Temporal E2F Dynamics and Cell-Fate Decisions in Single Mammalian Cells

描述(由申请人提供)：我们研究的主要目标是建立一个综合的计算/实验框架，以深入了解哺乳动物细胞周期调控的核心信号通路。我们的直接目标是对Myc/Rb/E2F网络的时间动力学有一个定量的了解。这个网络代表了一个理想的系统，用于将关键生物反应的建模与实验和定量分析相结合。首先，网络具有上游激活和下游执行功能，为探索复杂生物网络的设计原理提供了一个明确的背景。其次，鉴于Myc/Rb/E2F通路在控制细胞增殖和细胞命运决策中的关键作用，了解该网络的调制和动力学具有直接的医学意义。因此，该系统提供了一张蓝图，以开发数学模型，帮助揭示关键的监管属性，然后指导实验验证。人们普遍认为，E2F在调节多种细胞反应中起着核心作用，包括增殖、凋亡和分化。实验证据还表明，E2F时间动力学是下游反应的关键线索。然而，对E2F如何控制这种不同的结果缺乏定量的理解。回答这个问题需要对单个哺乳动物细胞在正常和扰动条件下的E2F动力学进行定量分析。为此，该提案的中心目标是开发一套实验和计算工具来量化单细胞中的时间E2F动力学，并了解这些动力学在调节细胞周期进展和凋亡方面的相应含义。拟议的研究是高度多学科的；它利用了参与调查人员的互补专业知识。这项拟议工作的预期成果包括：(1)定量了解对正常细胞生理学和癌症发展具有深远影响的网络的动力学，(2)能够精确调节和量化细胞周期动力学的遗传结构和细胞系，(3)用于Myc/Rb/E2F网络随机分析的基于机械的、受实验约束的数学模型。这些生物学见解以及实验和计算工具可能对系统生物学和细胞生物学的研究界具有广泛的实用价值。

DESCRIPTION (provided by applicant): The overarching goal of our research is to establish an integrated computational/experimental framework for an in-depth view of signaling pathways central to mammalian cell cycle regulation. Our immediate goal is to develop a quantitative understanding of the temporal dynamics of the Myc/Rb/E2F network. This network represents an ideal system for combining modeling with experimental and quantitative analysis of a key biological response. First, the network, with its upstream activation and downstream execution function, provides a well-defined context for exploring design principles of complex biological networks. Second, understanding the modulation and dynamics of this network has direct medical implications given the critical role of Myc/Rb/E2F circuit in controlling cellular proliferation and cell-fate decisions. As such, the system provides a blueprint to develop mathematical models that help revealing critical regulatory properties, which can then guide experimental validation. It is commonly accepted that E2F plays a central role in regulating diverse cellular responses, including proliferation, apoptosis, and differentiation. Experimental evidence also suggests that E2F temporal dynamics serves as a critical cue for downstream responses. However, a quantitative understanding of how E2F controls such diverse outcomes is lacking. Answering this question requires quantitative analysis of E2F dynamics in single mammalian cells under normal and perturbed conditions. To this end, the central goal of the proposal is to develop a set of experimental and computational tools to quantify the temporal E2F dynamics in single cells, and to understand the corresponding implications of these dynamics in regulating cell cycle progression and apoptosis. The proposed research is highly multidisciplinary; it takes advantages of the complementary expertise by the participating investigators. Expected outcomes for the proposed work include: (1) quantitative understanding of the dynamics of a network with profound implications for normal cell physiology and cancer development, (2) genetic constructs and cell lines that enable precise modulation and quantitation of cell cycle dynamics, (3) mechanistically based, experimentally constrained mathematical models for the stochastic analysis of the Myc/Rb/E2F network. These biological insights and experimental and computational tools are likely of broad utility for the research communities of systems biology and cell biology.