Regulation of β-Catenin Nuclear Dynamics by GSK-3β Involves a LEF-1 Positive Feedback Loop

Nuclear localization of beta-catenin is integral to its role in Wnt signaling and cancer. Cellular stimulation by Wnt or lithium chloride (LiCl) inactivates glycogen synthase kinase-3 beta (GSK-3 beta), causing nuclear accumulation of beta-catenin and transactivation of genes that transform cells. beta-catenin is a shuttling protein; however, the mechanism by which GSK-3 beta regulates beta-catenin nuclear dynamics is poorly understood. Here, fluorescence recovery after photobleaching assays were used to measure the beta-catenin-green fluorescent protein dynamics in NIH 3T3 cells before and after GSK-3 beta inhibition. We show for the first time that LiCl and Wnt3a cause a specific increase in beta-catenin nuclear retention in live cells and in fixed cells after detergent extraction. Moreover, LiCl reduced the rate of nuclear export but did not affect import, hence biasing beta-catenin transport toward the nucleus. Interestingly, the S45A mutation, which blocks beta-catenin phosphorylation by GSK-3 beta, did not alter nuclear retention or transport, implying that GSK-3 beta acts through an independent regulator. We compared five nuclear binding partners and identified LEF-1 as the key mediator of Wnt3a and LiCl-induced nuclear retention of beta-catenin. Thus, Wnt stimulation triggered a LEF-1 positive feedback loop to enhance the nuclear chromatin-retained pool of beta-catenin by 100-300%. These findings shed new light on regulation of beta-catenin nuclear dynamics.