TY - JOUR TI - Quantitative live-cell imaging and computational modeling shed new light on endogenous WNT/CTNNB1 signaling dynamics AU - de Man, Saskia MA AU - Zwanenburg, Gooitzen AU - van der Wal, Tanne AU - Hink, Mark A AU - van Amerongen, Renée A2 - Campelo, Felix A2 - Walczak, Aleksandra M A2 - Peifer, Mark VL - 10 PY - 2021 DA - 2021/06/30 SP - e66440 C1 - eLife 2021;10:e66440 DO - 10.7554/eLife.66440 UR - https://doi.org/10.7554/eLife.66440 AB - WNT/CTNNB1 signaling regulates tissue development and homeostasis in all multicellular animals, but the underlying molecular mechanism remains incompletely understood. Specifically, quantitative insight into endogenous protein behavior is missing. Here, we combine CRISPR/Cas9-mediated genome editing and quantitative live-cell microscopy to measure the dynamics, diffusion characteristics and absolute concentrations of fluorescently tagged, endogenous CTNNB1 in human cells under both physiological and oncogenic conditions. State-of-the-art imaging reveals that a substantial fraction of CTNNB1 resides in slow-diffusing cytoplasmic complexes, irrespective of the activation status of the pathway. This cytoplasmic CTNNB1 complex undergoes a major reduction in size when WNT/CTNNB1 is (hyper)activated. Based on our biophysical measurements, we build a computational model of WNT/CTNNB1 signaling. Our integrated experimental and computational approach reveals that WNT pathway activation regulates the dynamic distribution of free and complexed CTNNB1 across different subcellular compartments through three regulatory nodes: the destruction complex, nucleocytoplasmic shuttling, and nuclear retention. KW - WNT/CTNNB1 signaling KW - wnt signal transduction KW - beta-catenin KW - quantitative microscopy KW - fluorescence fluctuation spectroscopy KW - computational modeling JF - eLife SN - 2050-084X PB - eLife Sciences Publications, Ltd ER -