Department of Histology & Embryology, School of Veterinary Medicine, Bursa Uludag University, 16059, Bursa, Turkey.
Department of Biochemistry, School of Veterinary Medicine, Bursa Uludag University, 16059, Bursa, Turkey.
Mol Biol Rep. 2022 Nov;49(11):10195-10204. doi: 10.1007/s11033-022-07893-1. Epub 2022 Sep 2.
Transforming Growth Factor β (TGFβ) proteins are potent inducers of the epithelial-mesenchymal transition (EMT) in tumor cells. Although mitogen-activated protein kinase (MAPK) family has been shown to be involved in TGFβ-induced EMT, role of Dual Specificity Phosphatases (DUSP), key regulators of MAPK activity, in TGFβ-induced EMT is largely unkonwn.
Real-time qPCR analyses were performed to determine the effect of TGFβ1 on expression of EMT genes and DUSP proteins in the non-small cell lung cancer model A549 and pancreatic adenocarcinoma model PANC1 cells. Western blot analyses were conducted to study the changes in protein levels of EMT proteins and select DUSP proteins, as well as phosphorylations of MAPK proteins upon TGFβ1 stimulation. Small interfering RNA (siRNA) was utilized to reduce expressions of DUSP genes. We observed that the EMT phenotype coincided with increases in phosphorylations of the MAPK proteins ERK1/2, p38MAPK, and JNK upon TGFβ1 stimulation. Real-time qPCR analysis showed increases in DUSP15 and DUSP26 mRNA levels and Western blot analysis confirmed the increase in DUSP26 protein levels in both A549 and PANC1 cells treated with TGFβ1 relative to control. Silencing of DUSP26 expression by siRNA markedly suppressed the effect of TGFβ1 on E-cadherin and mesenchymal genes in the cells.
Data provided suggest that TGFβ1 modulates the expression of DUSP genes and that upregulation of DUSP26 may be required for TGFβ1-promoted EMT in A549 and PANC1 cells. Further studies should be carried out to elucidate the requirement of individual DUSPs in TGFβ1-associated EMT in tumor cells.
转化生长因子 β（TGFβ）蛋白是肿瘤细胞上皮-间充质转化（EMT）的有效诱导剂。虽然丝裂原活化蛋白激酶（MAPK）家族已被证明参与 TGFβ 诱导的 EMT，但 Dual Specificity Phosphatases（DUSP），即 MAPK 活性的关键调节剂，在 TGFβ 诱导的 EMT 中的作用在很大程度上仍不清楚。
实时 qPCR 分析用于确定 TGFβ1 对非小细胞肺癌模型 A549 和胰腺腺癌细胞模型 PANC1 中 EMT 基因和 DUSP 蛋白表达的影响。Western blot 分析用于研究 EMT 蛋白和选定的 DUSP 蛋白的蛋白水平变化，以及 TGFβ1 刺激后 MAPK 蛋白的磷酸化变化。利用小干扰 RNA（siRNA）降低 DUSP 基因的表达。我们观察到，EMT 表型与 TGFβ1 刺激后 MAPK 蛋白 ERK1/2、p38MAPK 和 JNK 的磷酸化增加相一致。实时 qPCR 分析显示，与对照组相比，TGFβ1 处理的 A549 和 PANC1 细胞中 DUSP15 和 DUSP26 mRNA 水平增加，Western blot 分析证实 DUSP26 蛋白水平增加。siRNA 沉默 DUSP26 表达显著抑制了 TGFβ1 对细胞中 E-钙粘蛋白和间充质基因的作用。
提供的数据表明，TGFβ1 调节 DUSP 基因的表达，上调 DUSP26 可能是 TGFβ1 促进 A549 和 PANC1 细胞 EMT 所必需的。应进一步研究以阐明个体 DUSPs 在肿瘤细胞中 TGFβ1 相关 EMT 中的需求。
