蛋白激酶CβⅡ通过调控翻译起始介导上皮-间充质转化促进肝细胞癌转移  

Protein kinase CβⅡ mediates epithelial-mesenchymal transition and he⁃patocellular carcinoma metastasis by modulating translation initiation

作　　者：刘敏 赖敏 曾云 祝珊珊 尤梅桂 高畅 LIU Min;LAI Min;ZENG Yun;ZHU Shanshan;YOU Meigui;GAO Chang(Department of Basic Medicine,Xiamen Medical College,Xiamen 361023,China;Department of Pharmacy,Xiamen Medical College,Xiamen 361023,China;Department of Hepatobiliary Surgery,Xiang'an Hospital of Xiamen University,Xiamen 361023,China)

机构地区：[1]厦门医学院基础医学部,福建厦门361023 [2]厦门医学院药学系,福建厦门361023 [3]厦门大学附属翔安医院肝胆外科,福建厦门361023

出　　处：《中国病理生理杂志》2023年第9期1547-1554,共8页Chinese Journal of Pathophysiology

基　　金：厦门市自然科学基金资助项目(No.3502Z20227229);福建省自然科学基金面上项目(No.2022J011403);福建省大学生创新训练计划项目(No.S202212631001)。

摘　　要：目的:研究蛋白激酶C β Ⅱ(protein kinase C β Ⅱ, PKCβⅡ)上调Snail和Twist蛋白表达介导上皮-间充质转化的分子机制。方法:采用[35S]-甲硫氨酸掺入实验和核糖体分离实验观察PKCβⅡ对Snail和Twist mRNA翻译的影响。敲减真核翻译起始因子4E(eukaryotic translation initiation factor 4E, eIF4E)通过核糖体分离实验、Western blot及RT-qPCR观察对PKCβⅡ调控Snail和Twist表达的影响。利用Western blot观察PKCβⅡ对调控eIF4E活性的相关信号通路的影响。利用Western blot观察应用丝裂原活化蛋白激酶相互作用激酶1(mitogen-activated protein kinase-interacting kinase 1, MNK1)抑制剂或雷帕霉素对PKCβⅡ调控Snail和Twist蛋白表达的影响。利用Transwell实验观察敲减eIF4E对PKCβⅡ调控肝癌细胞侵袭的影响。结果:[35S]-甲硫氨酸掺入实验和核糖体分离实验表明,相比β-actin,PKCβⅡ显著上调Snail和Twist mRNA的翻译(P<0.01)。敲减eIF4E抑制PKCβⅡ介导的Snail和Twist mRNA翻译和蛋白表达的上调(P<0.01),但对Snail和Twist的转录没有影响。高表达PKCβⅡ激活细胞外信号调节激酶(extracellular signal-regulated kinase, ERK)/MNK1及蛋白激酶B(protein kinase B, PKB/AKT)/哺乳动物雷帕霉素靶蛋白(mammalian target of rapamycin, mTOR)/eIF4E结合蛋白1(eIF4E-binding protein 1, 4E-BP1)通路上调eIF4E的活性(P<0.01)。应用MNK1抑制剂或雷帕霉素可抑制PKCβⅡ介导的Snail和Twist蛋白表达上调(P<0.01)。敲减eIF4E抑制PKCβⅡ介导的肝癌细胞侵袭能力的增强(P<0.01)。结论:PKCβⅡ通过激活ERK/MNK1通路和AKT/mTOR/4E-BP1通路上调eIF4E的活性,优先促进Snail和Twist mRNA的翻译,介导上皮-间充质转化,从而促进肝细胞癌的转移。这提示PKCβⅡ作为肝细胞癌治疗靶点的潜力。AIM:To investigate the molecular mechanism of protein kinase CβII(PKCβII)mediating epi-thelial-mesenchymal transition by up-regulation of Snail and Twist expression.METHODS:[35S]-methionine incorpora-tion assay and polysome profiling were performed to observe the effect of PKCβII on the translation of Snail and Twist mRNA.The impact of eukaryotic translation initiation factor 4E(eIF4E)knockdown on the expression of Snail and Twist regulated by PKCβII was examined by polysome profiling,Western blot and RT-qPCR.Western blot was used to observe the effect of PKCβII on signaling pathways modulating the eIF4E activity.Western blot was used to observe the effect of a mitogen-activated protein kinase-interacting kinase 1(MNK1)inhibitor or rapamycin on PKCβII-regulated expression of Snail and Twist.Transwell assay was used to observe the impact of eIF4E knockdown on PKCβII-regulated invasion of he-patocellular carcinoma cells.RESULTS:[35S]-methionine incorporation assay and polysome profiling showed that PKCβII significantly promoted the translation of Snail and Twist compared with that ofβ-actin(P<0.01).Knockdown of eIF4E inhibited PKCβII-mediated up-regulation of Snail and Twist mRNA translation and protein expression(P<0.01).However,it caused no statistically significant changes in Snail and Twist mRNA levels.Overexpression of PKCβII acti-vated extracellular signal-regulated kinase(ERK)/MNK1 and protein kinase B(PKB/AKT)/mammalian target of rapamy-cin(mTOR)/eIF4E-binding protein 1(4E-BP1)pathways to up-regulate the eIF4E activity(P<0.01).The application of MNK1 inhibitor or rapamycin blocked PKCβII-induced up-regulation of Snail and Twist protein expression(P<0.01).Knockdown of eIF4E inhibited the increased invasive ability of hepatocellular carcinoma cells induced by PKCβII(P<0.01).CONCLUSION:PKCβII up-regulates the activity of eIF4E through ERK/MNK1 and AKT/mTOR/4E-BP1 path-ways,leading to preferential translation of Snail and Twist mRNA to mediate epithelial-mesenchymal transition and pro-mote hepatoc

关 键 词：肝细胞癌 蛋白激酶CβⅡ 翻译起始 上皮-间充质转化 转移 

分 类 号：R735.7[医药卫生—肿瘤] R363[医药卫生—临床医学]