PlantSynBio: A Novel CRISPR SynBio Tool for Investigating and Reprogramming the Regulation of Alkaloid Biosynthesis in Catharanthus roseus

植物已经发展出复杂的机制，可以对环境的变化做出快速而特异的反应。例如，为了应对病原体、昆虫或食草动物的攻击，植物战略性地将其能量和资源从生长和发育转向生产防御相关化合物。与国防有关的化合物的生产也受到工厂的严格控制，以限制能源和资源的昂贵支出。这项研究的基本科学贡献是了解植物如何在分子水平上快速有效地协调和控制这种防御反应。该项目开发并实施了新的实验工具，以表征和研究调节植物长春花中萜类吲哚生物碱生产的过程。这项研究有可能加快调查，从而解决工程哈代作物和生产重要药物的挑战，同时提供对植物的基本了解。有广泛的跨学科传播这门科学通过教学和指导高中，本科生，和研究生。为了了解植物如何在分子水平上快速有效地协调和控制其防御反应，本研究选择的模式系统是长春花植物。 这种植物产生有价值的化合物，称为萜类吲哚生物碱（TIA），从这种植物的重要TIA包括化疗药物，长春碱和长春新碱。 为了阐明TIA生物合成的调控，本研究的第一个目的是鉴定参与控制TIA生物合成的潜在转录因子。第二个目标是设计、构建和测试一种基于CRISPR-dCas的新型合成生物学工具，以同时但独立地调节多种转录因子的表达。 然后将这种经过验证的合成生物学工具应用于重新编程转基因植物或组织培养物中TIA的调节和产生。 这项研究提供了一种新的合成生物学工具，将可用于植物群落的各种应用，将揭示植物如何快速有效地协调和控制防御反应，该奖项反映了NSF的法定使命，并被认为值得通过使用基金会的知识价值和更广泛的影响审查进行评估来支持的搜索.

Plants have developed elaborate mechanisms for responding rapidly and specifically to changes in their environment. For instance, in response to attack by pathogens, insects, or herbivores, plants strategically redirect their energy and resources from growth and development towards the production of defense-related compounds. The production of defense-related compounds is also carefully controlled by the plant to limit the costly expenditure of energy and resources. The fundamental scientific contribution of this research is to understand how plants rapidly and efficiently coordinate and control this defense response at the molecular level. This project develops and implements novel experimental tools to characterize and investigate the processes that regulate the production of terpenoid indole alkaloids in the plant Catharanthus roseus. The study has the potential to hasten the investigation and consequently the solution to challenges in engineering hardy crops and producing important medicines while providing fundamental understanding of plants. There is broad interdisciplinary dissemination of this science through teaching and the mentoring of high school, undergraduate, and graduate students.To understand how plants rapidly and efficiently coordinate and control their defense response at the molecular level, the model system selected for this research is the Catharanthus roseus plant. This plant produces the valuable compounds known as terpenoid indole alkaloids (TIAs) upon herbivory; important TIAs from this plant include the chemotherapeutic drugs, vinblastine and vincristine. To elucidate the regulation of TIA biosynthesis, the first aim of this research identifies potential transcription factors involved in controlling TIA biosynthesis. The second aim designs, builds, and tests a novel synthetic biology tool based on CRISPR-dCas, to simultaneously but independently regulate the expression of multiple transcription factors. This validated synthetic biology tool is then applied to reprogram the regulation and production of TIAs in transgenic plants or tissue cultures. This research contributes a novel synthetic biology tool that will be made available for diverse applications in the plant community, will reveal how plants rapidly and efficiently coordinate and control the defense response, and will enable strategies for reprogramming and enhancing TIA production.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.