The role of dynamics in defining the limits of normal developmental signaling.

动力学在定义正常发育信号限制中的作用。

• 批准号: 9893735
• 负责人: John G. Albeck
• 金额: $ 5万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-08-01 至 2021-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9893735
• 关键词: Affect; Automobile Driving; Behavior; Cancer Biology; Cell Line; Cell Proliferation; Cell model; Cell physiology; Cells; Clustered Regularly Interspaced Short Palindromic Repeats; Communication; Complex; Complication; Computer Simulation; Data; Defect; Development; Disease; Dose; Drug Targeting; Drug usage; Engineering; Event; Fos-Related Antigens; Frequencies; Gene Expression; Gene Expression Process; Gene Expression Profile; Genes; Genetic Transcription; Genome; Germ-Line Mutation; Heart Abnormalities; Homeostasis; Human; Human Development; Hyperactive behavior; Imaging Techniques; Impaired cognition; In Vitro; Individual; Inherited; Kinetics; Knock-in; Lead; Link; Live Birth; Malignant Neoplasms; Measurement; Measures; Methods; Modeling; Molecular; Monitor; Mutation; Nature; Outcome; Pathologic; Pathology; Pathway interactions; Patients; Pattern; Pharmaceutical Preparations; Pharmacology; Phenotype; Phosphotransferases; Play; Probability; Process; Proteins; Regulation; Reporter; Role; Series; Signal Transduction; Surveys; Syndrome; System; Testing; Therapeutic; Time; Tissues; Translations; Variant; Work; cancer genetics; cancer risk; cell behavior; cell motility; cognitive development; developmental disease; disease-causing mutation; genetic regulatory protein; human disease; imaging platform; inhibitor/antagonist; live cell imaging; mRNA Differential Displays; mathematical model; migration; mutant; preference; programs; repaired; response; single cell analysis; transmission process

Signaling by the Ras/ERK pathway controls cell proliferation, migration, and differentiation. Proper function of this pathway is essential for human development and homeostasis. Sporadic mutations in the pathway play a central role in many cancers, while hereditary mutations cause a series of congenital syndromes, termed RASopathies, which result in impaired cognitive development, cardiac malformations, and increased risk of cancer. A major unanswered question is what differentiates normal from pathological Ras/ERK signaling. It is known that the dynamic pattern of ERK activity – including the strength, frequency, and duration of its activity – are essential to proper signaling. However, the standard methods for measuring ERK activity lack the single-cell precision needed to resolve these essential details. In this project, we will use live-cell imaging, which allows nearly continuous monitoring of thousands of cells simultaneously, to collect data on mutant-driven ERK signaling that is far more accurate and detailed than was previously available. We will focus on the mutations found in the RASopathies, where their role as a single gene driving pathological effects in development is more clearly defined than in cancer, where mutations in many other genes are a complication. We will use our imaging platform to compare for the first time the changes in ERK signaling resulting from disease-causing mutations at the single cell level. Using multiple in vitro systems to replicate cellular processes involved in development, we will determine how these changes modify cell proliferation, migration, and differentiation. We will then dissect the mechanisms underlying these phenotypic changes at the level of gene expression, using a new class of reporters that are integrated directly into the genomes of human cells. At the levels of kinase kinetics, gene expression, and cell behavior, we will quantify how mutant cells respond to multiple Ras pathway inhibitors, which are now being considered as treatments for the RASopathies. This work will have several important outcomes. First, it will reveal the quantitative boundaries of signal behavior that are compatible with normal function, allowing us to understand how Ras pathway mutations lead to disease, and why some mutations are more severe than others. Secondly, it will allow us to make rational choices about which drugs to give to patients with different mutations, so that treatment can be personalized to best normalize each individual's specific signaling patterns. Finally, it will result in a mathematical model of the link between kinase activity and downstream gene expression programs that will allow us to better understand developmental programs and engineer desired cellular responses using existing drugs that target kinase activity.

Ras/ERK 通路的信号传导控制细胞增殖、迁移和 该途径的正常功能对于人类的发育和分化至关重要。 该通路中的零星突变在许多癌症中发挥着核心作用，而 遗传性突变会导致一系列先天性综合征，称为 RASopathies，从而导致 认知发育受损、心脏畸形和癌症风险增加 A 尚未解答的主要问题是正常 Ras/ERK 信号传导与病理性 Ras/ERK 信号传导有何区别。 众所周知，ERK 活动的动态模式——包括强度、频率和 其活动的持续时间——对于正确的信号传递至关重要，但是，标准方法。 测量 ERK 活性缺乏解决这些基本细节所需的单细胞精度。 在这个项目中，我们将使用活细胞成像，它可以近乎连续地监测 同时收集数千个细胞，收集突变驱动的 ERK 信号传导数据 我们将重点关注发现的突变。 在 RASopathies 中，它们作为单个基因驱动病理效应 与癌症相比，发育的定义更为明确，在癌症中，许多其他基因的突变是一个重要因素。 我们将使用我们的成像平台首次比较 ERK 的变化。 在单细胞水平上使用多种致病突变产生的信号传导。 系统来复制参与发育的细胞过程，我们将确定这些如何 然后我们将剖析细胞增殖、迁移和分化的变化。 使用新的方法在基因表达水平上研究这些表型变化的机制 直接整合到人类细胞基因组中的一类产品。 激酶动力学、基因表达和细胞行为，我们将量化突变细胞如何响应 多种 Ras 通路抑制剂，目前被认为是治疗 这项工作将产生几个重要成果。 与正常功能兼容的信号行为的定量边界，使我们能够 了解 Ras 通路突变如何导致疾病，以及为什么某些突变更容易发生 其次，它将使我们能够合理选择给予哪些药物。 针对具有不同突变的患者，以便可以进行个性化治疗以最好地正常化 最后，它将产生一个数学模型。 激酶活性和下游基因表达程序之间的联系将使我们能够 使用以下方法更好地理解程序的发育和设计所需的细胞反应 现有的靶向激酶活性的药物。