetPhosK鉴定Twist的潜在激酶。将MEK 1/2、JNK、p38 MAPK和PI 3 K的抑制剂应用于人乳腺癌细胞MDA-MB 231（OPN高）。24小时后，将Twist免疫沉淀并与磷酸丝氨酸孵育。Twist靶蛋白Bmi-1的表达在骨桥蛋白适体（APT）处理24小时后测定;突变体适体（MuAPT）用作对照。在APT和MuAPT处理后12、24和48 h对痉挛伤口测定进行成像。MEK 1/2抑制引起Twist丝氨酸磷酸化的相似的两倍降低（P <0.05）。在MB 231中，APT阻断OPN使Bmi 1蛋白降低两倍（P <0.05）。在划痕试验中，适体处理的细胞在细胞迁移和伤口闭合方面显著降低（P <0.001）。我们证明，OPN细胞外结合MB 231激活自分泌MAPK细胞内信号通路，导致Twist激活和促进Bmi 1表达，以进一步EMT启动和细胞迁移。我们的研究结果阐明了OPN作为人乳腺癌细胞中EMT的主要调节因子的先前未描述的作用。
Background. Twist is an epithelial-mesenchymal transition (EMT) transcription factor that instigates cell invasion. Our research has shown that osteopontin (OPN) regulates the EMT factor Twist. The underlying signaling pathway is unknown. We hypothesized that OPN activates Twist to induce EMT in human breast cancer.Methods. Potential kinases for Twist were identified using NetPhosK. Inhibitors of MEK1/2, JNK, p38 MAPK, and PI3K were applied to human breast cancer cells MDA-MB231 (OPN high). After 24 h, Twist was immunoprecipitated and incubated with phosphoserine. Expression of the Twist target protein, Bmi-1, was determined following 24-h osteopontin aptamer (APT) treatment; mutant aptamer (MuAPT) was used as the control. Scratch-wound assay was imaged 12, 24, and 48 h after APT and MuAPT treatment.Results. MEK1/2 inhibition caused similar to twofold decrease in Twist serine phosphorylation (P < .05). APT blockade of OPN in MB231 decreased Bmi1 protein twofold (P < .05). Aptamer-treated cells were significantly decreased in cell migration and wound closure in the scratch wound-assay (P < .001).Conclusion. We demonstrate that OPN extracellular binding to MB231 activates an autocrine MAPK intracellular signaling pathway resulting in Twist activation and promoting Bmi1 expression to further EMT initiation and cellular migration. Our results elucidate a previously undescribed role for OPN as a prime regulator of EMT in human breast cancer cells.