Regulation of TCF3 by Wnt signaling

描述（由申请人提供）：t细胞因子（TCF）家族在响应分泌Wnt蛋白的基因表达控制中起关键作用。Wnt通路的作用是调节细胞增殖、迁移行为、细胞命运决定和细胞死亡。本研究旨在研究TCF3的调控，TCF3是一种转录抑制因子，在细胞分化和肿瘤发生过程中涉及Wnt信号传导。我们在非洲爪蟾外胚层和哺乳动物细胞中进行的初步实验发现，TCF3蛋白在翻译后的特异性变化与Wnt靶基因的激活有关。这种效应已被证明与同源域相互作用蛋白激酶（HIPK）有关，这导致了一种假设，即Wnt蛋白通过HIPK2和TCF3来调节其转录靶点。这一假设将在哺乳动物细胞和爪蟾卵中进行测试，其中生化和细胞生物学方法可以与潜在机制的快速功能分析相结合。为了研究TCF3翻译后修饰在wnt依赖性靶基因激活中的作用，将在形态学和分子分析中比较非磷酸化和拟磷TCF3突变蛋白。其他实验将阐明Wnt信号如何激活HIPK并触发对靶基因调控至关重要的TCF3翻译后修饰。这些研究将有助于描述通常导致细胞增殖和分化的基本信号网络，但这些信号网络在疾病期间被错误调节。鉴于Wnt通路与癌症的关联，了解TCF3如何被调控将作为新药筛选的基础，并应导致关键治疗策略的发展。

DESCRIPTION (provided by applicant): The T-cell factor (TCF) family plays key roles in the control of gene expression in response to secreted Wnt proteins. Wnt pathways operate to modulate cell proliferation, migratory behavior, cell fate determination and cell death. This proposal aims to study the regulation of TCF3, a transcriptional repressor which has been implicated in Wnt signaling during cell differentiation and tumorigenesis. Our preliminary experiments in Xenopus ectoderm and mammalian cells identified a specific post-translational change in TCF3 protein mobility that correlates with Wnt target gene activation. This effect has been demonstrated to involve homeodomain-interacting protein kinase (HIPK), leading to a hypothesis that Wnt proteins act via HIPK2 and TCF3 to regulate their transcriptional targets. This hypothesis will be tested in mammalian cells and Xenopus eggs, in which biochemical and cell biological approaches can be combined with rapid functional analysis of underlying mechanisms. To investigate roles of TCF3 postranslational modification for Wnt-dependent activation of target genes, non- phosphorylatable and phosphomimetic TCF3 mutant proteins will be compared in morphological and molecular assays. Other experiments will elucidate how Wnt signals activate HIPK and trigger TCF3 post-translational modifications that are critical for target gene regulation. These studies will help characterize basic signaling networks normally leading to cell proliferation and differentiation, but which are misregulated during disease. Given the association of Wnt pathways with cancer, the knowledge of how TCF3 is regulated will serve as a basis for new drug screens and should lead to the development of crucial therapeutic strategies. PUBLIC HEALTH RELEVANCE: This application concerns the function of TCF3, a Wnt-dependent transcriptional repressor that is essential for the regulation of cell proliferation. Since Wnt pathways have been found activated in a variety of tumors, the proposed studies are highly relevant to human cancer.