利用布氏锥虫BBCP 蛋白研究真核生物中心体的蛋白组成及蛋白功能

In eukaryotes, the microtubule-organizing center is, to a large extent, a central organelle in the cell cycle events, named centrosome. In flagellated cells, centrosome migrates to plasma membrane, further extends and elongates as cilia/flagella. In this case, the centrosome is thus renamed as basal body. In Trypanosoma brucei, a flagellated parasitic protozoon causing African sleeping sickness, is an ideal model for centrosome/basal body studies..In a pioneer study, we have identified a novel centrosome protein termed BBCP in T. brucei, and determined its function to maintain centrosome organization of mother and daughter centrioles. Depletion of BBCP via RNAi results in errors of centrosome replication and lateral death of abnormal cells. However, the mechanism of BBCP working remains unclear..In this proposal, we are planning to characterize the cellular localization signal and functional domains / active sites of BBCP. We are also planning to identify novel centrosome components, which interact with BBCP either directly or indirectly, by applying proteomic techniques, such as immuno-co-precipitation, mass spectrometry, etc. The candidates will be firstly selected by bioinformatics approaches for downstream analysis. In situ tagging and RNAi methods will be preformed to reveal the function of these proteins. Based on these data, an interaction map of BBCP and relevant proteins will be generated, by which a hypothesis would arise to explain how the centrosome organization is maintained and what components are involved. In addition, the data of centrosome mutation could also provide hits for researches for centrosomal diseases, such as polycystic kidney disease, Bardet-Biedl Syndrome and etc. These informations could be helpful for the novel drugs design against trypanosomes, as they are pathogens causing serious diseases in tropical area, monitored by World Health Organization.

中心体是正常细胞复制分裂的关键细胞器。在具有鞭毛/纤毛的细胞，中心体特化成基体，并长出鞭毛/纤毛行使运动感知等重要生物学功能。布氏锥虫是一种重要的人兽共患病病原体，也是目前公认的重要模式生物。申请者在锥虫中发现了一种新的称作BBCP 的蛋白。该蛋白负责维持基体间的正常配对，缺失该蛋白将引起基体复制异常，导致死亡。但是BBCP行使功能的具体机制还不清楚。本项目计划深入研究BBCP 蛋白的定位功能结构域和活性位点。通过免疫共沉淀，质谱分析等分析技术，筛选出与BBCP 相互作用的蛋白，构建基体蛋白相互作用的网络图谱，并用RNA 干扰等分子生物学技术证实相互作用网络上关键蛋白的功能。本研究结果可获得丰富的信息和证据，解释基体蛋白功能行使的分子机制，揭示真核生物中心体及基体的运作机理。并为中心体和基体功能缺陷引起的相关疾病的研究和治疗提供重要的理论基础。同时还能为锥虫病的治疗寻找新的药物靶标。

布氏锥虫是一种重要的人兽共患病病原体，也是目前公认的重要模式生物。申请者在锥虫中发现了一种新的称作BBCP 的蛋白。该蛋白负责维持基体间的正常配对，缺失该蛋白将引起基体复制异常，导致死亡。但是BBCP行使功能的具体机制还不清楚。本项目深入研究BBCP 蛋白的定位功能结构域和活性位点。通过免疫共沉淀，质谱分析等分析技术，筛选出与BBCP 相互作用的蛋白，并用RNA 干扰等分子生物学技术证实相互作用网络上关键蛋白的功能。本研究项目共发表SCI论文7篇，国内核心期刊论文1篇，申请专利3份。培养博士两名，硕士两名。为解释基体蛋白功能行使的分子机制，揭示真核生物中心体及基体的运作机理，锥虫及其他病原寄生虫的检测及防治提供重要的理论基础。