Quantifying β-catenin subcellular dynamics and cyclin D1 mRNA transcription during Wnt signaling in single living cells

Abstract

Signal propagation from the cell membrane to a promoter can induce gene expression. To examine signal transmission through sub-cellular compartments and its effect on transcription levels in individual cells within a population, we used the Wnt/β-catenin signaling pathway as a model system. Wnt signaling orchestrates a response through nuclear accumulation of β-catenin in the cell population. However, quantitative live-cell measurements in individual cells showed variability in nuclear β-catenin accumulation, which could occur in two waves, followed by slow clearance. Nuclear accumulation dynamics were initially rapid, cell cycle independent and differed substantially from LiCl stimulation, presumed to mimic Wnt signaling. β-catenin levels increased simultaneously at adherens junctions and the centrosome, and a membrane-centrosome transport system was revealed. Correlating β-catenin nuclear dynamics to cyclin D1 transcriptional activation showed that the nuclear accumulation rate of change of the signaling factor, and not actual protein levels, correlated with the transcriptional output of the pathway.DOI: http://dx.doi.org/10.7554/eLife.16748.001

β-catenin-Mediated Wnt Signal Transduction Proceeds Through an Endocytosis-Independent Mechanism

“…Single cells live imaged for up to 10 hours were used to quantify cellular accumulation and nuclear translocation of βcatenin. Consistent with previous reports of β-catenin dynamics upon pathway activation, individual cells presented significant variability in their response in terms of the rate and extent of change in β-catenin (8,25,26). Some cells exhibited substantial increase in total and nuclear β-catenin within an hour while others showed slow or minor increase, reflect-ing the inherent variability in the cells' molecular machinery to respond to Wnt signaling.…”

“…As mentioned previously, individual cells have been shown to present significant variability in absolute level changes in β-catenin (8,25,26). Indeed, we observed variability in absolute levels and dynamics of β-catenin accumulation among both wildtype and Ap2α knockout cells ( Supplementary Fig 6A).…”

Retracted: Inhibition of microRNA‐940 suppresses the migration and invasion of human osteosarcoma cells through the secreted frizzled‐related protein 1–mediated Wnt/β‐catenin signaling pathway

“…SFRP1 can combine with Wnt receptor through its CRD and suppress the interaction between FZD receptor and the Wnt ligand, causing β‐catenin phosphorylation and reducing its content . When Wnt signaling pathway was activated, signals from the Wnt could release β‐catenin from its binding protein, which would accumulate in the cytoplasm and then shift into the cell nucleus . In the cell nucleus, β‐catenin would bind to TCF/LEF protein and activate the transcription and expression of target gene cyclinD1, thereby upregulating miR‐940 and suppressing the expression of SFRP1 .…”

Synergy with TGFß ligands switches WNT pathway dynamics from transient to sustained during human pluripotent cell differentiation

“…Our GFP-ß-catenin knock-in labeling strategy is quantitative, contains high temporal resolution, conserves spatial information, and in contrast to ß-catenin-fluorescent-protein over-expression reporters (38), maintains pathway stoichiometry. In the future we will be able to combine this approach with self-patterning differentiation assays (e.g., organoids, gastruloids, and embryoids) (39,40), providing us with the signaling trajectories required for each cell-fate.…”