Function of Arabidopsis Small RNA-ARGONAUTE Complexes

智力价值：植物和动物的基因组已经进化到控制发育、对环境变化的反应和入侵病原体所需的基因的功能。这些生物体内的许多基因都是由微小的RNA分子控制的，这种分子被称为微RNA(MiRNA)和短干扰RNA(SiRNA)，通过一种称为基因沉默的过程。小RNA通过充当序列特异的向导，将ArgAerte(AGO)家族的蛋白质输送到靶向RNA分子。然后，AGO蛋白可以导致目标RNA的降解、错误定位或抑制，从而实现基因调控。模式植物拟南芥含有多达10个具有功能的AGO蛋白，其中大约一半被分配了至少一些功能，或被证明与一组特定的小RNA相互作用。广泛的分析揭示了与植物基因组中的序列相对应的多个小RNA(通常为21-24个核苷酸)的高度丰富、复杂的景观。然而，小RNA的活动，而不是简单的积累，是在全基因组范围内理解基于小RNA的沉默的关键。一旦形成了小RNA，以前的小RNA复合体是如何形成的，以及形成的程度如何？一旦这些复合体形成，哪些对其他RNA分子具有靶向作用？对于主要的AGO蛋白(Ago1和AGO10)，靶RNA池在稳定和翻译抑制水平上是如何受到影响的？本项目通过关注如何处理和管理沉默信息来解决这三个问题中的每一个。了解基因沉默是如何在分子和基因组水平上发挥作用的，将有助于阐明植物如何利用和控制大量遗传信息。更广泛的影响：该项目将产生比实际科学成果更广泛的重大影响。该项目涉及本科生在一线参与研究。在项目期间，两名学生研究人员将得到支持。该项目还为两名博士后科学家提供了明确的指导计划，并参与了与科学家和教师教育伙伴关系(STEPS，www.steps.org)的外联合作，该伙伴关系是俄勒冈州立大学的一个项目，为K-12教师提供培训，并让学生接触实验科学。P.I.与6-12年级的STEPS项目协调员Kari Van Zee博士合作，开发生物技术进步系列的夏季研讨会。它为教师提供适合学校的实验室实验的专业发展研讨会、讲座和动手培训。拟南芥小RNA项目数据库是前一个由国家科学基金会资助的项目产生的信息和数据的主要门户，将继续在公共领域为任何寻求全基因组小RNA数据的人提供服务。为处理和分析高通量测序数据而开发的程序，如CASHX和TargetFinder，将继续通过ASRP网站提供。最后，该项目正在引导更好的方法来利用植物固有的遗传潜力来满足我们的食物、纤维和能源需求。

Intellectual Merit:The genomes of plants and animals have evolved to control the function of genes required for development, response to environmental change, and invasive pathogens. Many of the genes within these organisms are controlled by tiny RNA molecules, called microRNA (miRNA) and short-interfering RNA (siRNA), through a process called gene silencing. The small RNA delivers proteins in the ARGONAUTE (AGO) family to target RNA molecules by functioning as sequence-specific guides. The AGO protein can then cause degradation, mislocalization, or repression of the target RNA, thereby achieving gene regulation. The model plant Arabidopsis contains up to 10 functional AGO proteins, roughly one-half of which have been assigned at least some function or shown to interact with a set of specific small RNA. Extensive analyses have revealed highly abundant, complex landscape of multiple small RNAs (generally 21-24 nucleotides) corresponding to sequences across the plant genome. However, small RNA activity, rather than simply accumulation, is key to understanding small RNA-based silencing on a genome-wide scale. Once small RNA are formed, how and to what extent are AGO-small RNA complexes formed? Once these complexes form, which are active to target other RNA molecules? For the major AGO proteins (AGO1 and AGO10), how are target RNA pools affected at the stabilization and translational repression levels? This project addresses each of these three questions by focusing on how silencing information is processed and managed. Understanding how gene silencing works at the molecular and genome-wide levels will shed light on how plants use and control a large amount of genetic information. Broader Impacts:The project will have significant broader impact beyond the actual scientific results. The project involves front-line participation of undergraduate students in research. Two student researchers will be supported for the duration of the project. The project also provides a clear mentoring plan for two postdoctoral scientists, and involves participation in an outreach collaboration with the Scientists and Teachers Education Partnerships (STEPS, www.steps.org), an Oregon State University-based program that provides training to K-12 teachers and exposure of students to experimental science. The P.I.s work with the STEPS Program Coordinator for Grades 6-12, Dr. Kari Van Zee, to develop summer workshops in the Advances in Biotechnology series. This provides teachers with professional development workshops, lectures, and hands-on training for school-appropriate laboratory experiments. The Arabidopsis Small RNA Project Database, the primary portal for information and data generated by the previous NSF-funded project, will continue to operate in the public domain for anyone seeking genome-wide small RNA data. Programs developed to process and analyze high-throughput sequencing data, such as CASHX and TargetFinder, will continue to be made available through the ASRP website. Finally, this project is leading to better ways to use the inherent genetic potential of plants for our food, fiber, and energy needs.