Collaborative Research: PlantSynBio: Deciphering the roles of genetic and biochemical redundancy and pathway regulation via refactoring the protective plant cuticle

作为面对不断变化的环境生存的固定生物体，植物进化出了特殊的功能来抵御环境压力。 这些特征之一是气生植物表面的外部保护屏障，称为角质层。角质层充当植物与其环境之间的物理屏障，起到限制水和气体损失的作用。尽管许多在角质层形成过程中发挥作用的关键基因已被识别，但仍缺乏对角质层如何构建的整体了解。该项目将设计两个通常没有角质层（酵母细胞和植物根）的新型并行合成生物学系统，从头开始构建角质层，并破译这种独特植物特征背后的生化途径的复杂性。系统地确定角质层的构建方式将带来重要的应用，例如培育具有定制角质层的农作物，这些角质层可能会增强对环境压力的耐受性，以及用于生物可再生行业的受角质层启发的化学品。此外，该项目将培训下一代多学科科学家，并建立教学和研究计划，最终目标是增加来自 STEM 背景不足的科学劳动力的比例。该多学科项目将在不自然产生角质层的系统中构建和测试两个协同合成生物学底盘（即植物根和酿酒酵母），以系统地重构转录调控网络以及组装保护性、疏水角质层屏障。这两种合成底盘将用于全面建模和定量理解组装功能性植物角质层的集成机制。根底盘将用于研究植物转录因子对角质层组装的协调激活。该底盘将提供时间转录和代谢数据，以实现动态预测模型的开发，从而提供角质层代谢及其相关调节的整体视图。第二个底盘依赖于多个工程酵母即插即用系统，表达不同的遗传互补物，捕获途径内的基因冗余，将评估合成角质层成分的生产。这些菌株产生的代谢数据将作为动力学建模的输入，这将提供对这一复杂途径的第一个动力学理解。植物根和酵母底盘的协调发展与所提出的计算框架相结合，将为角质层组装的发现和系统分析提供一个新的平台。该奖项由分子和细胞生物科学部门的系统和合成生物学集群以及综合有机系统部门的植物基因组研究计划共同资助。该奖项反映了 NSF 的法定使命，并通过使用基金会的智力价值和更广泛的影响审查标准进行评估，被认为值得支持。

As stationary organisms that are faced with surviving constantly changing environments, plants have evolved specialized features to protect against environmental stresses. One of these features is an exterior protective barrier on aerial plant surfaces called the cuticle. The cuticle acts as a physical barrier between the plant and its environment, functioning to limit the loss of water and gasses. Although many key genes that function in making the cuticle have been identified, a holistic view of how the cuticle is built is missing. This project will engineer two novel, parallel synthetic biology systems that are normally devoid of a cuticle (yeast cells and plant roots) to build a cuticle from scratch and decipher the complexities of the biochemical pathways underlying this unique plant feature. Systematically determining how a cuticle is built will lead to important applications such as the breeding of crop plants with customized cuticles that may have enhanced tolerance to environmental stresses, as well as cuticle-inspired chemicals for the biorenewables industry. Moreover, this project will train the next generation of multi-disciplinary scientists, and build teaching and research initiatives with the ultimate goal of increasing the proportion of the scientific workforce who are from STEM-underrepresented backgrounds.This multi-disciplinary project will build and test two synergistic synthetic biology chassis in systems that do not naturally produce a cuticle (i.e., plant roots and the yeast Saccharomyces cerevisiae) to systematically refactor the transcriptional regulatory network, and the metabolic pathways that assemble the protective, hydrophobic cuticle barrier. These two synthetic chassis will be used to comprehensively model and quantitatively understand the integrated mechanisms that assemble a functional plant cuticle. The root chassis will be used to study the coordinated activation of cuticle assembly by plant transcription factors. This chassis will provide temporal transcriptional and metabolic data to enable the development of dynamic predictive models that provide a holistic view of cuticle metabolism and its associated regulation. A second chassis relies on multiple engineered yeast plug-and-play systems expressing different genetic complements capturing the gene redundancies within the pathway that will be assessed for the production of synthetic cuticle constituents. The metabolic data generated from these strains will be the inputs for kinetic modeling, which will provide the first kinetic understanding of this complex pathway. The coordinated development of the plant root and yeast chassis in combination with the proposed computational framework will provide a novel platform for discovery, and systematic analysis of cuticle assembly.This award was co-funded by the Systems and Synthetic Biology Cluster in the Division of Molecular and Cellular Biosciences and by the Plant Genome Research 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.