Department of Anatomy and Cell Biology, Carver College of Medicine, The University of Iowa, Iowa City, Iowa 52240, USA.
Am J Physiol Lung Cell Mol Physiol. 2010 Nov;299(5):L694-710. doi: 10.1152/ajplung.00140.2010. Epub 2010 Aug 27.
Wnt/β-catenin-dependent activation of lymphoid enhancer factor 1 (Lef-1) plays an important role in numerous developmental processes. In this context, transcription of the Lef-1 gene is increased by Wnt-mediated TCF4/β-catenin activation on the Lef-1 promoter through mechanisms that remain poorly defined. In mouse airway submucosal gland progenitor cells, Wnt3A transiently induces Lef-1 gene expression, and this process is required for epithelial cell proliferation and glandular morphogenesis. In the present study, we sought to identify additional candidate transcriptional regulators of the Lef-1 gene during glandular morphogenesis. To this end, we found that Sox17 expression is dramatically downregulated in early glandular progenitor cells that induce Lef-1 expression. Wnt stimulation of undifferentiated primary airway epithelial cells induced similar changes in Sox17 and Lef-1 expression. Reporter assays revealed that ectopic expression of Sox17 suppresses Wnt3A/β-catenin activation of the Lef-1 promoter in cell lines. EMSA and ChIP analyses defined several Sox17- and TCF4-binding sites that collaborate in transcriptional control of the Lef-1 promoter. More specifically, Sox17 bound to four sites in the Lef-1 promoter, either directly or indirectly through TCF complexes. The DNA- or β-catenin-binding domains of Sox17 controlled context-specific binding of Sox17/TCF complexes on the Lef-1 promoter. Combinatorial site-directed mutagenesis of Sox17- or TCF-binding sites in the Lef-1 promoter demonstrated that these sites control Wnt/β-catenin-mediated induction and/or repression. These findings demonstrate for the first time that Sox17 can directly regulate Wnt/β-catenin-dependent transcription of the Lef-1 promoter and reveal new context-dependent binding sites in the Lef-1 promoter that facilitate protein-protein interactions between Sox17 and TCF4.
Wnt/β-catenin 依赖性激活淋巴增强因子 1（Lef-1）在许多发育过程中发挥重要作用。在这种情况下，Lef-1 基因的转录通过仍然定义不明确的机制，通过 Wnt 介导的 TCF4/β-catenin 激活在 Lef-1 启动子上增加。在小鼠气道粘膜下腺祖细胞中，Wnt3A 短暂诱导 Lef-1 基因表达，并且这个过程是上皮细胞增殖和腺体形态发生所必需的。在本研究中，我们试图在腺体形态发生过程中鉴定 Lef-1 基因的其他候选转录调节剂。为此，我们发现 Sox17 表达在诱导 Lef-1 表达的早期腺祖细胞中显著下调。未分化的原代气道上皮细胞的 Wnt 刺激诱导 Sox17 和 Lef-1 表达发生类似变化。报告基因检测表明，Sox17 的异位表达抑制细胞系中 Wnt3A/β-catenin 对 Lef-1 启动子的激活。EMSA 和 ChIP 分析定义了几个 Sox17 和 TCF4 结合位点，它们协同转录控制 Lef-1 启动子。更具体地说，Sox17 直接或间接通过 TCF 复合物结合到 Lef-1 启动子的四个位点。Sox17 的 DNA 或 β-catenin 结合域控制 Sox17/TCF 复合物在 Lef-1 启动子上的特定结合。在 Lef-1 启动子上的 Sox17 或 TCF 结合位点的组合定点突变显示，这些位点控制 Wnt/β-catenin 介导的诱导和/或抑制。这些发现首次表明 Sox17 可以直接调节 Lef-1 启动子的 Wnt/β-catenin 依赖性转录，并揭示了 Lef-1 启动子中促进 Sox17 和 TCF4 之间蛋白-蛋白相互作用的新的上下文相关结合位点。
