Elucidation of translational regulatory mechanisms of plant immune responses

在过去的二十年里，在理解植物如何控制编码防御蛋白的信使RNAs的产生方面取得了重大进展，但对于这些信使RNAs的翻译是否以及如何调节以产生蛋白质的了解很少。随着基于测序的测量信使RNA翻译速率的技术的发展，很明显，防御蛋白的合成在植物中受到严格的调控。本项目旨在了解这一调控是如何进行的，并确定信使RNA的功能和参与这一调控的重要分子。该项目的成功不仅将填补植物免疫机制方面的知识空白，而且将通过更精确地控制防御蛋白的生产来彻底改变抗病作物的工程策略。这一新策略将能够在不影响植物适应性的情况下培育广谱抗病能力，从而减少杀虫剂的使用，减轻抗药性病原体的选择压力。该项目还将对从高中生到博士后研究人员进行前沿研究的科学家进行培训。该项目的长期目标是阐明植物免疫反应的翻译调控机制。对植物免疫转录因子TBF1的研究表明，其翻译受两个上游开放阅读框(UORFs)控制。这些uORF通常会阻止核糖体到达主要的ORF。在免疫诱导后，这种抑制效应迅速减轻，导致TBF1翻译。这表明，去抑制已存在的免疫组织因子mRNA的翻译可能是一种快速响应病原体攻击的策略。利用核糖体足迹技术进行全球翻译组谱分析表明，在模式触发免疫(PTI)期间，翻译的变化与转录没有很好的相关性。PTI过程中翻译效率(TE)改变的基因属于不同的功能群，包含这种免疫反应的新调节因子。此外，在PTI过程中，随着TE的增加，转录产物的5‘前导区显著富含mRNA共识(ur基序)。该项目旨在建立更多的翻译组图谱和第一个植物tRNAome数据集，这对于研究植物防御反应过程中的全球翻译变化至关重要，并阐明tRNA水平的变化、mRNA序列特征(如uORF和R基序)以及反式作用因子如何调节翻译。该项目将深入了解植物如何选择性地翻译与防御相关的蛋白质，并识别不同免疫反应的新调节器。该项目还将培训两名博士后研究人员，并使外联活动得以继续，包括为当地高中和社区大学生举办实验室之旅，以及为SROP计划的少数族裔本科生暑期研究提供指导。

In the past two decades, significant progress has been made in understanding how plants control the production of messenger RNAs encoding defense proteins when challenged by pathogens, but very little is known about whether and how translation of these messenger RNAs is regulated to produce proteins. With the development of a sequencing-based technology for measuring the rate of messenger RNA translation, it becomes clear that defense protein synthesis is tightly regulated in plants. This project aims to understand how this regulation is carried out and to identify messenger RNA features and important molecules involved in this regulation. The success of this project will not only fill a gap in the knowledge of the plant immune mechanisms, but also revolutionize strategies of engineering disease resistant crops by more precisely control defense proteins production. This new strategy will enable engineering broad-spectrum disease resistance without compromising plant fitness, leading to reduced use of pesticides and lightening of selective pressure for resistant pathogens. This project will also result in training of scientists from high school students to postdoctoral researchers in performing frontier research. The long-term goal of this project is to elucidate the translational regulatory mechanisms of plant immune responses. Studies of TBF1, a key plant immune transcription factor (TF), showed that its translation is controlled by two upstream open reading frames (uORFs). These uORFs normally block ribosomes from reaching the major ORF. Upon immune induction, this inhibitory effect is rapidly alleviated, leading to TBF1 translation. This suggests that de-repressing translation of pre-existing immune TF mRNA may be a strategy for rapid response to pathogen challenge. Global translatome profiling using ribosome footprinting showed that during pattern-triggered immunity (PTI), changes in translation did not correlate well with transcription. Genes with altered translational efficiency (TE) during PTI belong to diverse functional groups and contain novel regulators of this immune response. Moreover, an mRNA consensus (uR-motif) was found to be significantly enriched in the 5' leader regions of transcripts with increased TE during PTI. This project aims to generate more translatome profiles and the first plant tRNAome datasets, essential for studying global translational changes during plant defense responses, and elucidate how tRNA level changes, mRNA sequence features (e.g., uORFs and R-motif), and trans-acting factors regulate translation. The project will lead to insight into how plants selectively translate defense-related proteins and identify new regulators of different immune responses. The project will also involve training of two postdoctoral researchers and enable continuation in outreach activities, including hosting lab tours for local high school and community college students, and mentoring for the SROP Program for minority undergraduate summer research.