BAG3 蛋白Ser187位点磷酸化诱导人甲状腺癌FRO 细胞上皮间质转化

Bcl-2相关抗凋亡蛋白3（Bcl-2 associated athanogene 3，BAG3）是BAG家族的重要成员，调节肿瘤细胞的粘附、迁移和侵袭，促进恶性肿瘤的复发和转移.本室前期工作证明，PKCδ可催化BAG3的Ser187位点磷酸化.本文研究BAG3蛋白磷酸化修饰对甲状腺癌FRO 细胞EMT表型转化的影响.稳定转染野生型WT-BAG3、模拟磷酸化型S187D-BAG3、阻碍磷酸化型S187A-BAG3 FRO细胞后，观察细胞形态的变化.结果显示，稳定转染模拟磷酸化型S187D-BAG3引起甲状腺癌FRO细胞呈现明显的间质细胞形态.实时PCR 和Western印迹，结果显示，稳定表达S187D-BAG3显著上调间质细胞标记物N-cadherin和波形蛋白mRNA与蛋白质在FRO细胞的表达，但下调上皮细胞标记物E-cadherin的mRNA和蛋白质的表达.同时，免疫荧光结果显示，稳定过表达S187D-BAG3的FRO细胞，E-cadherin和β-catenin出现向核周的内化.本文结果提示，BAG3蛋白磷酸化修饰可诱导甲状腺癌FRO细胞上皮间质转化.

Phosphorylation of BAG3 at Ser187 Induces Epithelial-to-Mesenchymal Transition in Human Thyroid Cancer FRO Cells

Bcl-2 associated athanogene 3 (BAG3) is one of the important members of the BAG family. BAG3 adjusts the adhesion, migration and invasion of tumor cells, promotes the recurrence and metastasis of malignant tumor. Our preliminary work has proved that PKC delta phosphorylates BAG3 at Ser187 sites. In this paper, we studied the effect of BAG3 phosphorylation on epithelial-to-mesenchymal transition(EMT)phenotype transformation in thyroid cancer FRO cells. FRO cells were stably transfeced WT-BAG3, S187D-BAG3, S187A-BAG3, then we observed the changes of cell morphology. Results showed that S187D-BAG3 cells exhibited obvious mesenchymal cell morphology. RT-PCR and Western blot results demonstrated higher mRNA and protein expression level of mesenchymal markers,N-cadherin and vimentin and lower epithelial marker, E-cadherin mRNA and protein expression levels in S187D-BAG3 cells. Internalization of E-cadherin and β-catenin were observed in S187D-BAG3 cells by IF results. Our results indicated that the phosphorylation of BAG3 at Ser187 induced EMT of FRO cells.