YAP1-FOS促进人口腔鳞状上皮癌细胞增殖，迁移及EMT转换的机制研究

摘要/Abstract

摘要： 目的　探讨YAP1-FOS在促进口腔鳞状上皮细胞癌（OSCC）细胞的增殖，迁移及EMT转换过程中发挥的功能。方法　通过组织免疫荧光实验检测OSCC癌组织及周围正常组织中YAP1的表达及亚细胞定位，细胞克隆形成实验检测YAP1对OSCC细胞增殖情况的影响，细胞划痕实验检测YAP1对OSCC细胞迁移的影响，Western Blot检测YAP1对EMT相关分子标记基因蛋白表达的影响，RT-PCR检测YAP1对EMT相关分子标记基因转录水平变化的影响，CoIP及免疫印迹实验检测YAP1与FOS互作，共同调控下游基因表达以控制EMT转换过程。结果　组织免疫荧光实验结果显示，OSCC癌组织细胞中YAP1表达增加，且由细胞质转移至细胞核；细胞克隆形成实验结果显示，YAP1上调后克隆形成数目显著增加（P<0.01）；细胞划痕实验结果显示，过表达YAP1后，细胞划痕间距显著减小（P<0.01）；Western Blot及RT-PCR结果显示，上皮细胞关键基因E-cadherin表达水平在YAP1上调后被显著抑制（P<0.01），而间质细胞关键基因β- catenin，Vimentin及N-cadherin表达水平则显著上调（P<0.01）。CoIP及免疫印迹结果显示，YAP1与核转录因子FOS互作，抑制FOS表达后YAP1的促细胞增殖能力显著减弱（P<0.01），同时EMT相关分子标记表达显著下调（P<0.01）。结论　YAP1结合FOS基因促进人口腔鳞状上皮细胞癌（OSCC）细胞的增殖，迁移及EMT转换过程，这一结果可为人口腔鳞状上皮细胞癌的发生发展提供理论基础，YAP1-FOS或可成为新的人口腔癌药物靶点。

关键词: YAP1-FOS, 细胞增殖, EMT, 口腔鳞状上皮细胞癌

Abstract: Objective　To explore the roles of YAP1-FOS in promoting the process of cell proliferation, migration and EMT in oral squamous cell carcinoma (OSCC). Methods　The YAP1 expression and subcellular localization in the OSCC cancer tissue and the surrounding normal tissue were detected by tissue immunofluorescence assay. The effect of YAP1 on cell proliferation in OSCC cells was tested by cell clonogenic assay. The effect of YAP1 on cell migration in OSCC cells was detected by cell scratch assay. The effects of YAP1 on the marker gene protein expression and changes in marker gene transcription level of EMT related molecule were respectively detected by Western blot and RT-PCR. The process in which YAP1 and FOS interacted with each other to control the conversion process of EMT through mutual regulation of downstream genes was detected by CoIP and immunoblot. Results　The immunofluorescence assay results showed that the YAP1 expression increased in OSCC cancer tissue and transferred from cytoplasm to cell nucleus. Clonogenic assay results showed that the clone number increased significantly after over-expression of YAP1 (P < 0.01). The scratch assay results showed that the wound distance significantly decreased after over-expression of YAP1(P < 0.01). Western blot and RT-PCR showed that the expression levels of the key gene of epithelial cells E-cadherin were downregulated (P < 0.01), while the expression levels of the key genes of mesenchymal cells, β-catenin, Vimentin and N-cadherin, were upregulated significantly after YAP1 over-. CoIP and immunoblot results showed that YAP1 interacted with FOS, then the ability of YAP1 promoting cell proliferation significantly weakened afte the depression of FOS expression, meanwhile the marker expression of EMT related molecule was significant down-regulated(P < 0.01). Conclusion　YAP1 combined with FOS genes promotes the cell proliferation, migration and EMT transcription in human OSCC, which provides a theoretical basis for the occurrence and development of human OSCC. YAP1-FOS may become the new drug target of human oral cancers.

Key words: Keywords: YAP1-FOS, cell proliferation, EMT, oral squamous cell carcinoma

中图分类号:

王毅刘红胡杨. YAP1-FOS促进人口腔鳞状上皮癌细胞增殖，迁移及EMT转换的机制研究[J]. , 2019, 39(7): 581-586.

参考文献 20

[1]	王俊杰, 赵腾达, 刘兴光, 等.基因在口腔鳞状细胞癌中的表达及意义[J].口腔医学, 2016, 36(4):333-336
[2]	李金, 华红.无创检测技术用于口腔鳞状细胞癌早期诊断国内研究现状[J].现代口腔医学杂志, 2016, 30(6):358-362
[3]	陈浩, 马洪, 段晓峰, 等.基因在口腔鳞状细胞癌中的表达及意义[J].实用口腔医学杂志, 2016, 32(3):436-438
[4]	陈新, 徐文华, 周健, 等.口腔鳞状细胞癌现状[J].口腔医学, 2017, 37(5):462-465
[5]	张斌.口腔鳞状细胞癌的表观遗传学研究现状和进展[J].口腔颌面外科杂志, 2016, 26(2):77-82
[6]	Camargo FD, Gokhale S, Johnnidis JB, et al.YAP1 increases organ size and expands undifferentiated progenitor cells[J].CurrBiol, 2007, 17(23):2054-2060
[7]	Lu L, Li Y, Kim SM, et al.Hippo signaling is a potent in vivo growth and tumor suppressor pathway in the mammalian liver[J].ProcNatlAcadSci USA, 2010, 107(4):1437-1442
[8]	Yu FX, Guan KL.The Hippo pathway: regulators and regulations[J].Genes Dev, 2013, 27(4):355-371
[9]	Zhao B, Wei X, Li W, et al.Inactivation of YAP oncoprotein by the Hippo pathway is involved in cell contact inhibition and tissue growth control[J].Genes Dev, 2007, 21(21):2747-2761
[10]	Tulchinsky E.Fos family members: regulation,structure and role in oncogenic transformation[J].HistolHistopathol, 2000, 15(3):921-928
[11]	Fialka I, Schwarz H, Reichmann E, et al.The estrogen-dependent c-JunER protein causes a reversible loss of mammary epithelial cell polarity involving a destabilization of adherensjunctions[J].J Cell Biol, 1996, 132(6):1115-1132
[12]	朱皓, 任光玉, 吴琴, 等.子宫颈癌中高危型HPV感染与c-jun和c-fos蛋白表达的关系[J].临床与实验病理学杂志. 2017，33(1):27-31.
[13]	李建成.EGFR、Her-2及c-Fos联合检测在食管鳞癌中的意义[A]. .中国中药杂志2015/专集：基层医疗机构从业人员科技论文写作培训会议论文集[C]. 2016, 3.
[14]	Shao DD, Xue W, Krall EB, et al.KRAS and YAP1 converge to regulate EMT and tumor survival[J].Cell, 2014, 158(1):171-184
[15]	Gupta B, Johnson NW, Kumar N.Global Epidemiology of Head and Neck Cancers: A Continuing Challenge[J].Oncology, 2016, 91(1):13-23
[16]	Zanconato F, Cordenonsi M, Piccolo S.YAPTAZ at the Roots of Cancer[J].Cancer Cell, 2016, 29(6):783-803
[17]	Kodaka M, Hata Y.The mammalian Hippo pathway: regulation and function of YAP1 and TAZ[J].Cell Mol Life Sci, 2015, 72(2):285-306
[18]	Huang J, Wu S, Barrera J, et al.The Hippo signaling pathway coordinately regulates cell proliferation and apoptosis by inactivating Yorkie,the Drosophila Homolog of YAP[J].Cell, 2005, 122(3):421-434
[19]	Zhang N, Bai H, David KK, et al.The MerlinNF2 tumor suppressor functions through the YAP oncoprotein to regulate tissue homeostasis in mammals[J].Dev Cell, 2010, 19(1):27-38
[20]	Nishio M, Hamada K, Kawahara K, et al.Cancer susceptibility and embryonic lethality in Mob1a1b double-mutant mice[J].J Clin Invest, 2012, 122(12):4505-4518