Mechanisms of Ligand-Dependent Notch Activation

Notch蛋白是高度保守的信号转导系统中的受体，用于交流 相互联系的细胞之间的信息。本项目的总体目标是阐明 信号传导细胞上表达的正则配体激活Notch受体的机制 信号接收单元。具体地说，我们提出了两组互补的研究，将破译两个 配体诱导的Notch信号转导所需的关键事件： 目的1.确定精神炸弹如何诱导配体依赖的Notch信号 我们将利用MIB的模块化性质以及我们在结构和生化方法方面的专业知识来 确定心灵炸弹功能背后的分子逻辑。我们的首要任务将是i)确定 配体-尾结合的结构基础，以及ii)确定不同的MIB结构域如何合作 把泛素转移到这些尾巴上。 目的2.确定配体刺激如何诱导Notch受体金属蛋白酶的裂解 一种主要的Notch配体依赖激活模型假设结合配体的内吞作用 对受体施加机械力，释放自我抑制相互作用，保护 金属蛋白酶裂解位点。这些研究将结合强大的单分子方法和基于细胞的方法 评价Notch信号诱导的机械力模型的可行性。 区分机械转导模型和替代方案，如变构“构象” Switch“模型将极大地提高我们对传送缺口所需关键事件的理解 相邻细胞之间的信号，并将对靶向配体依赖的努力具有重要意义 凹槽信号。此外，还发展了探查缺口信号中机械力影响的方法 转导将在力在生物学中的作用的许多其他领域得到普遍应用。 调查。 总之，追求这些目标将大大促进我们对负责的关键步骤的理解 在相邻小区之间传送陷波信号。此外，他们还将确定新的潜在目标 旨在干扰多种人类癌症中致癌Notch信号的干预措施。

Notch proteins are the receptors in a highly conserved signal transduction system used to communicate information between cells that contact each other. The overarching goal of this Project is to elucidate the mechanism by which canonical ligands expressed on signal-sending cells activate Notch receptors on signal-receiving cells. Specifically, we propose two complementary sets of studies that will decipher two of the critical events that are required for ligand-induced Notch signaling: Aim 1. To determine how Mind bomb induces ligand-dependent Notch signaling We will exploit the modular nature of mib and our expertise in structural and biochemical methods to determine the molecular logic underlying mind bomb function. Our top priorities will be i) to determine the structural basis for ligand-tail binding, and ii) to determine how the different Mib domains cooperate to transfer ubiquitin onto these tails. Aim 2. To determine how ligand stimulation induces metalloprotease cleavage of Notch receptors One leading model for ligand-dependent activation of Notch posits that the endocytosis of bound ligand exerts a mechanical force on the receptor, releasing autoinhibitory interactions that protect the metalloprotease cleavage site. These studies will combine powerful single-molecule approaches and cellbased assays to evaluate the feasibility of the mechanical force model of Notch signal induction. Distinguishing between a mechanotransduction model and alternatives, such as allosteric "conformational switch" models, will substantially advance our understanding of the key events required for conveying Notch signals between adjacent cells, and will have important implications for efforts to target ligand-dependent Notch signaling. Moreover, the methods developed for probing the effect of mechanical force in Notch signal transduction will have general application in many other fields where the role of force in biology is under investigation. Together, pursuit of these aims will substantially advance our understanding of the key steps responsible for conveying Notch signals between adjacent cells. In addition, they will identify new potential targets for interventions designed to interfere with oncogenic Notch signaling in a wide spectrum of human cancers.