Stability and Robustness of Genetic Toggle Switches

由于生物体拥有的基因数量有限，每个基因能够执行的功能也有限，因此一组基因的协调活动对于细胞执行许多复杂的任务是必要的。然而，人们对这种基因网络的组织和进化知之甚少。在这项研究中，将使用体内/体外进化相结合的方法来“繁殖”各种特性的遗传回路。重点将放在最简单、特征最明确的一类遗传电路上，即双基因“触发开关”。将进行分子和进化实验，以研究与该系统的功能和进化有关的一些问题。具体地说，提出的进化方法将被用来产生相对于蜂窝噪声自发切换具有不同程度稳定性的开关。将对进化的调控序列进行分析，以确定与不同程度的电路稳定性相关的特征。还将描述这些开关对突变的稳健性，并将研究系统对噪声的稳定性与其对突变的稳健性之间的可能关系。最后，将通过对进化过程中各个中间阶段的进化区域进行测序来探索开关进化的动力学。这项研究的更广泛影响包括在生物物理学、分子生物学和进化论的界面和前沿培养新一代学生，以及开发实验方案以进化用于各种生物工程和生物医学应用的设计者基因电路。

Because the number of genes an organism possesses is limited and the number of functions each gene can perform is also limited, coordinated activities of a group of genes are necessary for a cell to execute many complex tasks. However, very little is known about the organization and evolution of such gene networks. In this research, a combined in vivo/in vitro evolution approach will be used to "breed" genetic circuits of various properties. The focus will be on the simplest, and the best-characterized class of genetic circuits, the two-gene "toggle switch". Molecular and evolution experiments will be performed to investigate a number of issues concerning the function and evolution of this system. Specifically, the proposed evolutionary method will be used to generate switches with varying degrees of stability with respect to spontaneous switching by cellular noise. The evolved regulatory sequences will be analyzed to identify features that correlate with different degrees of circuit stability. The robustness of these switches to mutation will also be characterized, and possible relationship between the system's stability to noise and its robustness to mutation will be investigated. Finally, the dynamics of the switch evolution will be probed by sequencing the evolved regions at various intermediate stages of the evolution process. Broader impacts of this research include the training of a new generation of students at the interface and the forefront of biophysics, molecular biology, and evolution, and the development of experimental protocols to evolve designer genetic circuits for a variety of bioengineering and biomedical applications.