Collaborative Research: RESEARCH-PGR: Extracellular RNA Produced By Plants: What, Where, How, Who, and Why?

合作研究：RESEARCH-PGR：植物产生的细胞外 RNA：什么、在哪里、如何、谁以及为什么？

• 批准号: 2141970
• 负责人: Blake Meyers
• 金额: $ 120万
• 依托单位: Donald Danforth Plant Science Center
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-04-01 至 2026-03-31
• 项目状态: 未结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2141970&HistoricalAwards=false
• 关键词: Collaborative; Research; RESEARCH; PGR; Extracellular

This project investigates the role of secreted RNA in the immune system of plants. The Innes and Meyers laboratories recently discovered that the leaves of plants accumulate RNA in the spaces between cells and on their surfaces. Although we usually think of RNA as a molecule that can direct cells to synthesize specific proteins (e.g., the mRNA in COVID vaccines directs our cells to make SARS-CoV2 spike protein), some RNAs serve other functions. Analysis of the base sequences of plant extracellular RNAs revealed that these RNAs are diverse in sequence, but do not appear to encode proteins. The discovery of extracellular non-coding RNA in plants raises two fundamental questions that this project will address: 1) how do plants secrete RNA? and 2) what is the function of this RNA? It takes a large amount of energy for cells to secrete RNA, thus this secreted RNA must benefit the plant in some way. This project will test the hypothesis that secreted RNA functions to protect plants from infection by fungi and bacteria. If this hypothesis is correct, the proposed research will enable generation of crop plants with improved immune systems that are more resistant to disease. Such crops are needed to feed a growing global population in a sustainable manner, while reducing the environmental impacts of agriculture.The Innes and Meyers laboratories recently discovered that the apoplast of Arabidopsis leaves contains abundant long non-coding RNAs, including circular RNAs, as well as small RNAs. These RNAs are bound to protein particles, which protects them against degradation. Notably, this extracellular RNA (exRNA) is highly enriched in the post-transcriptional modification N6-methlyadenine (m6A). These discoveries raise fundamental questions about plant biology: Are there specific exRNAs that are broadly conserved across plant species? How are exRNAs secreted, and are post-transcriptional modifications central to this process? And why do plants produce exRNAs? Do they play a fundamental role in plant-microbe interactions? To answer these questions, exRNA will be purified from the apoplast and leaf surfaces of seven diverse species: Arabidopsis, soybean, tomato, lettuce, pineapple, rice, and maize, which were chosen based on their phylogenetic diversity, genomic resources, importance as crops, and diversity in physiology. These exRNAs will be analyzed using both RNA-seq and sRNA-seq, which will allow identification of RNAs that are conserved between species. To assess whether m6A or other modifications are required for secretion, transgenic plants that express exRNAs that lack modification sites will be tested for their secretion efficiency. To investigate additional requirements for exRNA secretion, the exRNA content in Arabidopsis and rice plants with mutations in known RNA binding proteins and secretory pathway genes will be analyzed. Lastly, to assess whether exRNAs contribute to immunity, mutants compromised in exRNA secretion will be tested for resistance to fungal and bacterial pathogens.This award was co-funded by the Plant Genome Research Program and the Plant Biotic Interactions Program in the Division of Integrative Organismal Systems.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

该项目研究了分泌RNA在植物免疫系统中的作用。 Innes和Meyers实验室最近发现，植物的叶子在细胞和表面之间的空间中积聚RNA。 尽管我们通常将RNA视为可以指导细胞合成特定蛋白质的分子（例如，COVID疫苗中的mRNA指示我们的细胞制造SARS-COV2 Spike蛋白），但一些RNA具有其他功能。 对植物细胞外RNA的碱基序列的分析表明，这些RNA在顺序上是多种多样的，但似乎没有编码蛋白质。 植物中细胞外非编码RNA的发现提出了两个基本问题，该项目将解决：1）植物如何分泌RNA？ 2）此RNA的功能是什么？ 细胞分泌RNA需要大量能量，因此该分泌的RNA必须以某种方式使植物受益。 该项目将测试分泌的RNA功能以保护植物免受真菌和细菌感染的假设。 如果该假设是正确的，则拟议的研究将使生成具有更高耐药性的免疫系统的作物植物。 需要这样的农作物来以可持续的方式养活不断增长的全球人口，同时减少农业的环境影响。世伊Innes和Meyers实验室最近发现，拟南芥叶片的占中心包含丰富的长期非编码RNA，包括环形RNA，以及小型RNA。这些RNA与蛋白质颗粒结合，从而保护它们免受降解。值得注意的是，该细胞外RNA（EXRNA）高度富集在转录后修饰N6-甲乙酰胺（M6A）中。这些发现提出了有关植物生物学的基本问题：在植物物种中是否存在广泛保守的特定exrnas？ Exrnas如何分泌，并且在此过程的转录后修改是核心的吗？为什么植物会产生exrnas？他们在植物 - 微生物相互作用中起着基本作用吗？为了回答这些问题，将从七种多种物种的凋亡和叶片表面纯化：拟南芥，大豆，番茄，生菜，菠萝，菠萝，大米和玉米，这些杂物是根据其系统发育多样性，基因组资源，农作物的重要性以及在物理学中的多样性而选择的。这些EXRNA将使用RNA-Seq和SRNA-Seq分析，这将允许鉴定物种之间保守的RNA。为了评估分泌分泌所需的M6a或其他修饰，表达缺乏修饰位点的Exrnas的转基因植物将用于分泌效率。为了研究ExRNA分泌的其他要求，将分析拟南芥和水稻植物中具有突变的ExRNA含量，其中已知的RNA结合蛋白和分泌途径基因将进行分析。最后，为了评估埃克雷纳斯是否有助于免疫力，将在EXRNA分泌中妥协的突变体对真菌和细菌病原体的抵抗力进行测试。该奖项由植物基因组研究计划和植物生物互动计划共同资助，而植物生物互动计划则在International System of International Infornation Internation Supporiation nsf the NSF的授权措施中，该奖项反映了NSF的合法宣传措施。更广泛的影响审查标准。