Enhancer pioneering activity of Wnt/β-catenin signaling

Wnt/β-catenin signaling is widely understood as a molecular switch that activates preprogrammed regulatory states. Whether it also actively reshapes chromatin to establish those states has remained unclear. To address this, we performed time-resolved ATAC-seq and CUT&RUN analyses in HEK293T cells and monitored β-catenin-dependent changes in chromatin accessibility and complex assembly following pathway activation. In addition to conventional Wnt-responsive elements (WREs), which are constitutively accessible and pre-bound by TCF/LEF transcription factors (TFs), we identified a second class of WREs, that are initially inaccessible but become activated upon stimulus. At these latent enhancers, β-catenin functions as a pioneer-like factor together with HDAC1 and CBP, mediating chromatin remodeling and TCF/LEF occupancy. Together, these results extend the current view of Wnt/β-catenin signaling by showing that it can establish regulatory elements de novo . Here, β-catenin emerges as the central regulator of transcriptional control and remodeling, providing a framework for the dynamic and context-dependent nature of Wnt-driven gene regulation in development and disease.