RECEPTOR TYROSINE KINASE SIGNALING IN THE LIVER

描述：(改编自申请者的摘要)一个重要的成长群体 因子和激素通过受体酪氨酸激酶作用于肝细胞 (RTK)，如表皮生长因子(EGF)受体，肝细胞生长 因子/分散因子(HGF/SF)受体和胰岛素受体。尽管有一个 我们对酪氨酸激酶分子机制的了解呈爆炸性增长 在过去的十年中，RTK信号通路的调节 细胞水平仍然知之甚少。RTK激活多个信令 在早期接头/靶蛋白水平上经常重叠的通路 丝裂原活化蛋白激酶(MAPK)的激活、刺激， 以及由它们的刺激触发的转录事件。最近的文献数据， Kholodenko博士的研究小组的研究结果也支持这一观点 信号的特异性由信号的时序、持续时间和幅度产生 激活不同的组件进程。然而，缺乏一种 RTK信号转导途径调控的定量和综合描述 妨碍了我们对复杂的特殊变化的原因的理解 信号响应及其对下游通路的直接影响。 这项提议旨在通过定量的动力学监测和 RTK的磷酸化和活化水平的计算分析 EGF、HGF/SF和胰岛素刺激肝细胞途径的中间产物。 Kholodenko博士将使用生长因子的动力学和控制分析 完整肝细胞中识别主要分子和动力学的信号 控制RTK信号通路的因素。具体目标是：(1) 分析控制瞬变和持续响应模式的因素 EGF刺激新分离的细胞内多种信号蛋白的靶向 肝细胞；(2)分析早期信号的动力学和调控 EGF刺激的MAPK级联反应；(3)扩大实验范围 和计算分析，以调查对其他RTK中介的响应 信号，特别是HGF/SF和胰岛素，单独或联合使用。

DESCRIPTION: (adapted from applicant's abstract) An important group of growth factors and hormones acts on liver cells through receptor tyrosine kinases (RTKs) such as the epidermal growth factor (EGF) receptor, hepatocyte growth factor/scatter factor (HGF/SF) receptor and insulin receptor. Despite an explosive growth of our knowledge of molecular mechanisms of tyrosine kinase signaling during the past decade, the regulation of RTK pathways at the cellular level remains poorly understood. RTKs activate multiple signaling pathways that often overlap at the level of early adapter/target protein activation, stimulation of mitogen activated protein kinase (MAPK) cascades, and transcriptional events triggered by their stimuli. Recent literature data, also supported by findings from Dr. Kholodenko's group, suggest that specificity of signaling is generated by the timing, duration and amplitude of activation of the different component processes. However, the lack of a quantitative and integrative description of the regulation of RTK pathways hampers our understanding of the causes of particular alterations in complex signaling responses and their immediate consequences for downstream pathways. This proposal aims to fill this gap by quantitative kinetic monitoring and computational analysis of the levels of phosphorylation and activation of RTK pathway intermediates in hepatocytes stimulated with EGF, HGF/SF and insulin. Dr. Kholodenko will employ kinetic and control analyses of growth factor signaling in intact hepatocytes to identify the principal molecular and kinetic factors controlling RTK signaling pathways. The Specific Aims are: (1) To analyze the factors controlling transient and sustained response patterns in multiple signaling proteins targeted by EGF stimulation in freshly isolated hepatocytes; (2) To analyze the kinetics and control of early signaling responses of the EGF-stimulated MAPK cascade; (3) To extend the experimental and computational analyses to investigate the responses to other RTK-mediated signals, specifically HGF/SF and insulin, singly or in combination.