Mechanism of Wnt signal transduction

Wnt通路是一个进化保守的信号通路，存在于从果蝇到人类的后生动物中。Wnt信号已被证明在发育过程中发挥重要作用。鉴于Wnt通路参与多种人类疾病的发生（例如，超过90%的结直肠癌），人们正在努力开发针对该通路的治疗方法。不幸的是，部分由于我们对Wnt信号转导的详细机制了解不完全，针对这一途径的治疗方法的开发进展缓慢，目前还没有Wnt抑制剂用于临床。我们的首要目标是了解Wnt信号传播的基本生化机制，从而最终协调组织、器官和肢体的形成，并了解其失调如何导致疾病状态。在十多年的时间里，我的实验室已经1)开发了第一个生化系统（爪虫卵提取物），它概括了Wnt通路的关键反应，2)开发了Wnt通路的第一个数学模型（Lee- Heinrich模型），3)为涉及受体介导的细胞质信号传导（通过直接抑制GSK3活性）的机制提供了证据，4)发现了一种Wnt通路的小分子抑制剂，该抑制剂已被FDA指定为治疗家族性癌前病变（家族性腺瘤性息肉病）的孤儿药。我们最近资助的工作主要集中在1)我们新提出的模型，该模型描述了β-catenin（一种Wnt转录辅激活因子）如何通过E3连接酶XIAP促进的过程与辅抑制因子Gro/TLE竞争，以结合转录因子TCF/LEF； 2)新发现的APC肿瘤抑制因子缺失导致Wnt细胞表面受体激活的机制。对于MIRA拨款申请，我建议通过以下方式扩展这些研究：1)确定APC调节受体激活的分子机制，特别是在此过程中囊泡运输的作用；2)追求与XIAP相互作用以促进-catenin介导的Wnt信号传导的去泛素酶USP47的表征。我们已经进行了几个基因组规模的筛选，以确定新的Wnt通路基因。基于这些筛选结果，我建议研究1)涉及跨膜和囊泡转运蛋白Arl4c、ITSN1、Syndecan-2和Cdh13的Wnt信号体形成；2)USP46/UAF1/WDR20去泛素酶复合物对Wnt受体的稳态调节；3)核激酶STK38对Wnt基因转录的调控。

The Wnt pathway is an evolutionarily conserved signaling pathway present in metazoans from Drosophila to humans. Wnt signaling has been shown to play important roles in development. Given that the Wnt pathway is involved in the genesis of a wide variety of human diseases (e.g., over 90% of all colorectal cancers), there is an intense effort to develop therapeutics that target this pathway. Unfortunately, in part due to our incomplete understanding of the detailed mechanism of Wnt signal transduction, progress in developing therapeutics that target this pathway has been slow, and no Wnt inhibitors are currently in clinical use. Our overarching goal is to understand the basic biochemical mechanisms by which a Wnt signal is propagated to ultimately coordinate the formation of tissues, organs, and limbs and to understand how its misregulation can lead to disease states. In over more than a decade, my laboratory has 1) developed the first biochemical system (Xenopus egg extract) that recapitulated key reactions of the Wnt pathway, 2) developed the first mathematical model (Lee- Heinrich model) of the Wnt pathway, 3) provided evidence for a mechanism involving receptor-mediated signaling (via direct inhibition of GSK3 activity) to the cytoplasm, and 4) identified a small molecule inhibitor of the Wnt pathway that has been designated by the FDA as an orphan drug for a familial precancerous disease (familial adenomatous polyposis). Our recently funded work focuses on 1) our newly proposed model describing how β-catenin, a Wnt transcriptional coactivator, competes with the corepressor Gro/TLE for binding to the transcriptional factor, TCF/LEF, via a process facilitated by the E3 ligase XIAP and 2) a newly identified mechanism by which loss of the APC tumor suppressor leads to activation of Wnt cell surface receptors. For the MIRA grant application, I propose to extend these studies by 1) determining the molecular mechanism by which APC regulates receptor activation, specifically the role of vesicle transport in this process, and 2) pursuing the characterization of a deubiqutinase, USP47, which interacts with XIAP to promote -catenin-mediated Wnt signaling. We have undertaken several genome-scale screens to identify new Wnt pathway genes. Based on the results of these screens, I propose to study 1) Wnt signalosome formation involving the transmembrane and vesicular transport proteins: Arl4c, ITSN1, Syndecan-2, and Cdh13, 2) Wnt receptor homeostasis by the USP46/UAF1/WDR20 deubiquitinase complex, and 3) regulation of Wnt gene transcription by the nuclear kinase STK38.