背景:粘附连接(AJ)促进细胞-细胞接触,并在生理和病理条件下促进细胞通讯以及信号传导。在人类癌症中经常观察到AJ蛋白的异常表达;然而,这些因素对肿瘤发生的影响尚不清楚.此外,对于一些因素,如α-catenin矛盾的数据已经被描述。在这项研究中,我们旨在破译AJ成分α-catenin如何促进肝癌的形成。
方法:TCGA数据用于检测23种人类肿瘤类型的转录变化。为了检测蛋白质,通过免疫组织化学分析肝癌组织微阵列。肝癌细胞系(HLF,Hep3B,HepG2)用于生存力,扩散,RNA干扰介导的基因沉默后的迁移分析。为了研究肿瘤的启动潜力,通过流体动力学基因递送将编码α-catenin和肉豆蔻酰化AKT的载体注射到小鼠中。进行结合质谱的BioID测定以鉴定α-连环蛋白结合配偶体。通过邻近连接和共免疫沉淀测定证实结果。使用染色质免疫沉淀研究了转录调节因子在基因启动子处的结合。
结果:在许多人类恶性肿瘤中,α-cateninmRNA显着降低(例如,结肠腺癌)。相比之下,在其他癌症实体中升高的α-连环蛋白表达与不良临床结果相关(例如,用于肝细胞癌;HCC)。在HCC细胞中,α-连环蛋白在支持肿瘤细胞增殖和迁移的膜和细胞质中都可检测到。在体内,α-连环蛋白与AKT过表达一起促进了中度致癌特性。细胞分裂调节中心体蛋白55(CEP55)被鉴定为HCC细胞胞浆中的新型α-连环蛋白结合蛋白。α-连环蛋白和CEP55之间的物理相互作用与CEP55稳定相关。CEP55在人类HCC组织中高表达,其过度表达与不良的总体生存率和癌症复发相关。除了依赖α-连环蛋白的蛋白质稳定,CEP55由TEA结构域转录因子(TEAD)组成的复合物转录诱导,叉头箱M1(FoxM1),和Yes相关蛋白(YAP)。令人惊讶的是,CEP55不影响HCC细胞增殖,但与α-catenin联合显着支持迁移。
结论:支持迁移的CEP55在HCC细胞中由两种独立的机制诱导:通过与AJ蛋白α-catenin相互作用的稳定和通过FoxM1/TEAD/YAP复合物的转录激活。
上皮细胞中的细胞-细胞接触对于细胞极性很重要,细胞分隔,以及发育过程中的组织结构,稳态,和后生动物成年组织的再生。在这种情况下,粘附连接(AJ)机械地感测细胞接触信息,直接影响细胞骨架重塑,信号通路的调节,最终是细胞生物学。的确,细胞间接触的丧失和细胞极性是人类癌变的关键特征,也是鉴定许多上皮肿瘤的重要病理参数。我们在这项研究中证明,在人肝细胞癌(HCC)中经常观察到AJ成分α-连环蛋白的过表达。α-连环蛋白支持肝癌细胞增殖和迁移。连同癌基因AKT,α-连环蛋白适度促进小鼠肝脏肿瘤的发生。使用质谱,我们在肝癌细胞的胞质溶胶中发现了几种新的α-catenin相互作用伴侣,包括胞质分裂调节蛋白中心体蛋白55(CEP55)。CEP55介导促迁移效应,其在HCC细胞中的过表达受两种分子机制控制:α连环蛋白依赖性蛋白稳定和TEA域转录因子(TEAD)/叉头盒M1(FoxM1)/Yes相关蛋白(YAP)复合物的转录诱导。总之,我们在这里描述了一种新的机制,即细胞间接触的变化如何支持肝癌的形成和进展。这项研究表明,AJ成分α-连环蛋白的失调通过不同的分子机制促进肝癌的发生。视频摘要。
Adherens junctions (AJs) facilitate cell-cell contact and contribute to cellular communication as well as signaling under physiological and pathological conditions. Aberrant expression of AJ proteins is frequently observed in human cancers; however, how these factors contribute to tumorigenesis is poorly understood. In addition, for some factors such as α-catenin contradicting data has been described. In this study we aim to decipher how the AJ constituent α-catenin contributes to liver cancer formation.
TCGA data was used to detect transcript changes in 23 human tumor types. For the detection of proteins, liver cancer tissue microarrays were analyzed by immunohistochemistry. Liver cancer cell lines (HLF, Hep3B, HepG2) were used for viability, proliferation, and migration analyses after RNAinterference-mediated gene silencing. To investigate the tumor initiating potential, vectors coding for α-catenin and myristoylated AKT were injected in mice by hydrodynamic gene delivery. A BioID assay combined with mass spectrometry was performed to identify α-catenin binding partners. Results were confirmed by proximity ligation and co-immunoprecipitation assays. Binding of transcriptional regulators at gene promoters was investigated using chromatin-immunoprecipitation.
α-catenin mRNA was significantly reduced in many human malignancies (e.g., colon adenocarcinoma). In contrast, elevated α-catenin expression in other cancer entities was associated with poor clinical outcome (e.g., for hepatocellular carcinoma; HCC). In HCC cells, α-catenin was detectable at the membrane as well as cytoplasm where it supported tumor cell proliferation and migration. In vivo, α-catenin facilitated moderate oncogenic properties in conjunction with AKT overexpression. Cytokinesis regulator centrosomal protein 55 (CEP55) was identified as a novel α-catenin-binding protein in the cytoplasm of HCC cells. The physical interaction between α-catenin and CEP55 was associated with CEP55 stabilization. CEP55 was highly expressed in human HCC tissues and its overexpression correlated with poor overall survival and cancer recurrence. Next to the α-catenin-dependent protein stabilization, CEP55 was transcriptionally induced by a complex consisting of TEA domain transcription factors (TEADs), forkhead box M1 (FoxM1), and yes-associated protein (YAP). Surprisingly, CEP55 did not affect HCC cell proliferation but significantly supported migration in conjunction with α-catenin.
Migration-supporting CEP55 is induced by two independent mechanisms in HCC cells: stabilization through interaction with the AJ protein α-catenin and transcriptional activation via the FoxM1/TEAD/YAP complex.
Cell–cell contact in epithelial cells is important for cell polarity, cellular compartmentalisation, as well as tissue architecture during development, homeostasis, and regeneration of adult tissues in metazoans. In this context, adherens junctions (AJs) mechanically sense cell contact information with direct impact on cytoskeletal remodelling, the regulation of signalling pathways, and eventually cell biology. Indeed, the loss of cell–cell contact and cellular polarity are key features in human carcinogenesis and important pathological parameters for the identification of many epithelial tumors.We demonstrate in this study, that overexpression of the AJ constituent α‐catenin is frequently observed in human hepatocellular carcinoma (HCC). α‐catenin supports HCC cell proliferation and migration. Together with the oncogene AKT, α‐catenin moderately facilitates tumor initiation in mouse livers. Using mass spectrometry, we identified several new α‐catenin interaction partners in the cytosol of liver cancer cells, including the cytokinesis regulator centrosomal protein 55 (CEP55). CEP55 mediates pro-migratory effects and its overexpression in HCC cells is controlled by two molecular mechanisms: α‐catenin-dependent protein stabilization and transcriptional induction by the TEA domain transcription factors (TEADs)/forkhead box M1 (FoxM1)/yes-associated protein (YAP) complex.In summary, we here describe a new mechanism how changes in cell–cell contact support liver cancer formation and progression. This study demonstrates that dysregulation of the AJ component α‐catenin contributes to liver carcinogenesis via distinct molecular mechanisms. Video Abstract.