Systems-level analysis of the regulation and function of c-Myc

目的：在这个项目中，我们将使用计算和实验技术相结合的方法来表征c-Myc在各种细胞系统中对c-Myc激活信号的反应。通过允许我们研究在小规模相互作用水平上不明显的新特性，这种类型的方法将为操纵电路功能提供新策略，以及对抗c-Myc动力学解除管制的癌症的新方法。材料和方法：为了测量电路元件的动态，我们将主要使用活细胞的长期延时荧光显微镜，其中c-Myc已被荧光标记。我们将使用化学和遗传扰动来改变c-Myc动力学并确定对细胞功能的影响。使用计算模型，我们将把这些数据与细胞结果的测量相结合，以预测响应特定扰动的途径行为。2014财年进展：利用小鼠胚胎成纤维细胞，将编码绿色荧光蛋白GFP的基因插入到两个c-Myc等位基因的内源性位点，我们发现了刺激后c-Myc激活的新的复杂动态。我们用产生响应的一般网络结构来描述这些动态。目前的工作重点是获得对这种调控的更详细的机制理解，以及确定观察到的c-Myc动力学的表型后果。这项工作是与David Levens （LP/CCR）合作进行的。

PURPOSE: In this project, we will use a combination of computational and experimental techniques to characterize c-Myc dynamics in a variety of cell systems in response to c-Myc activating cues. By allowing us to study emergent properties that are not evident at the level of smaller-scale interactions, this type of approach will provide novel strategies for manipulating circuit functions, as well as new ways to combat cancers in which c-Myc dynamics are deregulated. MATERIALS AND METHODS: To measure the dynamics of circuit components, we will primarily use long-term time-lapse fluorescence microscopy of living individual cells in which c-Myc has been fluorescently tagged. We will use chemical and genetic perturbations to alter c-Myc dynamics and determine the effects on cellular functions. Using computational modeling, we will integrate these data with measurements of cellular outcomes to predict pathway behavior in response to specific perturbations. PROGRESS IN FY2014: Using mouse embryonic fibroblasts in which the gene encoding the green fluorescent protein GFP has been inserted into the endogenous loci of both c-Myc alleles, we have identified novel, complex dynamics of c-Myc activation following stimulation. We have characterized these dynamics in terms of general network structure responsible for generating the response. Current work is focused on obtaining a more detailed mechanistic understanding of this regulation, as well as identifying phenotypic comsequences of the observed c-Myc dynamics. This work is being performed in collaboration with David Levens (LP/CCR).