Overexpression and nuclear enrichment of the oncogene yes-associated protein (YAP) cause tumor initiation and support tumor progression in human hepatocellular carcinoma (HCC) via cell autonomous mechanisms. However, how YAP expression in tumor cells affects intercellular communication within the tumor microenvironment is not well understood. To investigate how tumor cell-derived YAP is changing the paracrine communication network between tumor cells and non-neoplastic cells in hepatocarcinogenesis, the expression and secretion of cytokines, growth factors and chemokines were analyzed in transgenic mice with liver-specific and inducible expression of constitutively active YAP (YAPS127A). Transcriptomic and proteomic analyses were performed using primary isolated hepatocytes and blood plasma. In vitro, RNAinterference (RNAi), expression profiling, functional analyses and chromatin immunoprecipitation (ChIP) analyses of YAP and the transcription factor TEA domain transcription factor 4 (TEAD4) were performed using immortalized cell lines. Findings were confirmed in cohorts of HCC patients at the transcript and protein levels. YAP overexpression induced the expression and secretion of many paracrine-acting factors with potential impact on tumorous or non-neoplastic cells (e.g. plasminogen activator inhibitor-1 (PAI-1), C-X-C motif chemokine ligand 13 (CXCL13), CXCL16). Expression analyses of human HCC patients showed an overexpression of PAI-1 in human HCC tissues and a correlation with poor overall survival as well as early cancer recurrence. PAI-1 statistically correlated with genes typically induced by YAP, such as connective tissue growth factor (CTGF) and cysteine rich angiogenic inducer 61 (CYR61) or YAP-dependent gene signatures (CIN4/25). In vitro, YAP inhibition diminished the expression and secretion of PAI-1 in murine and human liver cancer cell lines. PAI-1 affected the expression of genes involved in cellular senescence and oncogene-induced senescence was confirmed in YAPS127A transgenic mice. Silencing of TEAD4 as well as treatment with the YAP/TEAD interfering substance Verteporfin reduced PAI-1 expression. ChIP analyses confirmed the binding of YAP and TEAD4 to the gene promoter of PAI-1 (SERPINE1). These results demonstrate that the oncogene YAP changes the secretome response of hepatocytes and hepatocyte-derived tumor cells. In this context, the secreted protein PAI-1 is transcriptionally regulated by YAP in hepatocarcinogenesis. Perturbation of these YAP-dependent communication hubs including PAI-1 may represent a promising pharmacological approach in tumors with YAP overexpression.

中文翻译：

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Yes 相关蛋白 (YAP) 诱导分泌组表型并转录调节纤溶酶原激活剂 Inhibitor-1 (PAI-1) 在肝癌发生中的表达

癌基因 yes 相关蛋白 (YAP) 的过度表达和核富集通过细胞自主机制导致肿瘤发生并支持人类肝细胞癌 (HCC) 的肿瘤进展。然而，肿瘤细胞中的 YAP 表达如何影响肿瘤微环境内的细胞间通讯尚不清楚。为了研究肿瘤细胞衍生的 YAP 如何改变肝癌发生过程中肿瘤细胞和非肿瘤细胞之间的旁分泌通讯网络，在肝脏特异性和可诱导表达的转基因小鼠中分析了细胞因子、生长因子和趋化因子的表达和分泌。主动 YAP (YAPS127A)。使用原代分离的肝细胞和血浆进行转录组学和蛋白质组学分析。体外、RNA 干扰 (RNAi)、表达谱、使用永生化细胞系对 YAP 和转录因子 TEA 结构域转录因子 4 (TEAD4) 进行功能分析和染色质免疫沉淀 (ChIP) 分析。结果在转录和蛋白质水平的 HCC 患者队列中得到证实。YAP 过表达诱导了许多旁分泌作用因子的表达和分泌，对肿瘤或非肿瘤细胞有潜在影响（例如纤溶酶原激活剂抑制剂-1 (PAI-1)、CXC 基序趋化因子配体 13 (CXCL13)、CXCL16）。人类 HCC 患者的表达分析表明，PAI-1 在人类 HCC 组织中过度表达，并且与较差的总体存活率和早期癌症复发相关。PAI-1 与通常由 YAP 诱导的基因在统计学上相关，例如结缔组织生长因子 (CTGF) 和富含半胱氨酸的血管生成诱导剂 61 (CYR61) 或 YAP 依赖性基因特征 (CIN4/25)。在体外，YAP 抑制减少了 PAI-1 在鼠和人肝癌细胞系中的表达和分泌。PAI-1 影响参与细胞衰老的基因的表达，在 YAPS127A 转基因小鼠中证实了癌基因诱导的衰老。TEAD4 的沉默以及用 YAP/TEAD 干扰物质 Verteporfin 处理降低了 PAI-1 的表达。ChIP 分析证实了 YAP 和 TEAD4 与 PAI-1 (SERPINE1) 的基因启动子的结合。这些结果表明，致癌基因 YAP 改变了肝细胞和肝细胞来源的肿瘤细胞的分泌组反应。在这种情况下，分泌蛋白 PAI-1 在肝癌发生过程中受到 YAP 的转录调控。