一个新的mRNA-like非编码RNA功能研究

项目名称： 一个新的mRNA-like非编码RNA功能研究

项目编号： No.30871386

项目类型： 面上项目

立项/批准年度： 2009

项目学科： 医药、卫生

项目作者： 刘木根

作者单位： 华中科技大学

项目金额： 33万元

中文摘要： 通过研究一位染色体平衡易位的KTS病人发病的遗传基础，克隆了一个位于8号染色体易位断裂点附近的新基因，我们将之命名为lncRNA-9q。该基因也是一个进化上的新基因，只在灵长类动物中才发现有同源基因。生物信息学分析和体外翻译实验表明：lncRNA-9q可能是一个mRNA-like non-coding RNA基因，在RNA水平上行使生理功能。染色体易位发生在该基因上游距转录起始点1.3kb处。为分析染色体易位对该基因表达的影响及KTS发病的分子机制，我们对该基因启动子结构进行分析，发现存在5个可能的启动子元件，其中MER61A-LT可能起主要的调控作用；荧光素酶检测实验揭示染色体易位可显著降低lncRNA-9q表达，可能是患者患有疾病的分子基础。通过构建带有链霉素标签的lncRNA-9q融合RNA及pull-down分析，及酵母三杂交的方法，我们分离出一批可能与该RNA基因存在相互作用的蛋白质。我们的研究不仅有助于揭示lncRNA-9q这一mRNA-like non-coding RNA新基因的功能，阐明KTS这一血管发育异常疾病的病理机制，且可为预防和治疗血管相关疾病打下良好基础。

中文关键词： 信使RNA样非编码RNA；易位；转录;启动子

英文摘要： By study a patient associated with Klippel-Trenaunay syndrome, we identified a novel gene which we named it as lncRNA-9q. Bioinformatics studies revealed it is also a new gene evolved only from primates. No whole open read frame is found in the full-length mRNA, and in vitro translation assay failed to identified any protein product from different lncRNA-9q transcripts, suggested that lncRNA-9q may be one new member of gene family of mRNA-like non-coding RNA. The promotor of lncRNA-9q contains endogenous retroviruses repeat elements including MER61A-LTR, MER90A-LTR, LINE1, LIME1-ORF2, and MER110A. Luciferase assay experiments showed that the MER61A-LTR may be the crucial element to regulate expression of lncRNA-9q. To investgate the mechanism by which balance translocation between chromosome 8 and 14 causing KTS, we PCR amplified the 3kb, and 1.3kb of the lncRNA-9q promotor regions, and the promotor region from KTS patient contain t(8:14) translocation, respectively, and fused these fragments to pGL3-basic vector. By analyzed the lucferase activities, we revealed that transocation may significantly down-regulated the expression of lncRNA-9q. In order to elucidate the biological function of lncRNA-9q, we performed experiments to isolate of specific RNA-binding proteins by using methods both "StreptoTag" and Yeast three hybridization, and got some candidate proteins. Our findings will not only be a great help to development effective methods for investigation long non-coding RNA, understand the pathologic mechanism of KTS, but also to develop treatment for blood vessel diseases in the future.

英文关键词： mRNA-like non-coding RNA; translocation; transcription; promoter;bioinformtics