Regulatory mechanisms of dynamic Wnt signaling in vivo

体内动态Wnt信号传导的调控机制

• 批准号: 8576948
• 负责人: MARCEL WEHRLI
• 金额: $ 29.26万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-30 至 2017-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8576948
• 关键词: APC2 gene; Address; Adult; Affect; Behavior; Binding; Biological Assay; Biological Models; Cell Differentiation process; Cell Maintenance; Cell Nucleus; Cells; Colorectal Cancer; Complex; Defect; Detection; Development; Disease; Dissociation; Drosophila genus; Elements; Event; Family Dasypodidae; Fluorescence; Gene Targeting; Genetic; Goals; Health; Homologous Gene; Human; Kinetics; Knowledge; Ligands; Malignant Neoplasms; Methods; Mission; Modeling; Molecular; Mutation; Nature; Organ; Organism; Osteoporosis; Pathway interactions; Pattern; Phosphorylation; Phosphotransferases; Physiological; Positioning Attribute; Process; Protein Family; Protein Overexpression; Proteins; Protocols documentation; Public Health; Receptor Activation; Regulation; Regulatory Element; Relative (related person); Research; Resolution; Signal Pathway; Signal Transduction; Testing; Time; Tissues; Tumor Suppressor Proteins; Work; arm; base; casein kinase; in vitro Assay; in vivo; innovation; kinase inhibitor; novel; public health relevance; receptor; regional difference; research study; response; scaffold; stem cells; tool

DESCRIPTION (provided by applicant): Wnt/??catenin signaling is critically important to development and disease. How this signaling pathway functions normally in vivo remains unclear. At least a dozen different models have been postulated to explain ?-catenin regulation; most are based on studies requiring protein overexpression in highly artificial in vitro assays. Many contradictions may be resolved by a unifying model built on the hypothesis that Wnt signaling in different contexts always uses the same 'core' Wnt pathway, which is fine-tuned by context-specific 'accessory modules'. A new paradigm for Wnt signaling will be developed by determining how components of the core pathway are assembled and regulated by different accessory modules, and what constitutes an accessory module. The goal is to identify these mechanisms by generating a dynamic model of the pathway (Aim 1). The hypothesis is that the core pathway responds to Wnt signal with faster kinetics than accessory mechanisms. An innovative assay has been established in the Drosophila model system to detect interacting Wnt pathway components at molecular resolution and in real time. This has already revealed several highly significant findings, namely the detection, for the first time in cells or in vivo, of the ky regulator in the pathway, termed the 'destruction complex'. This complex is deemed part of the core pathway in contrast to an accessory mechanism represented by the interaction between the receptor Arrow and the destruction complex scaffold Axin, based work by the PI. The molecular mechanism of this accessory module will be probed by functional tests of model-based predictions (Aim 2). In addition, the hypothesis that a set of mutations in Arrow selectively affects Arrow's function in the same accessory module will be tested. Similarities to defects associated with osteoporosis caused by mutations in a human homologue of Arrow, LRP5, raise the possibility that this disorder may be caused by defects in a Wnt accessory mechanism, rather than in the core mechanism, as assumed. Aim 3 tests the hypothesis that the Wnt receptor may initiate signaling by dissociation of the receptor subunits Arrow and Frizzled, contrary to current models for the assembly and activation of this receptor complex. Whether the strong negative regulation of the Frizzled2-Arrow interaction is part of the core pathway or represents a previously unrecognized mechanism for modulating Wnt-dependent processes in a context- specific manner will also be determined. Taken together, the analyses proposed here will significantly advance the understanding of Wnt signaling by generating a dynamic model of normal signaling in vivo at unprecedented resolution. The emerging model is expected to provide a clear distinction between universal regulation by the core pathway and tissue-specific modulation of the Wnt signal by accessory mechanisms. Such a paradigm shift will reveal, in many instances, that Wnt-associated diseases are defective in tissue-specific accessory mechanisms and such a distinction will greatly facilitate the development of targeted therapies.

描述（由申请人提供）：Wnt/?? Catenin信号对发育和疾病至关重要。该信号通路通常在体内运作的功能尚不清楚。假定至少有十几个不同的模型来解释 - 蛋白酶调节；大多数基于需要在高度人为的体外测定中过表达的研究。许多矛盾可以通过构建的统一模型来解决以下假设：在不同上下文中Wnt信号始终使用相同的“核心” Wnt途径，该途径由特定于上下文特定的“附件模块”进行了微调。通过确定如何通过不同的附件模块组装和调节核心途径的组件以及什么构成附件模块来开发新的Wnt信号范式。目的是通过产生途径的动态模型来识别这些机制（AIM 1）。假设是，核心途径对Wnt信号的响应比附件机制更快。在果蝇模型系统中已经建立了一种创新的测定，以在分子分辨率和实时检测相互作用的Wnt途径成分。这已经揭示了几个高度显着的发现，即第一次在细胞或体内检测到途径中的ky调节剂，称为“破坏复合物”。与由受体箭头与破坏复合物支架Axin之间的相互作用表示的辅助机理相比，该复合物被认为是核心途径的一部分，这是PI的基于PI的工作。该附件模块的分子机制将通过基于模型的预测的功能测试来探测（AIM 2）。另外，有选择性地中的一组突变的假设 将测试在同一附件模块中影响箭头的功能。与箭头同源物LRP5中突变引起的与骨质疏松症相关的缺陷相似，这增加了这种疾病可能是由Wnt附属机制中的缺陷引起的，而不是在核心机制中引起的。 AIM 3检验了以下假设：Wnt受体可以通过解离受体亚基箭头和卷曲来启动信号传导，这与当前用于组装和激活该受体复合物的模型相反。卷曲2-箭头相互作用的强负调节是否是核心途径的一部分，还是代表以上语境调节Wnt依赖性过程的先前未识别的机制 - 也将被确定。 综上所述，这里提出的分析将通过在前所未有的分辨率下生成正常信号传导的动态模型，从而显着提高对Wnt信号的理解。预计新兴模型将通过核心途径与辅助机制对WNT信号的组织特异性调节之间的通用调节之间有明显的区别。在许多情况下，这种范式转移将表明与Wnt相关的疾病在组织特异性的辅助机制中有缺陷，这种区别将极大地促进靶向疗法的发展。