Mechanisms of Ligand-Dependent Notch Activation

Notch蛋白是一个高度保守的信号转导系统中的受体， 细胞之间相互接触的信息。本项目的总体目标是阐明 信号发送细胞上表达的典型配体激活Notch受体的机制 信号接收细胞具体来说，我们提出了两套互补的研究，将破译两个 配体诱导的Notch信号传导所需的关键事件： 目标1.为了确定思维炸弹如何诱导配体依赖性Notch信号传导， 我们将利用mib的模块化特性以及我们在结构和生物化学方法方面的专业知识， 确定思维炸弹功能背后的分子逻辑我们的首要任务是i）确定 ii）确定不同的Mib结构域如何合作， 将泛素转移到这些尾巴上。 目标2.确定配体刺激如何诱导Notch受体的金属蛋白酶切割 Notch的配体依赖性激活的一个主要模型假定结合配体的内吞作用 对受体施加机械力，释放保护受体的自抑制相互作用。 金属蛋白酶切割位点。这些研究将结合联合收割机强大的单分子方法和基于细胞的 分析以评估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模型，如变构“构象”模型， 开关”模型，将大大提高我们的理解所需的关键事件，为输送缺口 信号之间的相邻细胞，并将有重要的影响，努力靶向配体依赖性 陷波信号。此外，还提出了探测机械力对Notch信号影响的方法， 在生物学中力的作用受到限制的许多其他领域，转导将具有普遍的应用。 调查 总之，追求这些目标将大大促进我们对负责实现这些目标的关键步骤的理解。 在相邻小区之间传送Notch信号。此外，他们还将确定新的潜在目标， 这些干预措施旨在干扰广谱人类癌症中的致癌Notch信号传导。

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.