TCL1/WTX/β-Catenin信号通路调控上皮间质转化促进结直肠癌侵袭转移的分子机制研究

Distant metastasis is one of the main causes of death in colorectal cancer (CRC). Although CRC metastasis recently attracts extensive attention, its molecular mechanisms still remain to be determined. Our preliminary data indicates that: (1) The relative expression of TCL1 is significantly elevated in CRC cell lines and tissues, which confers enhanced invasiveness and metastasis; (2) TCL1 knockdown in CRC cells inhibits epithelial-mesenchymal transition (EMT). (3)Mass spectrometry analysis suggests that TCL1 interacts with WTX and recruits AKT to phosphorylate WTX, which disrupts the association of WTX with β-catenin; (4) Overexpression of TCL1 not only upregulates the level of β-Catenin and but also promotes its nuclear localization, whereas β-Catenin/TCF4/LEF1 complex transcriptionally transactivates the promoter of TCL1, suggesting a positive feedback loop. Therefore, we hypothesized that TCL1/WTX/β-Catenin signal pathway plays a critical role in CRC invasiveness and metastasis by regulating EMT. By using multiple molecular biology tools, cell culture, and mice model, we comprehensively investigated the role of TCL1/WTX/β-Catenin feedback loop in CRC metastasis. Together, this study will provide not only a novel theoretical basis on the mechanism of molecular biology of CRC metastasis, but also new drug targets for the precise treatment of CRC.

远处转移是结直肠癌患者的主要死亡原因，虽近年来得到广泛关注，但其具体的分子机制仍不明确。我们前期研究发现：（1）TCL1在结直肠癌中高表达，且与转移相关；（2）在结直肠癌细胞中敲低TCL1可抑制肿瘤的上皮间质转化（EMT）；（3）TCL1可与WTX结合，募集AKT磷酸化WTX，从而使WTX脱离β-Catenin；（4）TCL1过表达促使β-Catenin表达增加并入核，同时TCL1启动子区域存在TCF4/LEF1复合物结合位点。因此我们推测：TCL1/WTX/β-Catenin信号通路及自身环路通过调控EMT促进结直肠癌的侵袭转移。本课题将应用多种生物学研究方法，在细胞水平、分子机制水平、动物水平和临床水平阐述TCL1/WTX/β-Catenin信号通路及自身正反馈环路通过影响EMT相关基因表达调控结直肠癌侵袭转移这一机制。本研究有望为结直肠癌远处转移的治疗提供新的治疗靶点。

背景:T细胞白血病/淋巴瘤1 (T-cell leukemia/lymphoma 1, Tcl1)是恶性血液病中已确定的致癌基因。我们之前发现Tcl1与肝细胞癌的戊糖磷酸途径有关。然而，Tcl1在结直肠癌(CRC)发展中的作用仍不清楚。.方法:比较Tcl1在人结直肠癌和正常结肠组织中的表达。我们对小鼠的CRC细胞和肿瘤的形成进行了增殖实验、菌落形成实验、球形成实验和凋亡实验。我们探讨了Tcl1 / mtx /β-catenin轴使用质谱分析,co-immunoprecipitation,免疫荧光染色,荧光素酶记者分析,芯片分析。.结果:我们发现Tcl1在CRC标本中上调，与CRC患者生存不良呈正相关。Tcl1在体内外促进CRC细胞生长、EMT和stemness。从力学上看,Tcl1直接绑定到mtx,β-catenin负监管机构。此外，这种相互作用依赖于Tcl1的共同激活剂Akt对WTX的磷酸化。一种蛋白激酶磷酸化mtx在ser - 895年之间的交互mtx和β-catenin受损,从而促进β-catenin的转录活动。然而，抗磷酸化的WTX-S895A突变体具有相反的作用。有趣的是,启动子的活性Tcl1正受β-catenin / TCF4复杂,揭露一个积极的反馈回路。重要的是，tcl1水平与人类CRC标本中磷酸化- wtx - s895的表达呈正相关。.结论:我们的研究阐明了一个新颖的机制Tcl1 / mtx /β-catenin轴协调增长,EMT和CRC具备干细胞。这些结果提示了CRC潜在的临床意义。