Fig. 7: Both FER kinase and PIEZO1 are required for the cell front retraction wave. | Nature Communications

Fig. 7: Both FER kinase and PIEZO1 are required for the cell front retraction wave.

From: Spatiotemporal H2O2 flashes coordinate actin cytoskeletal remodeling and regulate cell migration and wound healing

Fig. 7

a FER (red) localization during wound healing in BxPC3 DUOX1/2 WT HyPer7-MEM cells, HyPer7 (merge, green; or intensity heatmap) and actin (purple), coverslips were fixed at the indicated timepoints post wounding. Cell front speed (b) and change in wound area (c) in BxPC3 DUOX1/2 and DUOX1/2 KO cell lines ± FER inhibition (E260 20 µM). d Colocalization of PIEZO1 (purple) and FER (red) at the wound edge during scratch assay in fixed BxPC3 DUOX1/2 WT cells. e Colocalization of PIEZO1 (purple) and DUOX (red) at the wound edge in fixed BxPC3 DUOX1/2 WT cells. f Cell front speed during PIEZO1 stimulation (Yoda1 5 µM) or calcium channel inhibition (GsMTx4 5 µM) in BxPC3 DUOX1/2 WT cells (blue, Yoda1; light blue, GsMTx4) or BxPC3 DUOX1/2 KO cells (red, Yoda1; light red, GsMTx4). Brightfield frames extracted from live cell scratch assay movies showing time-dependent wound closure for (g), Yoda1-treated BxPC3 DUOX1/2 KO cells, red arrows indicate areas of cell front arrest (Supplementary Movie 10), and h, GsMTx4-treated BxPC3 DUOX1/2 WT cells, green lines indicate initial wound edges at t = 0 h, red lines indicate wound edges at the indicated timepoints, red arrows show area of damaged cells. i Brightfield images extracted from live cell scratch assays at indicated timepoints, see elimination of damaged cell front in BxPC3 DUOX1/2 WT cells (upper panels, green arrows) and inefficient incorporation of the damaged cell front in BxPC3 DUOX1/2 KO cells (lower panels, red arrows). j Analysis of monolayer permeability using FITC-dextran (4 kDa) in BxPC3 DUOX1/2 WT (blue) and DUOX1/2 KO (red) cell lines for 24 h pre-wound followed by measurements at 3 h, 26 h, and 50 h post wounding, individual Welch’s t test at each time point, *P < 0.05. Each point represents the average from 3 independent experiments, each with 3 replicates, Data are presented as mean ± SEM. k, Scheme of PIEZO1-DUOX2-FER engagement during wound healing. b, c, f, n = each point represents the average cell front speed or wound area for every 3 h from 3 independent experiments, each with 6 replicates. a, d, e, g, h, i Scale bars 100 µm, microscopy analyses representative of n = 5. k Created in BioRender. Knaus, U. (2025) https://BioRender.com/putget1, modified in Adobe Illustrator.

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