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

目得：在这个项目中，我们将使用计算和实验技术相结合的特点c-Myc的动态在各种细胞系统中的c-Myc激活线索。通过允许我们研究在小规模相互作用水平上不明显的涌现特性，这种方法将提供操纵电路功能的新策略，以及对抗c-Myc动力学失调的癌症的新方法。材料和方法：为了测量电路组件的动态，我们将主要使用活的单个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。使用计算建模，我们将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通路行为。2015财年进展：使用小鼠胚胎成纤维细胞，其中编码绿色荧光蛋白GFP的基因已被插入到两个c-Myc等位基因的内源性基因座中，我们已经确定了新的，复杂的c-Myc激活动力学刺激后。我们的特点是这些动态的一般网络结构负责产生的响应。目前的工作重点是获得更详细的机械理解这种调节，以及确定所观察到的c-Myc动态的表型后果。我们还扩展了这项工作，以研究c-Myc网络和p53网络对各种刺激的共同调节。这项工作是与大卫莱文斯（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 FY2015: 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. We have also extended this work to look at co-regulation of the c-Myc network and the p53 network in response to various stimuli. This work is being performed in collaboration with David Levens (LP/CCR).