Quantitative Characterization & Modeling of sRNA-Mediated Gene Regulation

描述（由申请人提供）：近年来，人们越来越认识到小调控RNA（sRNA）在原核生物和真核生物中协调基因活性的复杂作用。目前对细菌中sRNA介导的基因调控的分子组成和机制有了基本的了解。我们实验室的理论分析表明，sRNA介导的基因调控具有许多独特的功能特征，包括阈值线性响应和抗噪声波动，对涉及sRNA的遗传电路的性质具有有趣的影响。在这里，我们提出了一个联合的实验/计算研究计划，以表征的功能，sRNA在E。大肠杆菌中。在分子水平上，我们将阐明最常见的RyhB类sRNA的sRNA-靶相互作用的序列决定因素，通过表征所选突变体的基因表达，并构建相互作用的生物物理学/生物信息学模型。在“设备”规模，我们将表征预测的sRNA介导的调节，包括不同的目标之间的分层串扰相同的sRNA的各种新的属性。在电路规模上，我们将建构合成电路，并研究常见的电路模体，如级联和开关的性质，并研究它们的时间特性。此外，我们将开发设计靶向和沉默特定宿主基因的反义sRNA的模型。我们希望分子相互作用的知识，导致预测工具，以确定潜在的大量的sRNA的目标和构建额外的层的基因调控网络，涉及sRNA在E。大肠杆菌，而设备和共同的电路图案的知识将有助于理解的特殊作用，sRNA调节器可能在协调更大规模的网络。此外，内源性靶标的沉默可能导致产生多个基因敲低菌株的有效工具，其可用于药物发现研究以及全基因组规模的基因-基因相互作用研究。

DESCRIPTION (provided by applicant): In recent years, there has been a growing appreciation for the complex roles that small regulatory RNA (sRNA) can play in coordinating gene activities in both prokaryotes and eukaryotes. There exists by now a basic understanding of the molecular components and mechanisms involved in sRNA-mediated gene regulation in bacteria. Theoretical analysis by our lab suggest a number of unique functional features for sRNA-mediated gene regulation, including a threshold-linear response and a resistance to noisy fluctuation, with interesting implications on properties of genetic circuits involving sRNA. Here we propose a joint experimental/computational research program to characterize the functions of sRNA in E. coli at multiple scales. At the molecular scale, we will elucidate the sequence determinant of sRNA-target interaction for the most common RyhB-class sRNA by characterizing the gene expression of selected mutants, and constructing biopysical/bioinformatic models of the interaction. At the "device" scale, we will characterize various novel properties predicted for sRNA-mediated regulation, including hierarchical cross talk between different targets of the same sRNA. At the circuit scale, we will construct synthetic circuits and study the properties of common circuit motifs such as the cascade and switches, and investigate their temporal characteristics. In addition we will develop models to design antisense sRNAthat target and silence specific host genes. We expect that the knowledge of the molecular interaction to lead to predictive tools to identify the potentially large number of sRNA targets and construct additional layers of the gene regulatory network involving sRNA in E. coli, while knowledge of the devices and common circuit motifs will shed light towards understanding the special roles that sRNA regulators may play in coordinating larger scale networks. Additionally, the silencing of endogenous target may lead to an effective tool to generate multiple gene knockdown strains which may be used for drug discover studies as well as gene- gene interaction studies at the genome-wide scale.