Axin-induced Wnt signaling in mouse embryogenesis and Wnt-related cancers

小鼠胚胎发生和 Wnt 相关癌症中轴蛋白诱导的 Wnt 信号传导

• 批准号: 8527327
• 负责人: Angela R Parrish
• 金额: $ 4.92万
• 依托单位: SLOAN-KETTERING INST CAN RESEARCH
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-06-01 至 2016-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8527327
• 关键词: Address; Adult; Alleles; Area; Attenuated; Axin protein; Binding; Biochemical; Biochemistry; Breast Cancer Model; Candidate Disease Gene; Cells; Colon; Colorectal Cancer; Complex; Congenital Abnormality; Cultured Cells; Data; Development; ES Cell Line; Embryo; Embryonic Development; Employee Strikes; Frequencies; Gastrointestinal tract structure; Gene Transfer Techniques; Genetic; Genetic Models for Cancer; Goals; Head; Heterozygote; Homozygote; Incidence; Intestines; Investigation; Lead; Ligands; Location; Maintenance; Malignant Neoplasms; Mammary gland; Mass Spectrum Analysis; Membrane; Methods; Modeling; Mouse Mammary Tumor Virus; Mus; Mutation; N-terminal; Nature; Neoplasm Metastasis; Organ; Outcome; Pathway interactions; Peptides; Pharmaceutical Preparations; Play; Population; Post-Translational Protein Processing; Primitive Streaks; Property; Proteins; Regulation; Reporting; Research; Role; Severities; Signal Pathway; Signal Transduction; Small Intestines; Staging; Stem cells; Testing; Therapeutic; Time; Tissues; Transgenic Mice; Transgenic Organisms; Up-Regulation; Western Blotting; base; body system; cancer therapy; cancer type; cellular imaging; embryonic stem cell; human disease; mammary epithelium; mutant; overexpression; prevent; protein complex; protein expression; public health relevance; research study; response; small hairpin RNA; stem cell niche; stem cell population; tumor; tumorigenesis

DESCRIPTION (provided by applicant): The canonical Wnt signaling pathway is required for the development of most organ systems and for the maintenance of stem cell populations in the adult. Aberrant signaling can drive the formation of tumors in several tissues. Axin proteins are considered negative regulators of the Wnt pathway due to their function in the destruction complex, which prevents signaling in the absence of Wnt ligand. Surprisingly, stabilization of Axin proteins by either genetic or pharmacological methods leads to an increase in signaling in the posterior of the e8.5 mouse embryo, whereas Wnt signaling is decreased in the head, suggesting that Wnt signaling is regulated in a tissue-specific manner. The goals of the proposed research are to understand how the positive and negative roles of Axin1 and Axin2 are tissue-specifically regulated during development. Specifically, the aims of this research are the characterization of the mechanisms controlling tissue-specific, Axin-stimulated Wnt signaling in the primitive streak, identification of the biochemical basis of Axin-stimulated Wnt signaling, and examination of whether stabilized Axin2 promotes tumorigenesis in the intestine and mammary gland. Using genetics and cell-based approaches, the proteins that establish Axin-stimulated Wnt signaling in the primitive streak will be identified. The role of high levels of Wnt ligand, the expression of core signaling components, the genetic interaction of potential modulators of the pathway, and intersecting signaling pathways expressed at this time and location in the mouse embryo will be examined for their contribution to Axin-stimulated Wnt signaling. These data will be used to recapitulate Axin-induced Wnt signaling in ES cells. To facilitate biochemical analysis of Axin-containing complexes and imaging of the cellular localization of Axin- containing complexes, a mouse that expresses a conditional tagged, stabilized Axin1 from the Rosa26 locus will be generated. Immunopurification of Axin-containing complexes from ES cells or tissues engaged in both Axin-inhibited and Axin-induced Wnt signaling will be assessed by Western blot to compare the binding of core pathway components. Mass spectrometry will be performed to identify whether unique proteins or post- translational modifications lead to different signaling outcomes in response to stabilized Axin. Finally, tissues in adults will be evaluated for increased Wnt signaling in response to stabilized Axin2, concentrating on populations maintained by Wnt-controlled stem cell niches. Additionally, the effect of stabilized Axin2 on two models of Wnt-related cancer, the Apcmin model of colorectal cancer and the MMTV-Wnt1 model of breast cancer, will be assessed, focusing on the incidence, onset, or invasiveness of the tumors that arise to determine whether some tumors increase in severity due to stabilized Axin proteins, addressing the utility of Axin-stabilizing drugs in cancer treatment.

描述（由申请人提供）：大多数器官系统的开发以及成人的干细胞种群的发展是必需的。异常信号传导可以驱动几个组织中肿瘤的形成。由于轴蛋白在破坏复合物中的功能，因此被认为是Wnt途径的负调节剂，该蛋白在没有Wnt配体的情况下阻止了信号传导。令人惊讶的是，通过遗传或药理方法对轴蛋白的稳定化导致E8.5小鼠胚胎后端的信号传导增加，而头部的WNT信号传导降低，这表明Wnt信号在组织特异性方式中受到调节。拟议的研究的目标是了解AXIN1和AXIN2的正和负作用如何在发育过程中特异性调节。具体而言，这项研究的目的是在原始条纹中控制组织特异性，轴刺激的Wnt信号的机制的表征，鉴定Wnt的Wnt信号的生化基础的鉴定，以及研究稳定的AXIN2是否促进了直肠和乳腺中的肿瘤发生。使用遗传学和基于细胞的方法，将确定在原始条纹中建立轴蛋白刺激的Wnt信号传导的蛋白质。高水平的Wnt的作用 配体，核心信号成分的表达，途径的电势调节剂的遗传相互作用以及在此时间和位置在小鼠胚胎中表达的相交信号通路的遗传相互作用，将检查其对Axin刺激的Wnt信号传导的贡献。这些数据将用于概括ES细胞中AXIN诱导的Wnt信号传导。为了促进含轴蛋白复合物的生化分析和含有轴突的复合物的细胞定位的成像，将生成从ROSA26基因座表达有条件标记的，稳定的Axin1的小鼠。将通过Western blot评估含Axin抑制和轴诱导的Wnt信号的ES细胞或组织的ES细胞或组织的含Axin的复合物的免疫功能，以比较核心途径成分的结合。将执行质谱法，以确定独特的蛋白质或转化后修饰是否导致响应稳定轴的不同信号传导结果。最后，组织 在稳定的AXIN2响应的响应中，将评估成年人的Wnt信号，并集中在Wnt控制的干细胞壁ni维持的种群上。此外，将评估稳定AXIN2对WNT相关癌症模型的影响，结直肠癌的APCMIN模型和MMTV-WNT1乳腺癌模型，将评估，重点关注肿瘤的发病或侵入性，这些肿瘤的发病率或侵入性在确定由于稳定的轴蛋白质而导致的肿瘤是否会增加Axpin cancer cancel的稳定性。