An Integrative Analysis of MAPK Signaling in Budding Yeast

描述(申请人提供)：调控两个单倍体细胞融合的交配反应通路或对外源添加的信息素的反应而激活的交配反应通路一直是典型的信号转导通路，其分析导致对丝裂原活化蛋白激酶(MAPK)级联的结构和功能的多个重要见解。大量的遗传和分子生物学工具已经开发出来，继续使这一途径成为一个非常有吸引力的系统，以逐步完善对来自细胞微环境的信号如何转化为基因转录和细胞形态的明确变化的理解。然而，我们对这一重要途径的了解还远未完全，这在很大程度上是因为缺乏实验工具来分析该途径在更多生理背景下的活性，如梯度感应和趋化性，以及饱和信息素梯度(默认反应)中的顺序投影形成，而不是对空间均一信息素浓度的相对短期反应。在这一应用中，我们建议使用新颖的实验设计和微制造器件在梯度感应和默认响应表型的背景下研究交配途径。我们建议研究途径活性的各种修饰物的作用，特别是支架蛋白和各种负调控因子。我们还将探索信息素途径和其他MAPK途径之间的串扰机制，也依赖于新的实验技术。这些定量实验分析的结果将被整合到芽殖酵母中激活的MAPK通路的综合计算模型中。MAPK通路跨物种的保守性以及这些通路在各种人类病理中的特殊重要性使这项研究对我们理解人类健康和疾病具有特别重要的意义。与公共健康相关：这项应用集中在信号转导的系统方法上，这是典型的和研究最多的途径之一，酵母中的交配途径。尽管表面上看起来很简单，但这一途径调节了过多的细胞反应，从细胞形态的变化和定向细胞生长，到周期性的形态发生和数百个基因的表达。我们建议使用一系列新的方法来研究该途径如何控制这一不同的表型反应，并建立对该途径功能的定量理解。我们预计，我们的发现将对酵母和高等生物中其他途径的功能提供信息。

DESCRIPTION (provided by applicant): The mating response pathway regulating fusion of two haploid cells of the budding yeast S. cerevisiae or activated in response to exogenously added pheromones has been a prototypical signal transduction pathway, whose analysis led to multiple important insights into the structure and function of mitogen-activated protein kinase (MAPK) cascades. A vast collection of genetic and molecular biology tools have been developed that continue to make this pathway a very attractive system for developing a progressively refined understanding of how the signals from the cell microenvironment are converted into well defined changes in gene transcription and cell morphology. However, our understanding of this important pathway is far from complete, in large part due to the lack of experimental tools for analysis of the pathway activity in more physiological contexts of gradient sensing and chemotropism, and the sequential projection formation in saturating pheromone gradients (the default response), as opposed to the relatively short term responses to spatially homogeneous pheromone concentrations. In this application, we propose to study the mating pathway within the contexts of gradient sensing and default response phenotypes using novel experimental designs and microfabricated devices. We propose to investigate the role of various modifiers of pathway activity, with a particular focus on the scaffold protein and various negative regulators. We will also explore the mechanisms of cross-talk between the pheromone pathway and other MAPK pathways, also relying on the novel experimental technology. The results of these quantitative experimental analyses will be integrated into a comprehensive computational model of the MAPK pathways activated in budding yeast. Conservation of MAPK pathways across species and the particular importance of these pathways in various human pathologies make this research especially significant for our understanding of human health and disease. PUBLIC HEALTH RELEVANCE: This application is focused on the systems approach to signal transduction in one of the prototypical and most studied pathways, the mating pathway in yeast. In spite of its apparent simplicity, this pathway regulates a plethora of cell responses, from changes in cell morphology and directed cell growth, to periodic morphogenesis and expression of hundreds of genes. We propose using a set of novel approaches to investigate the way the pathway can control this diverse set of phenotypic responses and build a quantitative understanding of the pathway function. We anticipate that our findings will be informative about the function of other pathways in yeast and higher organisms.