泛素E3连接酶HACE1调控核转录因子NRF2促进神经胶质瘤发生发展的机制研究

Glioma is the most common primary brain tumor in adults, and glioma patients always have a poor prognosis. Glioma tumorigenesis involves multiple genetic and epigenetic alterations, ultimately causing gain-of-function in oncogenes and loss-of-function in tumor suppressor genes. The previous studies have demonstrated that HECT and Ankyrin domain containing E3 ubiquitin ligase 1(HACE1), an E3 ligase, plays a tumor suppressor function in different types of cancer by targeting cyclin D1 and Rac1 for proteasome degradation. However, the biological role of HACE1 in gliomas remains unclear. Our preliminary data showed that HACE1 expression was up-regulated in a cohort of gliomas compared with meningomas, and patients with high HACE1 expression had a poor prognosis compared with those with low HACE1 expression. Moreover, knocking down HACE1 expression in glioma cell lines dramatically inhibited cell proliferation, colony formation, invasion and migration, and induced cell cycle arrest. On the other hand, ectopic expression of HACE1 significantly promoted cell growth and invasiveness, suggesting that HACE1 may play an oncogenic function in glioma tumorigenesis. Notably, we found that HACE1 up-regulated the expression of transcription factor NRF2 at post-transcriptional levels in an ubiquitination-independent mechanism. As a result of our preliminary data, the aims of this project were to explore the biological roles of HACE1 in glioma tumorigenesis and to reveal the molecular mechanism underlying the regulation of NRF2 by HACE1 using microRNA-Seq, RNA pull-down and protein mass spectrum approaches.

神经胶质细胞瘤是最常见的颅内恶性肿瘤，预后差，其发生发展涉及诸多的遗传和表观遗传学改变。泛素E3链接酶HACE1能够特异性泛素化降解癌基因cyclin D1及Rac1，在多种肿瘤中发挥抑癌作用，然而HACE1在胶质瘤中的生物学功能尚不明确。我们前期结果表明HACE1在胶质瘤中高表达，且高表达HACE1与胶质瘤患者的不良预后密切相关，下调胶质瘤细胞系中HACE1的表达能够显著抑制肿瘤细胞的恶性行为。相反，过表达HACE1则促进细胞的侵袭及增殖能力，并且这一作用不依赖于其泛素连接酶活性。随后，我们发现HACE1在转录后水平调控核转录因子NRF2的表达。本项目在此基础上旨在进一步明确HACE1在胶质瘤中的生物学功能，阐明HACE1通过调控NRF2参与胶质瘤发生发展的分子机制，为胶质瘤的治疗提供新的靶点和思路。

HACE1是HECT家族的泛素E3连接酶，有研究发现HACE1基因在多种恶性肿瘤中表达下调或发生突变，并证实其具有抑癌作用。然而，HACE1基因在神经胶质瘤中的生物学作用尚不明确。神经胶质瘤是中枢神经系统最常见的恶性肿瘤，目前治疗方案是以手术联合放、化疗的综合手段，但效果不容乐观。因其放疗敏感性的问题，放疗效果大打折扣。随着分子生物学的深入探索，寻找有效的分子靶点以阻断肿瘤细胞的恶性增殖、逆转多重耐药及如何提高放疗敏感性对神经胶质瘤的治疗具有重大意义。本研究发现HACE1在胶质瘤组织中表达上调，并且高表达的HACE1与低级别胶质瘤患者的远期不良预后相关，尤其是经过放疗的患者。此外，我们通过一系列体内外实验证实HACE1通过激活NRF2促进胶质瘤进展的分子机制，且这一促癌作用并不依赖其泛素连接酶活性。同时，我们的结果还证实HACE1可通过激活NRF2上调细胞内GSH水平，进而导致ROS水平降低，增加胶质瘤细胞对于放疗的抵抗。由此，我们推测HACE1可通过调控NRF2的活性促进胶质瘤进展及放疗抵抗。