多房棘球绦虫Argonaute蛋白新类群在小RNA诱导的沉默途径中的功能研究

项目名称： 多房棘球绦虫Argonaute蛋白新类群在小RNA诱导的沉默途径中的功能研究

项目编号： No.31472185

项目类型： 面上项目

立项/批准年度： 2015

项目学科： 水产学、兽医学

项目作者： 郑亚东

作者单位： 中国农业科学院兰州兽医研究所

项目金额： 85万元

中文摘要： Argonaute蛋白是组成小RNA诱导的沉默复合体的重要成分。绦虫和吸虫表达3~4个Argonaute蛋白，其中1个为主要结合miRNA的Ago亚家族，其余的则属于新类群Cluster 1，提示它们可能存在除miRNA外的其它小RNA诱导的沉默途径。本研究以多房棘球绦虫为研究模型，通过核酸原位杂交、荧光共聚焦、质谱和新一代测序等方法，分析其表达的3个新类群蛋白的组织分布特征；对比研究3个新类群蛋白所形成的沉默复合体的组成差异，解析它们的小RNA结合谱；体外验证3个新类群蛋白与其它沉默复合体分子和小RNA的相互作用；在沉默或过表达1个新类群蛋白的情况下，确定其它2个新类群蛋白和沉默复合体分子的表达水平，以及小RNA及其靶标分子的丰度，阐明这些新类群蛋白功能上的互补关系。这对揭示绦虫和吸虫小RNA诱导的沉默体系的复杂性和多样性有一定的科学意义，也为认识这些新类群蛋白的生物学功能提供科学证据。

中文关键词： 多房棘球绦虫；Argonaute；蛋白；小RNA

英文摘要： Argonaute protein is a key component of small RNA-induced silencing complex. Cestodes and trematodes express 3~4 Argonaute proteins, 1 of which is grouped into the Ago subfamily and the rest grouped into a novel subfamily Cluster 1, suggesting the presence of other small RNA-induced gene regulation pathways in these species. This study will use Echinococcus multilocularis as a model and, using in situ hybridization, immune confocal microscopy, mass spectrometry and next-generation sequencing techniques, focus on investigation of expression and tissue features of 3 novel Argonautes. The differences in the components of small RNA-induced silencing complex and the small RNA-binding spectra of these 3 novel Argonaute proteins will be comparatively studied. Then the interactions of these Argonautes with small RNAs and other components of the silencing complex will be in vitro verified. Furthermore, the expression levels of small RNAs, other Argonautes and components of the silencing complex will be analyzed in cases of silencing or overexpressing of Argonaute, aiming to illumination of the functional relationship of these Argonautes. These results have implications in revealing of the complexity and diversity of small RNA-induced gene silencing system in cestodes and trematodes, and also provide scientific evidence to understand the functions of these novel Argonautes.

英文关键词： Echinococcus multilocularis;Argonaute protein;small RNA