Department of Pharmacology, Howard Hughes Medical Institute, and the Institute for Stem Cell and Regenerative Medicine, University of Washington School of Medicine, Seattle, WA 98109, USA.
Proc Natl Acad Sci U S A. 2012 Mar 20;109(12):4485-90. doi: 10.1073/pnas.1118777109. Epub 2012 Mar 5.
Signal transduction pathways play diverse, context-dependent roles in vertebrate development. In studies of human embryonic stem cells (hESCs), conflicting reports claim Wnt/β-catenin signaling promotes either self-renewal or differentiation. We use a sensitive reporter to establish that Wnt/β-catenin signaling is not active during hESC self-renewal. Inhibiting this pathway over multiple passages has no detrimental effect on hESC maintenance, whereas activating signaling results in loss of self-renewal and induction of mesoderm lineage genes. Following exposure to pathway agonists, hESCs exhibit a delay in activation of β-catenin signaling, which led us to postulate that Wnt/β-catenin signaling is actively repressed during self-renewal. In support of this hypothesis, we demonstrate that OCT4 represses β-catenin signaling during self-renewal and that targeted knockdown of OCT4 activates β-catenin signaling in hESCs. Using a fluorescent reporter of β-catenin signaling in live hESCs, we observe that the reporter is activated in a very heterogeneous manner in response to stimulation with Wnt ligand. Sorting cells on the basis of their fluorescence reveals that hESCs with elevated β-catenin signaling express higher levels of differentiation markers. Together these data support a dominant role for Wnt/β-catenin signaling in the differentiation rather than self-renewal of hESCs.
信号转导通路在脊椎动物的发育过程中发挥着多样化、依赖于背景的作用。在对人类胚胎干细胞（hESC）的研究中，相互矛盾的报告声称 Wnt/β-连环蛋白信号通路促进自我更新或分化。我们使用一种敏感的报告基因来确定 Wnt/β-连环蛋白信号通路在 hESC 自我更新过程中不活跃。在多个传代过程中抑制该途径对 hESC 的维持没有不利影响，而激活信号通路则导致自我更新的丧失和中胚层谱系基因的诱导。在暴露于途径激动剂后，hESC 表现出 β-连环蛋白信号通路激活的延迟，这使我们假设 Wnt/β-连环蛋白信号通路在自我更新过程中被积极抑制。为了支持这一假设，我们证明 OCT4 在自我更新过程中抑制β-连环蛋白信号通路，并且靶向敲低 OCT4 可激活 hESC 中的β-连环蛋白信号通路。我们使用活 hESC 中的β-连环蛋白信号的荧光报告基因观察到，该报告基因在受到 Wnt 配体刺激时以非常异质的方式被激活。根据荧光强度对细胞进行分选，揭示了具有高β-连环蛋白信号的 hESC 表达更高水平的分化标志物。这些数据共同支持 Wnt/β-连环蛋白信号通路在 hESC 的分化而非自我更新中发挥主要作用。
