CAREER: Tunable Dynamics from Interlinked Feedback Loops in Synthetic and Natural Gene Circuits

摘要智力优势：生物体必须科普各种随时间变化的压力，这是由于营养物质供应、化学压力源和环境变化的波动。遗传上相同的细胞群体可以使用随机的基因表达来多样化它们的反应并对冲这种不确定性。然而，当环境变化缓慢，可以感觉到并作出反应时，就几乎不需要采取多样化的反应。这就提出了一个基本问题，即细胞如何协调这两个潜在矛盾的需求：需要使用表型多样性来预测压力，并采用定义明确的确定性反应。实现这一目标需要可调的基因调控网络，根据环境条件在随机和确定性反应之间切换。最近的理论研究表明，互连的正反馈和负反馈（“混合反馈”）网络可以提供这种类型的可调响应。利用单细胞延时显微镜和数学建模支持的微流体实验，该项目将量化合成和自然混合反馈回路的可调性，并测试细胞如何处理时变环境。这项工作意义重大，因为它将揭示遗传机制，使原本相同的细胞在某些情况下选择随机命运，并在其他情况下表现出确定性反应，使生物体能够科普环境的不确定性。更广泛的影响：研究计划与一个外展计划相结合，该计划专注于社区，高中和本科层次的系统和合成生物学教育：首先，通过与佛蒙特州伯灵顿的ECHO科学中心合作，PI将制作一个特别的夏季展览和讲座，旨在向社区观众传达尖端系统和合成生物学主题。其次，PI将在她的研究小组中接待本科生巴雷特学者，并将为州长工程学院的高中生提供学生演讲指导。第三，PI将引入系统和合成生物学的例子到两个核心工程课程，目前专注于传统的机械和电气工程应用。

ABSTRACTIntellectual Merit: Organisms must cope with a wide range of time-varying stresses, owing to fluctuations in nutrient availability, chemical stressors, and environmental changes. Genetically identical populations of cells can use random, stochastic expression of genes to diversify their response and hedge against this uncertainty. However, when environmental changes are slow and can be sensed and responded to, there is little need for a diversified response. This raises a fundamental question as to how cells reconcile these two potentially contradictory demands: the need to use phenotypic diversity to anticipate stress and also employ well-defined deterministic responses. Achieving this requires gene regulatory networks that are tunable, switching between stochastic and deterministic responses depending on environmental conditions. Recent theoretical studies have suggested that interlinked positive and negative feedback ("mixed feedback") networks, can provide this type of tunable response. Using single-cell time-lapse microscopy and microfluidic experiments supported by mathematical modeling, this project will quantify the tunability of synthetic and natural mixed feedback circuits and will test how cells handle time-varying environments. This work is significant because it will reveal genetic mechanisms that allow otherwise identical cells to choose random fates in some circumstances and exhibit deterministic responses in others, allowing organisms to cope with environmental uncertainty.Broader Impacts: The research plan is integrated with an outreach program focused on systems and synthetic biology education at the community, high school, and undergraduate levels: First, through a partnership with ECHO Science Center in Burlington, Vermont, the PI will produce a special summer exhibit and talk aimed at communicating cutting-edge systems and synthetic biology topics to community audiences. Second, the PI will host undergraduate Barrett Scholars in her research group and will provide mentorship for student presentations to high school students in the Governor's Institute in Engineering. Third, the PI will introduce systems and synthetic biology examples into two core engineering courses that currently focus on traditional mechanical and electrical engineering applications.