An Auxin Toolbox for Synthetic Multicellular Systems

该奖项由MCB的系统和合成生物学计划以及CBET的生物技术、生化和生物质工程计划资助，支持植物使用的生长素信号系统的研究。生长素是一种激素，几乎存在于植物的每一个部位，对各种细胞反应是必不可少的。然而，目前还不清楚植物中的每个细胞如何对生长素信号做出不同的反应，这取决于它是什么类型的细胞，它的位置和历史。了解植物如何利用生长素信号来协调多细胞行为，将有助于阐明植物如何生长和对刺激做出反应，并可能为设计定制的细胞-细胞通讯系统提供新的工程工具和方法。为了开发一个用生长素来编程多细胞行为的工具箱，关键基因将从植物拟南芥中的生长素感应途径转移到酵母中。实验室酵母菌是表征生长素途径的理想方法，因为它易于操作，并具备处理生长素所需的所有先决条件，但在其他方面基本上对生长素不敏感。对大量模拟植物中发现的生长素信号电路的合成生长素信号电路的严格定量表征将与数学建模和工程设计工具相结合，以建立对该途径的可编程性的理解。更广泛地说，这项工作可能会导致在人类组织工程中的新应用，以及改良重要作物物种的新方法。该项目还将支持开发合成生物学和细胞信号的新本科和K-12教育模块，使用生长素/酵母系统为例。技术描述：酵母中的合成细胞-细胞信号电路将使用遗传部分、途径和来自植物的灵感来构建。蛋白质成分将在不同的家庭成员之间变化，以探索电路是如何调节的。对电路调谐中细微差异的数学建模和表征将阐明生长素部分的可编程性。新的细胞-细胞行为的演示将作为该方法原理的证明。具体地说，将构建基于生长素受体(TIR1、AFB2)、转录因子(ARF)、辅助因子(AUX/IAA)的生长素信号处理电路。通过将这些组件适当地组合在一起，将构建简单的信号处理电路，如过滤器、中继器和脉冲发生器，并在酵母中表征它们的动态行为。在酵母中将构建一个生长素生物合成模块，由催化生长素产生的酶组成，并受生长素信号处理电路控制，以实现一种新的细胞间通信模式。生长素感应电路将控制生长素的产生，以实现过滤和阈值等信号功能。发送、接收和处理生长素的能力将使多细胞行为的研究成为可能，如波传播或共识。这一结果不仅将为设计多细胞系统提供新的工具，还将为植物如何以及为什么如此有效地利用生长素提供生物学假说。

The award funded by the Systems and Synthetic Biology Program in MCB and the Biotechnology, Biochemical and Biomass Engineering Program in CBET supports research into the auxin signaling system used by plants. Auxin is a hormone present in almost every part of the plant and is essential for diverse cellular responses. However, it remains unclear how each cell in the plant responds to auxin signals differently depending on what type of cell it is, its location, and its history. Understanding how plants use auxin signaling to coordinate multicellular behaviors will shed light on how plants grow and respond to stimuli and may also yield new engineering tools and methods for designing custom cell-cell communication systems. To develop a toolbox for programming multicellular behavior with auxin, key genes will be ported from the auxin sensing pathway in the plant Arabidopsis thaliana into the yeast Saccharomyces cerevisiae. Lab yeast is ideal for the characterization of the pathway because it is easy to manipulate and has all the prerequisites needed to process auxin, but is otherwise essentially insensitive to it. Rigorous quantitative characterization of a large variety of synthetic auxin signaling circuits that mimic those found in plants will be combined with mathematical modeling and engineering design tools to build an understanding of the programmability of the pathway. More broadly, the work may lead to new applications in human tissue engineering and new methods for improving important crop species. The project will also support the development of new undergraduate and K-12 educational modules on synthetic biology and cell signaling that use the auxin/yeast system as an example.Technical description: Synthetic cell-cell signaling circuits in yeast will be constructed using genetic parts, pathways, and inspiration from plants. Protein components will be varied across family members to explore how the circuits are tuned. Mathematical modeling and characterization of subtle differences in circuit tunings will elucidate the programmability of the auxin parts. Demonstration of novel cell-cell behaviors will serve as proof of principle of the approach. Specifically, auxin signal processing circuits based on auxin receptors (TIR1, AFB2), transcription factors (ARFs), co-factors (Aux/IAAs) will be constructed. By suitably combining these components, simple signal processing circuits such as filters, repeaters, and pulse generators will be constructed and their dynamic behaviors characterized in yeast. An auxin biosynthesis module, consisting of enzymes that catalyze the production of auxin and controlled by auxin signal processing circuits, will be constructed in yeast to enable a novel mode of cell-cell communication. Auxin sensing circuits will control the production of auxin to achieve signaling functions such as filters and thresholds. The capability to send, receive, and process auxin will enable the investigation of multicellular behaviors such as wave propagation or consensus. The results will not only provide new tools for engineering multicellular systems, but will also provide biological hypotheses about how and why auxin is used so effectively by plants.