Interaction between host RNA silencing and viral infection in plants

植物中宿主RNA沉默与病毒感染之间的相互作用

• 批准号: 0615154
• 负责人: Anne Simon
• 金额: $ 54万
• 依托单位: University of Maryland, College Park
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-08-01 至 2010-01-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0615154&HistoricalAwards=false
• 关键词: Interaction; between; host; RNA; silencing

One of the most exciting advances in modern molecular biology was the recent discovery that small RNAs regulate numerous cellular activities including development and virus infection through mechanisms referred to as RNA silencing. RNA silencing is an ancient, widespread mechanism for regulating eukaryotic gene expression, and is also used as a means of artificially reducing gene expression in a wide variety of important plants and animals. Based on very limited findings with plant and animal viruses, host small RNAs can be usurped to aid in virus replication or target viruses for destruction. Despite considerable efforts, little is understood about whether small RNAs derived from viral genomes can also control host gene expression, which could have substantial consequences for disease production. This project will combine the best studied viral replication system, Turnip crinkle virus (TCV)and its satellite RNA satC, with the best developed host silencing model, Arabidopsis thaliana. Preliminary experiments have already demonstrated that host enzymes dice TCV and satC into small RNAs that are mainly derived from "hotspot" regions of their genomes. One TCV small RNA tested was able to reduce the expression of at least six host mRNAs, which was related to symptom production and may be providing a more amenable environment for a successful infection. To gain an understanding of (1) the full range of genes targeted by TCV/satC; (2) how the enzymes that generate the viral small RNAs chose their target sequences, and (3) how the TCV coat protein, which naturally suppresses host RNA silencing, contributes to enhancing TCV replication while suppressing satC replication, this research will identify additional cellular targets of TCV and satC vsRNAs, determine the sequence and structural requirements for viral small RNA excision and analyze if the coat protein is responsible for altering the levels of specific small RNAs. This research will help blend together the currently separate fields of RNA virus replication and RNA silencing, dispel incorrect assumptions about RNA virus replication in plants and provide new ideas for the full role of viral RNA silencing suppressors. Successful completion of these experiments should significantly advance our understanding of the interplay between the host RNA silencing pathway and invading viruses with applications to both animal and plant viral pathogenesis, and will likely provide fundamental new information in the broad area of RNA silencing in plants.Broader Impact of the Project (education and outreach):Research and knowledge acquisition is being paired with promotion of teacher training in Prince George's County, a substantially minority school district adjacent to Washington DC. An effort was initiated by the PI in 2005 to improve teacher understanding of cutting edge Biology/Virology at DuVal High School. Teachers reported that their students improved both their understanding and enthusiasm for biology and midyear biology assessments improved by 100%. By also working weekly with AP Biology students, all students took the AP exam (compared with only 2 last year) and all applied to and were admitted to research-oriented universities, which were not their original choices (including two students who will join us at the University of Maryland- which is rare for this school district). This pilot program has led to the development of a two week summer workshop to work with 30 high school teachers selected from throughout the county (50 applications were received). The final lecture of the workshop will incorporate the development of the ideas that led to this research project and progress will also be presented at future planned summer workshops. Continued interaction is planned with AP biology students in the Fall, with each session leading off with the "results of the week", which will allow students to follow how this research is planned and how new ideas are generated. By pairing up all 30 teachers at the workshop with other investigators in the College of Chemical and Life Sciences, many with NSF funded research, the aspiration is to expand this model and excite AP-biology students throughout the county by describing how cutting-edge research is conducted and the role that NSF plays in advancing knowledge acquisition and science education.

现代分子生物学最令人兴奋的进展之一是最近发现小RNA通过称为RNA沉默的机制调节许多细胞活动，包括发育和病毒感染。 RNA沉默是一种古老而广泛的调节真核基因表达的机制，也被用作人为降低多种重要动植物基因表达的手段。基于对植物和动物病毒的非常有限的发现，宿主小RNA可以被篡夺来帮助病毒复制或针对病毒进行破坏。尽管付出了巨大的努力，但人们对源自病毒基因组的小RNA是否也能控制宿主基因表达知之甚少，这可能对疾病的产生产生重大影响。该项目将把研究最深入的病毒复制系统——芜菁皱病毒（TCV）及其卫星RNA satC，与最先进的宿主沉默模型——拟南芥结合起来。初步实验已经证明，宿主酶将 TCV 和 satC 切成小 RNA，这些小 RNA 主要源自其基因组的“热点”区域。测试的一种 TCV 小 RNA 能够减少至少 6 种宿主 mRNA 的表达，这与症状的产生有关，并且可能为成功感染提供更适宜的环境。了解 (1) TCV/satC 靶向的全部基因； (2) 生成病毒小 RNA 的酶如何选择其靶序列，以及 (3) TCV 外壳蛋白（自然抑制宿主 RNA 沉默）如何有助于增强 TCV 复制，同时抑制 satC 复制，这项研究将鉴定 TCV 和 satC vsRNA 的其他细胞靶点，确定病毒小 RNA 切除的序列和结构要求，并分析外壳是否 蛋白质负责改变特定小RNA的水平。这项研究将有助于将目前RNA病毒复制和RNA沉默这两个独立的领域融合在一起，消除关于RNA病毒在植物中复制的错误假设，并为病毒RNA沉默抑制子的全面作用提供新思路。成功完成这些实验将显着促进我们对宿主 RNA 沉默途径与入侵病毒之间相互作用的理解，并应用于动物和植物病毒发病机制，并可能在植物 RNA 沉默的广泛领域提供基本的新信息。 该项目的更广泛影响（教育和推广）：研究和知识获取与促进乔治王子县（一个主要是少数族裔学区）的教师培训相结合 毗邻华盛顿特区。 PI 于 2005 年发起了一项努力，旨在提高杜瓦尔高中教师对前沿生物学/病毒学的理解。老师们报告说，学生们对生物学的理解和热情都得到了提高，年中生物学评估提高了 100%。通过每周与AP生物学学生一起工作，所有学生都参加了AP考试（去年只有2人），并且全部申请并被研究型大学录取，这不是他们最初的选择（包括两名将加入我们马里兰大学的学生——这在这个学区是罕见的）。该试点计划催生了为期两周的夏季研讨会，与从全县选出的 30 名高中教师合作（已收到 50 份申请）。研讨会的最后一场讲座将纳入导致该研究项目的想法的发展，并且进展也将在未来计划的夏季研讨会上介绍。计划在秋季与 AP 生物学学生继续互动，每次会议都会以“本周的结果”开始，这将使学生能够了解这项研究是如何计划的以及新想法是如何产生的。通过将研讨会上的所有 30 名教师与化学与生命科学学院的其他研究人员（其中许多研究人员从事 NSF 资助的研究）配对，我们的愿望是通过描述前沿研究的进行方式以及 NSF 在推进知识获取和科学教育方面所发挥的作用来扩展这一模式，并激发全县 AP 生物学学生的兴趣。