线性泛素连接酶HOIP通过Hedgehog信号促进结肠癌细胞增殖和耐药的作用及分子机制

Aberrant activation of Hedgehog (Hh) signaling firmly relates with the initiation, development and drug resistance in multiple tumors, including colorectal cancer (CRC). Ubiquitination of Gli proteins, the key transcriptional factors in Hh signaling pathway, plays essential role in Hh signaling. HOIP is the only known E3 ligase mediating linear ubiquitination, however, the function of HOIP in mediating the ubiquitination of Gli proteins and facilitating tumorigenesis and drug resistance remains unknown..Our previous data claimed that the expression of HOIP is higher in CRC, compared with the adjacent tissue, and the highly expression of HOIP promotes cell proliferation and drug resistance in CRC cells. Further study revealed that HOIP would activate Hh signaling pathway via stabilizing transcriptional factor Gli..Thus, we hypothesized that HOIP might stabilize Gli through mediating its linear ubiquitination, activate Hh signaling pathway, and facilitate cell proliferation and tumor growth in CRC..Herein, we could, integrating various biochemical and molecular biological methods, investigate the molecular mechanism how HOIP stabilizes Gli via mediating its linear ubiquitination, and regulates Hh signaling pathway in the cellular level. And we would also declare the physical function of HOIP in facilitating tumor growth and drug resistance of CRC in vivo by xenograft model and AOM-DSS induced colon cancer in mice. Furthermore, the relationship between the expression level of HOIP and prognosis of CRC would be generated via clinical cohort..Through this research, we would claim a novel mechanism how Hh signaling is regulated in tumor tissues theoretically, and offer potential targets to CRC diagnosis and therapy.

Hedgehog（Hh）信号过度活化，与结肠癌等肿瘤的发生发展和耐药密切相关。Gli蛋白是Hh信号通路中的关键转录因子，其泛素化修饰在Hh信号转导中起到重要作用。HOIP是已知的唯一的介导线性泛素化的E3连接酶，但其对Gli的泛素化修饰及在结肠癌中的病理功能未见相关报道。.我们前期研究发现：1）HOIP在结肠癌组织中表达高于癌旁组织；2）HOIP高表达促进结肠癌细胞增殖和耐药；3）HOIP能通过稳定转录因子Gli激活Hh信号。据此提出科学假设：线性泛素连接酶HOIP通过稳定Gli并增强Hh信号，从而促进结肠癌细胞增殖和耐药。.本项目中，我们拟综合利用多学科技术，研究HOIP对Gli蛋白的线性泛素化修饰，构建细胞和动物模型探索HOIP通过Hh信号促进结肠癌细胞增殖和耐药的作用与机制，并通过临床队列分析建立HOIP与结肠癌患者预后的相关性，为结肠癌诊治提供新的靶点。

线性泛素链组装复合物（LUBAC，Linear ubiquitin chain assembly complex）是唯一能介导线性泛素化修饰的E3连接酶复合物，HOIP是该复合物的催化亚基。目前认为，线性泛素化修饰参与传递TNF信号并调控免疫过程。然而，线性泛素化修饰在肿瘤相关信号通路中的作用，以及其对肿瘤发生发展的影响研究较少。.本项目中，我们发现Hedgehog（Hh）信号通路的核心转录因子Gli蛋白是线性泛素化修饰的底物之一，HOIP和LUBAC可以介导Gli蛋白的线性泛素化，并稳定Gli蛋白，从而激活Hh信号。Hh信号在结肠癌等肿瘤的发展中有着重要的作用，因此我们也对HOIP在结肠癌中的作用进行了研究。我们发现，HOIP可以在体内和体外促进结肠癌细胞增殖生长。与此一致的是，干扰HOIP表达或使用小分子抑制剂（HOIPIN-1）抑制其活性，都可以抑制结肠癌细胞增殖生长。有趣的是，在正常的肠上皮细胞中，抑制HOIP的活性，对细胞增殖没有显著的影响。.综上所述，通过本项目的研究，我们发现HOIP和LUBAC可以通过介导Gli蛋白线性泛素化修饰激活Hh信号，促进结肠癌细胞增殖生长。该研究有助于加深对结肠癌发展机制的认识，为结肠癌的诊断和治疗提供理论依据。值得一提的是，我们发现HOIP的小分子抑制剂HOIPIN-1可以在不影响正常肠上皮细胞生长的同时，抑制结肠癌细胞增殖生长，为结肠癌的靶向治疗提供了新的潜在治疗靶点。