Gene silencing in schistosomes using RNAi

使用 RNAi 实现血吸虫基因沉默

• 批准号: 7013596
• 负责人: Patrick J Skelly
• 金额: $ 38.69万
• 依托单位: TUFTS UNIVERSITY BOSTON
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-03-15 至 2009-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7013596
• 关键词: RNA interference; Schistosoma; double stranded RNA; endopeptidases; functional /structural genomics; gene expression; gene induction /repression; hemoglobin; laboratory mouse; life cycle; metabolism; method development; nutrient intake activity; parasite infection mechanism; protein degradation

DESCRIPTION (provided by the applicant): Schistosomes are extracellular blood worms that infect over 200 million people globally and cause several thousand deaths annually. Large-scale genome sequencing projects for each of the three major human schistosome species: Schistosoma mansoni, S. haematobium and S. japonicum, are currently underway. Despite the wealth of new data to be generated by all of these undertakings, there is still no routine technique available that allows us to exploit the data through manipulation of the schistosome genome. Because of this, our understanding of the molecular and cellular biology of schistosomes has been severely hampered and lags behind that of most other important human pathogens. In this application, we propose to utilize a relatively new technology in gene manipulation called RNA-mediated interference (RNAi) to examine gene function in schistosomes. In preliminary experiments, we have achieved considerable inhibition of select genes involved in nutrient acquisition by the parasites. We plan to build on this success by first optimizing the protocol for gene suppression using RNAi in different schistosome life cycle stages by varying the amount and nature of the dsRNA employed, the mode of delivery and the duration of exposure. Such a systematic assessment of the relative importance of the factors that impinge on the phenomenon could have wider applicability for researchers using RNAi in other biological systems. Next we will utilize the optimized protocol to test specific hypotheses concerning an important area of schistosome biology: The involvement of proteases in hemoglobin degradation and nutrient acquisition. The work proposed here is designed to provide a simple, powerful and widely applicable protocol for the schistosome research community to facilitate functional schistosome genomics. In addition, our application of the technology is designed to provide significant new information about schistosome metabolism and biology.

描述(由申请人提供)：血吸虫是一种胞外血虫，每年感染全球2亿多人，并导致数千人死亡。目前正在对三种主要的人类血吸虫物种：曼氏血吸虫、血吸虫和日本血吸虫进行大规模基因组测序项目。尽管所有这些工作都将产生丰富的新数据，但仍然没有常规技术允许我们通过操纵血吸虫基因组来利用这些数据。正因为如此，我们对血吸虫的分子和细胞生物学的了解受到了严重的阻碍，并且落后于大多数其他重要的人类病原体。在这一应用中，我们建议利用一种相对较新的基因操作技术，称为RNA介导的干扰(RNAi)来检测血吸虫的基因功能。在初步实验中，我们已经实现了对参与寄生虫获取营养的选定基因的相当大的抑制。我们计划在这一成功的基础上，首先通过改变所使用的dsRNA的数量和性质、传递方式和暴露时间来优化在血吸虫不同生命周期阶段使用RNAi进行基因抑制的方案。这种对影响这一现象的因素的相对重要性的系统评估可能会对在其他生物系统中使用RNAi的研究人员具有更广泛的适用性。接下来，我们将利用优化的方案来测试有关血吸虫生物学的一个重要领域的特定假设：蛋白水解酶在血红蛋白降解和营养获取中的参与。这项工作旨在为血吸虫研究界提供一个简单、强大和广泛适用的方案，以促进血吸虫功能基因组学的发展。此外，我们应用这项技术是为了提供有关血吸虫新陈代谢和生物学的重要新信息。