Probing delays and memory in gene activation using a gene oscilloscope

这项由生物技术、生化和生物质工程项目颁发的NSF奖支持开发和应用一种新的工具--“基因示波器”，以在单细胞水平上测量基因激活的动力学。基因示波器由酵母菌株组成，酵母菌株含有可观察的、动态可控的输入转录因子和作为可观察输出的荧光报告器。输入/输出行为是从微流控设备中生长的单个细胞的电影中提取的。它可以用来研究基因调控网络中简单组件的动态。这项工作的智力价值是使用简单的集总动力学模型来量化启动子激活过程中的延迟和记忆，该模型将模型参数与启动子结构联系起来。这项工作将通过比较促进剂和激活剂的不同特征如何影响动力学来产生机械论的见解。此外，它还将通报和更新大规模的基因网络电子模拟工作，旨在了解网络组件的变化如何导致定性不同的动态、异常功能和疾病状态。更广泛的影响：除了对转录动力学的基本见解外，这项工作还应该促进基因示波器的概念，并潜在地使合成生物学家和代谢工程师能够表征转录设备‘部分’的动力学特性。除了传统的研究机会外，这个项目还将为PI实验室的毕业生和本科生提供机会，基因示波器是本科化学-生物工程实验室内一系列研究项目的基础。其目标是让下一代生物过程工程师和生物学家了解单细胞分析在生物技术中的重要性，生物技术传统上将培养物视为整体。

This NSF award by the Biotechnology, Biochemical and Biomass Engineering program supports the development and application of a novel tool, the 'gene oscilloscope', to measure the kinetics of gene activation at the single cell level. The 'gene oscilloscope' consists of a yeast strain containing observable, dynamically controllable input transcription factor and a fluorescent reporter as the observable output. Input/output behavior is extracted from movies of single cells grown in microfluidic devices. It can be used to examine the dynamics of simple components in a gene regulatory network. The intellectual merit of this work is to quantify delays and memory in the activation of a promoter using simple lumped kinetic models that connect model parameters to promoter architecture. This work will yield mechanistic insights by comparing how different features of promoters and activators affect kinetics. In addition, it will inform and update large scale in silico modeling efforts of gene networks aimed at understanding how changes in network components lead to qualitatively different dynamics, aberrant function, and disease states. Broader impacts: Apart from basic insights into transcriptional dynamics, this work should further the concept of a gene oscilloscope and potentially enable synthetic biologists and metabolic engineers to characterize the dynamical properties of transcriptional device 'parts'. Apart from traditional research opportunities this project has and will provide both graduates and undergraduates in the PI's lab, the gene oscilloscope serves as a foundation for a set of research projects within the undergraduate Chemical-Biological Engineering laboratory. The goal is to expose the next generation of bioprocess engineers and biologists to the importance of single cell analysis in biotechnology, which traditionally has treated cultures as monolithic.