Molecular mechanisms of Wnt and mechanical signaling through β-catenin

Wnt 的分子机制和通过 β-catenin 的机械信号传导

• 批准号: 10404076
• 负责人: William I Weis
• 金额: $ 73.31万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-06-07 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10404076
• 关键词: APC gene; Actins; Address; Adherens Junction; Adult; Affect; Area; Binding; Biochemical; Cadherins; Cell Fate Control; Cell Nucleus; Cell-Cell Adhesion; Cells; Complex; Cysteine-Rich Domain; Cytoplasmic Protein; Defect; Development; Dsh protein; E-Cadherin; Embryonic Development; Epithelial; Family; Frizzled Domain; Genes; Goals; Growth Factor; Homeostasis; Intercellular Junctions; Knowledge; Ligands; Link; Maintenance; Malignant Neoplasms; Microfilaments; Molecular; Molecular Conformation; Neoplasms; Pathway interactions; Phosphorylation; Phosphotransferases; Process; Property; Proteins; Role; Scaffolding Protein; Signal Transduction; Signaling Protein; Solid; Specific qualifier value; Tight Junctions; Tissues; Vinculin; WNT Signaling Pathway; Wnt proteins; afadin; alpha catenin; base; beta catenin; biophysical techniques; extracellular; mechanical force; mechanical signal; multicatalytic endopeptidase complex; novel strategies; protein B; receptor; recruit; tissue regeneration; transmission process; zonula occludens-1 protein

The development and homeostasis of solid tissues depends upon biochemical and mechanical signals that control cell fate and the resulting organization of cells in the tissue. The conserved protein β-catenin is a key effector of signals from both the Wnt family of secreted growth factors that specify cell fate during embryogenesis and tissue renewal in the adult, and mechanical force transmitted through cell-cell junctions in multicellular tissues. We hypothesize that mechanical force transmitted through β-catenin links Wnt signaling and cell-cell adhesion, and our overall goal is to understand the molecular mechanisms underlying these dual roles of β-catenin. Our strategy is to use biochemical, structural and biophysical methods to address critical knowledge gaps in these areas. In the absence of Wnts, the β-catenin is bound in a “destruction complex” that includes the proteins Axin and Adenomatous Polyposis Coli (APC), and kinases that phosphorylate β-catenin; phosphorylation leads to ubiquitylation and destruction of β-catenin by the proteasome. Wnt binding to the receptors Frizzled (Fzd) and LRP5/6 enables Fzd to recruit the cytoplasmic protein Dishevelled (Dvl), which in turn binds to Axin and thereby recruits the destruction complex to the activated receptor complex. This leads to phosphorylation of the LRP5/6 intracellular domain, which inhibits β-catenin destruction; the stabilized β-catenin enters the nucleus and activates target genes. We will address critical mechanistic aspects of this pathway that are not understood: 1) how secreted ligands “activate” the Fzd-Dvl interaction needed for β-catenin stabilization through interaction with the extracellular cysteine-rich domain of Fzd and LRP5/6; 2) how activated Dvl recruits Axin to turn off β-catenin destruction; 3) how the β-catenin destruction complex forms and interacts with the ubiquitylation/proteosomal machinery; 4) the essential role of APC in β-catenin destruction. Force transmission through cell-cell adherens junctions (AJ) requires a complex of E-cadherin, β-catenin, and α-catenin, which binds to actin filaments and forms a minimal force-sensing unit. Tension on cadherins can release β-catenin and cause its translocation to the nucleus independent of, but synergized by, Wnt signaling. Understanding such tension-triggered release of β-catenin requires understanding how force is transmitted through the AJ complex. α-Catenin additionally has a central role in organizing epithelial tissues based on its interactions with vinculin, Epithelial Protein Lost in Neoplasm (EPLIN), the tight junction (TJ) protein Zonula Occludens (ZO)-1, and afadin, all of which bind actin and recruit other scaffolding and signaling proteins. We will study: 1) The force-dependent conformational landscape and force responsivness of α-catenin alone and bound to its partners, including how β-catenin modifies α-catenin force responsiveness; 2) How αE-catenin conformation and force transmission properties are affected by its binding to its other junctional partners.

固体组织的发育和稳态取决于生化和机械信号 控制细胞命运和组织中细胞的组织组织。保守的蛋白β-catenin是关键 来自两个分泌生长因子的Wnt家族的信号的效应子，这些因子指定细胞命运 成人的胚胎发生和组织更新，以及通过细胞 - 细胞连接传播的机械力 在多细胞组织中。我们假设通过β-catenin链接传输的机械力wnt 信号传导和细胞细胞粘附，我们的总体目标是了解分子机制 这些双重作用的β-catenin。我们的策略是使用生化，结构和生物物理方法 解决这些领域的关键知识差距。 在不存在WNT的情况下，β-catenin与包括蛋白质轴的“破坏复合物”结合 磷酸化β-catenin的腺瘤性息肉病（APC）和激酶；磷酸化导致 蛋白酶体对β-catenin的泛素化和破坏。 Wnt与受体的结合（FZD） LRP5/6使FZD能够募集细胞质蛋白质染色（DVL），这又与Axin结合 从而向激活的接收器复合物报告了破坏复合物。这导致 LRP5/6细胞内结构域，抑制β-catenin毁灭；稳定的β-catenin进入 细胞核和激活靶基因。我们将解决该途径的关键机理方面 理解齿：1）分泌的配体如何“激活”β-catenin稳定所需的FZD-DVL相互作用 通过与FZD和LRP5/6的细胞外半胱氨酸域的相互作用； 2）激活的DVL如何 招募Axin关闭β-catenin的破坏； 3）β-catenin破坏如何形成和相互作用 带有泛素化/蛋白质体机械； 4）APC在β-catenin破坏中的基本作用。 通过细胞细胞粘附连接（AJ）的力传递需要E-钙粘蛋白的β-catenin，β-catenin， 和α-catenin，它与肌动蛋白丝结合并形成最小的力感应单元。钙粘蛋白的张力 可以释放β-catenin，并使其转移到核中，而与Wnt合成，但合成 信号。了解这种张力触发的β-catenin的释放需要了解力的力量 通过AJ复合物传输。 α-catenin在组织上皮组织中具有核心作用 基于其与杂种蛋白的相互作用，上皮蛋白在肿瘤（Eplin）中丧失，紧密连接（TJ） 蛋白质Zonula occludens（ZO）-1和Afadin，所有这些都结合肌动蛋白并募集其他脚手架， 信号蛋白。我们将研究：1）依赖力的会议景观和力量 单独α-catenin并与其伴侣结合，包括β-catenin如何修饰α-catenin 力量反应； 2）αe-catenin构象和力传递特性如何受其影响 与其其他连接伙伴结合。