Regulation of TCF3 by Wnt signaling

描述(由申请人提供)：T细胞因子(TCF)家族在控制分泌的Wnt蛋白的基因表达方面发挥关键作用。WNT通路调节细胞增殖、迁移行为、决定细胞命运和细胞死亡。这项建议旨在研究转录抑制因子TCF3的调控，该转录抑制因子参与了细胞分化和肿瘤发生过程中的Wnt信号。我们在非洲爪哇外胚层和哺乳动物细胞中的初步实验发现，TCF3蛋白迁移率在翻译后发生了特定的变化，与Wnt靶基因的激活相关。这一效应已被证明涉及同源结构域相互作用蛋白激酶(HIPK)，导致了一种假设，即Wnt蛋白通过HIPK2和TCF3调节其转录靶标。这一假设将在哺乳动物细胞和非洲爪哇卵中进行测试，在这些细胞中，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依赖的靶基因激活中的作用，我们将从形态和分子水平上比较非磷酸化和拟磷酸化的TCF3突变蛋白。其他实验将阐明Wnt信号如何激活HIPK并触发TCF3翻译后修饰，这对靶基因调控至关重要。这些研究将有助于确定通常导致细胞增殖和分化的基本信号网络的特征，但这些网络在疾病期间被错误地调控。鉴于Wnt通路与癌症的关联，了解TCF3是如何被调控的将成为新药筛选的基础，并应导致关键治疗策略的开发。

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.