Engineered Selective Pressure: Challenges and Opportunities

活细胞中复杂遗传电路的正向工程的快速进展证明，合成生物学这门年轻的学科有可能改变现代生物技术。对生命系统的预测性理解是生物工程设计操作和医学知情干预的先决条件。这种理解需要定量测量、数学分析和理论抽象。强大的测量技术和计算能力的出现使生物学能够推动下一次科学革命。合成生物学提供了一个天然的平台，用于使用来自工程遗传电路的定量数据开发和测试模型和一般网络原理。合成生物学的主要基本问题之一是合成电路倾向于进化并消除其预期功能。因此，重要的是要开发一种定量方法，使遗传网络更强大和稳定，这也将使它们在治疗应用中更有用。该项目的总体目标是探索挑战，并通过尖端实验和计算工具的结合，利用合成生物学中出现的不可避免的选择压力带来的机会。该项目的成功完成将导致在进化背景下理解合成电路和宿主基因组之间的相互作用的重大进展。 该项目将为新一代定量生物学家的跨学科培训提供充足的机会。此外，为了扩大该项目在学术界以外的影响，参与者将扩大一个非常成功的小学科学项目，该项目旨在促进合作伙伴小学的研究人员和教师之间的合作，以改善实践科学教育。为了实现该项目的总体目标，最近开发的“同步裂解电路”将被用作主要焦点。这种合成基因回路的功能是在阈值种群密度下触发大约90%的细菌死亡，留下剩余的10%的细胞生长回到阈值并重新开始循环。这产生了一种独特的表型，其特征在于可以使用微型和台式恒化器在宽范围的长度尺度上监测的群体循环。它还引入了强大的选择压力，可以导致快速进化。研究人员将开发定量测量技术和计算建模，这将导致电路主机进化过程的定量统计特征。沿着实验的时间进程对宿主基因组的电路和突变部分进行测序，将用于重建系统所采取的进化路径。数学建模将指导实验并帮助提取表征同步裂解回路的进化动力学的关键参数。利用所获得的知识，研究人员将设计一个“裂解回路稳定器”模块，杀死失去裂解效率的突变细菌，并探索如何使用裂解回路作为合成回路定向进化的工具。该奖项由分子和细胞生物科学（MCB）的系统和合成生物学（SSB）项目共同资助。生物科学理事会的生物技术和生物化学工程（BBE）计划以及工程理事会化学，生物工程，环境和运输系统（CBET）的生物技术和生物化学工程（BBE）计划。

Rapid advances in the forward engineering of complex genetic circuits in living cells proved that the young discipline of Synthetic Biology has the potential to transform modern biotechnology. A predictive understanding of living systems is a prerequisite for designed manipulation in bioengineering and informed intervention in medicine. Such an understanding requires quantitative measurements, mathematical analysis, and theoretical abstraction. The advent of powerful measurement technologies and computing capacity has positioned biology to drive the next scientific revolution. Synthetic Biology provides a natural platform for the development and testing of models and general network principles using quantitative data from engineered genetic circuits. One of the major fundamental problems of Synthetic Biology is the propensity of synthetic circuits to evolve and obliterate their intended functionality. Thus, it is important to develop a quantitative approach to make genetic networks more robust and stable which also will make them more useful in therapeutic applications. The overall goal of this project to explore the challenges and exploit the opportunities that accompany the inevitable selective pressure that arises in synthetic biology through a combination of cutting-edge experimental and computational tools. Successful accomplishment of this project will lead to significant advances in understanding the interaction between synthetic circuits and host genome in the evolutionary context. This project will provide ample opportunities for cross-disciplinary training of the new generation of quantitative biologists. Furthermore, to broaden the impact of this project beyond academia, its participants will expand a highly successful elementary school science program that that foster collaboration between researchers and teachers at partner elementary schools to improve hands-on science education.To reach the overall goal of the project, the recently developed "synchronized lysis circuit" will be used as the major focus. This synthetic gene circuit functions by triggering death of around 90% of the bacteria at a threshold population density, leaving the remaining 10% of the cells to grow back to threshold and restart the cycle. This generates a distinct phenotype characterized by population cycling that can be monitored across a wide range of length scales using micro- and bench-top chemostats. It also introduces a strong selective pressure that can lead to rapid evolution. The investigators will develop quantitative measurement technology and computational modeling that will lead to a quantitative statistical characterization of the circuit-host evolution process. Sequencing of the circuit and mutated sections of the host genome along the time course of the experiments will be used to reconstruct the evolutionary path the systems have taken. Mathematical modeling will guide experiments and help extract key parameters characterizing evolutionary dynamics of the synchronized lysis circuit. Using the gained knowledge, the researchers will engineer a "lysis circuit stabilizer" module that kills mutant bacteria losing lysis efficiency, and explore ways to use lysis circuit as a tool in directed evolution of synthetic circuits.This award was co-funded by the Systems and Synthetic Biology (SSB) program in the Molecular and Cellular Biosciences (MCB) Division in the Biological Sciences Directorate and the Biotechnology and Biochemical Engineering (BBE) program of the Division of Chemical, Bioengineering, Environmental and Transport Systems (CBET) in the Engineering Directorate.