Mechanism of Wnt signal transduction

Wnt通路是存在于从果蝇到人类的后生动物中的进化上保守的信号通路。Wnt信号已被证明在发育中发挥重要作用。鉴于Wnt途径参与多种人类疾病的发生（例如，超过90%的结直肠癌），人们正在努力开发靶向该途径的治疗方法。不幸的是，部分由于我们对Wnt信号转导的详细机制的不完全理解，开发靶向该途径的治疗方法的进展缓慢，并且目前没有Wnt抑制剂用于临床。我们的总体目标是了解Wnt信号传播的基本生化机制，以最终协调组织，器官和肢体的形成，并了解其失调如何导致疾病状态。十多年来，我的实验室开发出第一套生化系统，（非洲爪蟾卵提取物），概括了Wnt途径的关键反应，2）开发了第一个数学模型（Lee-Heinrich模型），3）为涉及受体介导的信号传导的机制提供了证据（通过直接抑制GSK 3活性）至细胞质，和4）鉴定了Wnt途径的小分子抑制剂，其已被FDA指定为用于家族性癌前疾病（家族性腺瘤性息肉病）的孤儿药。我们最近资助的工作集中在1）我们新提出的模型，描述了β-catenin，一种Wnt转录共激活因子，如何通过E3连接酶XIAP促进的过程与辅阻遏物Gro/TLE竞争结合转录因子TCF/LEF，2）一种新发现的机制，APC肿瘤抑制因子的缺失导致Wnt细胞表面受体的激活。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调节受体活化的分子机制，特别是囊泡转运在此过程中的作用，以及2）追求去泛素化酶USP 47的表征，USP 47与XIAP相互作用以促进β-连环蛋白介导的Wnt信号传导。我们已经进行了几个基因组规模的筛选，以确定新的Wnt途径基因。基于这些筛选的结果，我建议研究1）Wnt信号体的形成，涉及跨膜和囊泡转运蛋白：Arl 4c，ITSN 1，Syndecan-2，和Cdh 13，2）Wnt受体稳态的USP 46/UAF 1/WDR 20去泛素化酶复合物，和3）Wnt基因转录的调节核激酶STK 38。

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.