The biochemical mechanism of Wnt signaling

描述（由申请人提供）：该提案的长期目标是了解 Wnt 信号以受控方式传输的机制，以及该机制的破坏如何导致发育缺陷和癌症。 Wnt 分泌信号分子家族在动物发育过程中各种组织的分化和模式化中发挥着关键作用。 Wnt 信号传导的异常激活还会导致多种癌症。 Wnt 信号的传递由两种细胞表面受体介导，一种来自卷曲家族，另一种来自低密度脂蛋白受体相关蛋白 (LRP) 家族。然而，目前尚不清楚Wnt结合如何导致受体激活以及受体如何向下游传递Wnt信号。该提案将评估 Wnt 结合诱导的受体聚集激活下游信号传递的假设。为了建立 Wnt 受体激活的机制，本文提出的研究将采用生化和细胞生物学方法来检查在 Wnt 存在或不存在的情况下受体如何彼此相互作用以及与下游成分如何相互作用。由于 Wnt 和 Frizzled 家族均由大量家族成员组成，目前尚不清楚单个 Wnt 如何与各种 Frizzled 蛋白实现特异性相互作用。尽管各种配体-受体对之间的不同亲和力被认为在建立特异性方面发挥决定性作用，但由于缺乏合适的 Wnt 制剂，这一假设很难测试。为了评估这一假设，该提案将通过使用最近可用的可溶性和活性 Wnt 制剂来直接测量 Wnt-Frizzled 相互作用的特异性和亲和力。由于了解 Wnt 的生化特性对于阐明信号传导机制至关重要，因此该提案还将通过诱变、蛋白质纯化和生化表征来确定对其活性重要的 Wnt 的结构和生化特征。对该信号通路的完整理解将为开发高度选择性、基于机制的方法来预防、诊断和治疗由异常 Wnt 功能引起的疾病和癌症提供重要见解。

DESCRIPTION (provided by applicant): The long-term objective of this proposal is to understand the mechanisms by which Wnt signals are transmitted in a controlled manner and how disruption of this mechanism leads to developmental defects and cancers. The Wnt family of secreted signaling molecules plays pivotal roles in the differentiation and patterning of various tissues during animal development. Aberrant activation of Wnt signaling also causes several cancers. Transmission of Wnt signals is mediated by two cell surface receptors, one from the Frizzled family and the other from the low-density lipoprotein receptor-related protein (LRP) family. It is, however, unclear how Wnt binding leads to the activation of the receptors and how the receptors transmit the Wnt signal downstream. This proposal will evaluate the hypothesis that receptor clustering induced by Wnt binding activates the downstream signal transmission. To establish the mechanisms of Wnt receptor activation, the investigations proposed here will employ biochemical and cell biological approaches to examine how receptors interact with each other and with downstream components in the presence or absence of Wnts. Since both the Wnt and Frizzled families consist of a large number of family members, it remains unclear how individual Wnts achieve specific interactions with various Frizzled proteins. Although differing affinities among various ligand-receptor pairs are thought to play deciding roles in establishing the specificities, this assumption has been difficult to test due to the lack of suitable Wnt preparations. To evaluate this hypothesis, this proposal will directly measure the specificities and affinities of Wnt-Frizzled interactions by employing soluble and active Wnt preparations that have become available recently. Since a fundamental knowledge of the biochemical properties of Wnts is essential to the elucidation of the signaling mechanism, this proposal will also determine the structural and biochemical features of Wnts important for their activities through mutagenesis, protein purification and biochemical characterization. A complete understanding of this signaling pathway will provide important insights for the development of highly selective, mechanism-based approaches for the prevention, diagnosis, and treatment of diseases and cancers caused by aberrant Wnt functions.