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）：经典Wnt信号通路是大多数器官系统发育和维持成人干细胞群所必需的。异常信号传导可以驱动几种组织中肿瘤的形成。轴蛋白由于其在破坏复合物中的功能而被认为是Wnt途径的负调节剂，所述破坏复合物在Wnt配体不存在的情况下阻止信号传导。令人惊讶的是，通过遗传或药理学方法稳定Axin蛋白导致e8.5小鼠胚胎后部信号传导增加，而头部Wnt信号传导减少，表明Wnt信号传导以组织特异性方式调节。这项研究的目的是了解Axin 1和Axin 2的积极和消极作用在发育过程中是如何组织特异性调节的。具体而言，本研究的目的是表征控制组织特异性Axin刺激的原始条纹中Wnt信号传导的机制，鉴定Axin刺激的Wnt信号传导的生化基础，并检查稳定的Axin 2是否促进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Axin刺激的Wnt信号传导的蛋白质。高水平Wnt的作用 配体的表达，核心信号传导组分的表达，途径的潜在调节剂的遗传相互作用，以及在小鼠胚胎中此时和位置表达的交叉信号传导途径将被检查其对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Axin复合物的生物化学分析和含Axin复合物的细胞定位的成像，将产生从Rosa 26基因座表达条件性标记的、稳定的Axinl的小鼠。将通过Western印迹评估来自参与Axin抑制和Axin诱导的Wnt信号传导的ES细胞或组织的含Axin复合物的免疫纯化，以比较核心途径组分的结合。将进行质谱分析以确定独特的蛋白质或翻译后修饰是否会导致响应稳定的Axin的不同信号传导结果。最后是纸巾 将评估成年人中响应于稳定的Axin 2而增加的Wnt信号传导，集中于由Wnt控制的干细胞小生境维持的群体。此外，将评估稳定的Axin 2对两种Wnt相关癌症模型（结直肠癌的Apcmin模型和乳腺癌的MMTV-Wnt 1模型）的影响，重点关注肿瘤的发生率、发作或侵袭性，以确定某些肿瘤的严重程度是否因稳定的Axin蛋白而增加，从而解决Axin稳定药物在癌症治疗中的效用。