抑癌基因blu与纤毛发生、细胞周期的关系研究

blu is a tumor suppressor gene (TSG) and its expression loss or inactivation, due to promoter hypermethylation, has been reported in various different cancers， including Nasopharyngeal carcinoma(NPC) and lung cancer. As one of the earliest changes in tumor formation, however, the molecular mechanism of Blu is still poorly understood. Here, we plan to investigate its molecular function using zebrafish as a model system. Zebrafish has many advantages for genetic studies, such as fast development and body transparency. With whole mount in situ hybridization approach, we found that zebrafish blu gene is enriched in ciliated tissues, such as Kupffer's Vesicle, pronephric duct and olfactory placode. We hypothesize that Blu is involved in ciliogenesis and it may interact with ciliary proteins, such as HDAC6, to promote cilia formation and thus regulate cell cycle entry. We are going to test this hypothesis both in vitro and in vivo. The goals of this study include: (1) Screen zebrafish blu mutants and determine its function during ciliogenesis and cell cycle control; (2) Identify Blu binding partners. Through this works, it will help us to better understand the molecular mechanism of blu gene and also benefit the early diagnoses of various cancers.

blu基因是一个抑癌基因。在鼻咽癌、肺癌等常见肿瘤形成的初期，blu基因的启动子常发生超甲基化，从而导致该基因的表达降低或缺失。作为肿瘤形成的早期标记基因之一，目前对其具体的抑癌分子机制知之甚少。本立项拟以斑马鱼为模式生物，研究Blu抑制肿瘤形成的分子机理。斑马鱼Blu存在于富集纤毛的组织中，我们推测Blu的抑癌机制与纤毛发生及细胞周期调控密切相关，并将通过遗传及生化的分析方法对该假设进行验证。预期本项工作将取得以下研究成果：（1）筛选鉴定斑马鱼blu突变体，并阐明其表达缺失与斑马鱼肿瘤形成的联系；（2）明确Blu与纤毛发生、细胞周期调控的关系。通过本研究，将会使我们对Blu的具体抑癌分子机理有更深入的认识。同时，对多种肿瘤形成的早期分子鉴定提供理论依据。

纤毛的运动障碍可导致多种人类疾病，包括内脏反转，脑积水以及不孕不育等，但目前对纤毛的运动机制还不甚了解，近一半的不动纤毛综合症（PCD）患者的致病基因还不清楚。本研究以斑马鱼为模式生物，研究了一个与运动纤毛相关的基因blu/zmynd10的功能，并在此基础上深入探讨了纤毛运动所导致胚胎体轴发育缺陷的分子机制。本项目的主要研究发现包括：1）利用原位杂交筛选的方法鉴定出zmynd10为一个在运动纤毛富集组织中特异表达的基因；2）利用TALEN技术筛选出该基因的突变体，对突变体的分析表明zmynd10功能缺失可导致运动纤毛的运动障碍，进而导致胚胎多囊肾，体轴向腹部严重弯曲等表型；3）利用荧光染料示踪及转基因分析，发现突变体体轴的弯曲主要与神经底板处运动纤毛的缺陷有关，而与肾脏运动纤毛无关；4）通过对突变体基因表达的转录组分析，发现up1和up2是两个与体轴发育相关的关键基因。该基因主要在神经底板处神经元内表达，并受运动纤毛所调控，其缺陷可导致胚胎体轴弯曲；5）发现up1和up2通过运动神经元调节躯干肌肉细胞的收缩，进而控制胚胎体轴的发育。上述研究成果系统的阐述了运动纤毛通过调控神经系统，进而影响到肌肉收缩的分子机制，解决了长期以来困扰的有关斑马鱼纤毛突变体体轴弯曲的具体原因，为进一步利用斑马鱼研究人类纤毛疾病提供了理论支持。