PlantSynBio: Regulatory Systems to Tune Gene Expression in Synthetic Chloroplast Operons

在过去的50年里，对基础研究的投资得到了回报，在理解基本的生物过程方面取得了显著的进展，包括遗传、新陈代谢和生物发育的机制。合成生物学学科在工程设计原则的框架内将这种理解应用于实际问题。该项目将利用这一框架来应用在了解植物基因表达机制方面的最新进展，以改善农艺和营养性状，并促进将植物用作制药和其他有益生物化学品的“绿色”工厂。从基础研究中推断出的基本原则将被用于设计遗传“部分”，以优化赋予有益特性的基因的表达。该项目还将为本科生和博士后提供研究培训，并将邀请高危高中生参加营养生物化学的动手研讨会，这是俄勒冈大学旨在鼓励这些学生进入大学并取得成功的更广泛项目的一部分。该项目的总体目标是开发代谢工程、生物技术和增强植物农艺性状的工具。许多这样的应用需要多个基因的表达，最佳结果取决于它们适当的平衡表达。叶绿体遗传系统为多蛋白组合和多酶代谢途径的表达提供了特别的希望，因为叶绿体基因自然地以操纵子样的多顺反子转录单位表达，它们不经历表观遗传效应，并且它们可以达到非常高的表达水平。然而，目前的限制是缺乏特征化的遗传元件来编程在广泛的动态范围内可预测的表达，特别是为了多基因应用的目的。该项目旨在通过利用以下方面的最新进展来填补这一空白：(I)了解决定天然叶绿体操纵子表达水平的机制，(Ii)阐明五肽重复蛋白识别序列特异性RNA的基础，这是这一调控的关键，以及(Iii)对天然叶绿体基因的翻译效率进行编目。这一深厚的知识库将用于合理设计顺式元件和同源反式因子的调色板，以在合成的多基因转录单元中实现可预测、可调节和适当调整的基因输出。该奖项由整合组织系统部门的植物基因组研究计划以及分子和细胞生物科学部门的系统和合成生物学集群共同资助。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

Investments in basic research over the past five decades have been rewarded with remarkable advances in understanding fundamental biological processes, including mechanisms of heredity, metabolism, and organismal development. The discipline of synthetic biology applies this understanding to practical problems within a framework of engineering design principles. This project will use this framework to apply recent advances in understanding mechanisms of gene expression in plants to improve agronomic and nutritional traits, and to foster the use of plants as “green” factories for pharmaceuticals and other beneficial biochemicals. Foundational principles inferred from basic research will be used to design genetic “parts” to optimize the expression of genes that confer beneficial properties. This project will also provide research training for undergraduate and postdoctoral students, and it will engage at-risk high school students in hands-on workshops in nutritional biochemistry as part of a broader program at the University of Oregon that aims to encourage such students to enroll and succeed in college.The overarching goal of this project is to develop tools for metabolic engineering, biotechnology, and enhancement of agronomic traits in plants. Many such applications require expression of multiple genes, with optimal outcomes depending upon their suitably balanced expression. The chloroplast genetic system offers particular promise as a chassis for the expression of multiprotein assemblies and multi-enzyme metabolic pathways because chloroplast genes are naturally expressed in operon-like polycistronic transcription units, they do not experience epigenetic effects, and they can achieve very high expression levels. A current limitation, however, is a paucity of characterized genetic elements to program predictable expression over a wide dynamic range, especially for the purpose of multigene applications. This project aims to fill this gap by leveraging recent advances in (i) understanding mechanisms that determine expression levels in natural chloroplast operons, (ii) elucidating the basis for sequence-specific RNA recognition by pentatricopeptide repeat proteins, which are key to this regulation, and (iii) cataloging translational efficiencies of native chloroplast genes. This deep knowledge base will be used for the rational design of a palette of cis-elements and cognate trans-factors to achieve predictable, regulatable, and suitably tuned gene outputs in synthetic multigene transcription units.This award was co-funded by the Plant Genome Research Program in the Division of Integrative Organismal Systems and the Systems and Synthetic Biology Cluster in the Division of Molecular and Cellular Biosciences.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.