Transcriptional Activation by Wnt Signaling

描述（由申请人提供）： Wnt家族的分泌糖蛋白通过进化上保守的信号级联起作用，该信号级联促进β-连环蛋白的核积累，β-连环蛋白与DNA结合蛋白的TCF家族的成员相互作用以激活靶基因转录。TCF与特定序列结合，但共识是如此松散，以至于可以在整个基因组中找到潜在的TCF结合位点。尽管如此，我们发现TCF结合到Wnt靶标的特定位置，对应于Wnt应答元件（WRE）。WRE的系统诱变揭示了与TCF结合位点一起作用以介导Wnt信号传导的活化的额外基序的存在。这些基序（称为辅助位点）在其他几个WRE中也有发现，对保守的TCF/辅助位点簇的全基因组搜索已经确定了其他推定的WRE。将测试辅助站点在这些元素中的功能意义。将探索辅助位点如何与TCF结合位点相互作用的分子机制，并表征与其结合的反式作用因子。β-连环蛋白与TCF的结合将其从转录阻遏物转化为激活物。我们已经发现Wnt信号促进整个靶位点的组蛋白乙酰化，这与转录的激活相关。这种广泛的修饰似乎需要拮抗沉默Wnt靶的因子的作用。这些过程之间的机制和关系将被详细探讨。我们的数据表明，Wnt靶点的转录开关涉及的染色质结构变化远远超出了以前的认识。项目叙述：Wnt信号通路在发育过程中的细胞命运决定中起着重要作用，并且是维持成体组织中干细胞群体所必需的。该途径的失调在许多人类癌症中起着因果作用。我们的研究旨在了解除了TCF位点之外有助于WRE功能的基序的作用，这将导致更好的生物信息学方法来识别许多重要生物背景中的Wnt靶点。我们对染色质修饰在调节TCF转录开关中的作用的研究将作为研究哺乳动物系统中这些过程的范例。

DESCRIPTION (provided by applicant): Secreted glycoproteins of the Wnt family act through an evolutionarily conserved signaling cascade which promotes the nuclear accumulation of beta-catenin, which interacts with members of the TCF family of DNA-binding proteins to activate target gene transcription. TCFs binds to specific sequences, but the consensus is so loose that potential TCF binding sites can be found throughout the genome. Despite this, we find that TCF is bound to specific locations in Wnt targets, corresponding to Wnt response elements (WREs). Systematic mutagenesis of a WRE revealed the presence of additional motifs that act with TCF binding sites to mediate activation by Wnt signaling. These motifs (called Helper sites) are found in several other WREs, and genome-wide searches for conserved TCF/Helper site clusters have identified other putative WREs. The functional significance of the Helper sites in these elements will be tested. The molecular mechanism of how Helper sites interact with TCF binding sites will be explored, and the trans-acting factor(s) that bind to it will be characterized. Binding of beta-catenin to TCF converts it from a transcriptional repressor to an activator. We have found that Wnt signaling promotes histone acetylation throughout target loci, which is correlated with activation of transcription. This widespread modification appears to be required to antagonize the action of factors that silence Wnt targets. The mechanisms and relationship between these processes will be explored in detail. Our data indicates that the transcriptional switch at Wnt targets involves changes in chromatin structure far beyond what was previously recognized. PROJECT NARRATIVE: The Wnt signaling pathway plays important roles in cell fate decisions during development, and is required for the maintenance of stem cell populations in adult tissues. Misregulation of the pathway plays a causal role in many human cancers. Our studies to understand the role of motifs besides TCF sites that contribute to WRE function will lead to better bioinformatic methods to identify Wnt targets in many important biological contexts. Our work on the role of chromatin modifications in regulating the TCF transcriptional switch will serve as a paradigm to study these processes in mammalian systems.