Modeling Feedback Regulation of Cell Signaling

描述(由申请人提供)： 摘要细胞行为受环境信号的调节，包括营养物质、渗透压力、激素和神经递质。这些信号在剂量、持续时间和方向性上可能有很大的不同。这一建议旨在(I)在时间和空间上定量地测量酵母信息素反应途径，(Ii)设计描述观察到的行为的计算模型，以及(Iii)通过进一步的实验来测试每个模型的有效性。最重要的假设是，为研究动态系统而开发的数学技术可以解释通路组件如何解释和翻译细胞外的空间线索，以唤起细胞内的适当反应。重点将放在调节酵母中信息素途径的时间依赖行为的蛋白质上，特别是这些调节器如何对适当的信号转导做出贡献。模型将在分子和细胞水平上进行实验测试，其中包括使用创新的梯度流室。有三个特定的目标，集中在三种不同的蛋白质在该途径的三个不同的步骤中发挥作用。所有这三种蛋白质都是梯度感应活动所必需的。目的1将研究支架蛋白Ste5的时间依赖性调节。这一目标将检验这样的假设，即Ste5通过在MAP激酶Fus3完全激活所需的两个磷酸化事件之间施加延迟来调制信号。目的2将研究RGS蛋白Sst2对时间和空间的依赖调节。这一目标将检验这样一种假设，即Sst2作为支架蛋白发挥作用，协调G蛋白的激活(通过受体)和失活(G蛋白的失活)，从而将细胞定向到梯度刺激。目的3将研究信息素蛋白水解酶bar1对时间和空间的依赖调节。这一目标将检验Bar1重塑梯度的假设，从而协调两个细胞对相同刺激的反应。公共卫生相关性：项目叙述正确的细胞功能需要有能力检测荷尔蒙、神经递质和药物并对其做出适当反应。负责信号传递的细胞机制从人类到酵母都是保守的。该项目使用多学科方法，包括生物实验、微型制造的生长室和计算机模拟，以确定细胞如何解释刺激的方向和强度。更广泛的目标是了解空间和时间信息在细胞信号中的作用，并最终预测哪些药物治疗将在对抗人类疾病方面最有效。

DESCRIPTION (provided by applicant): SUMMARY Cellular behavior is modulated by environmental signals including nutrients, osmotic stress, hormones and neurotransmitters. These signals can vary considerably in dose, duration, and directionality. This proposal seeks to (i) quantitatively measure the yeast pheromone response pathway, both in time and space, (ii) devise computational models that describe the observed behaviors, and (iii) test the validity of each model through further experimentation. The over-arching hypothesis is that mathematical techniques developed for studying dynamical systems can explain how pathway components interpret and translate spatial cues outside the cell to evoke appropriate responses inside the cell. The focus will be on proteins that modulate the time-dependent behaviors of the pheromone pathway in yeast, and in particular how these modulators contribute to proper signal transduction. Models will be tested experimentally at the molecular and cellular level, and include the use of an innovative gradient flow chamber. There are three specific aims, focused on three different proteins acting at three distinct steps in the pathway. All three proteins are required for gradient-sensing activity. Aim 1 will investigate time-dependent regulation by the scaffold protein Ste5. This aim will test the hypothesis that Ste5 modulates signaling by imposing a delay between each of two phosphorylation events needed for full activation of a MAP kinase Fus3. Aim 2 will investigate time- and space-dependent regulation by the RGS protein Sst2. This aim will test the hypothesis that Sst2 functions as a scaffold protein that coordinates G protein activation (by the receptor) and inactivation (of the G protein), and thereby orients the cell towards a gradient stimulus. Aim 3 will investigate time- and space-dependent regulation by the pheromone protease Bar1. This aim will test the hypothesis that Bar1 remodels the gradient, and thereby coordinates the behavior of two cells responding to the same stimulus. PUBLIC HEALTH RELEVANCE: PROJECT NARRATIVE Proper cell function requires the ability to detect and respond appropriately to hormones, neurotransmitters and drugs. The cellular machinery responsible for signal transmission is conserved from humans to yeast. This project uses multi-disciplinary approaches, including biological experiments, microfabricated growth chambers and computer simulations, to establish how the direction and strength of a stimulus are interpreted by the cell. The broader objective is to understand the role of spatial and temporal information in cell signaling, and eventually predict what drug treatments will be most effective in combating human diseases.