(A) Left: Schematic representation of the single cell migration assay. Right: fluorescence image of nucleus-stained cells and their migration tracks in yellow. Scale bar = 100µm. (B) MSD(Δt) of WT cells fit with the Fürth model. (C) Cumulative distributions of the step length R for Δt = 10min. One blue curve per frame pair along a movie. The black line is a global fit of all blue curves with a cumulative Rayleigh distribution function. (D) Left: Schematic migration track and angular memory (cos θ(Δt), see Materials and Methods) at Δt = 10min (yellow) and Δt = 250min (blue). Right: Angular memory (cos θ(Δt)) between Δt = 10min (yellow dot) and Δt = 250min (blue dot). Data of (B-D) from 94 cell tracks of 2 independent experiments. Mean ± SEM.

(A) MSD(Δt) of vinculin FL and mutant cells fit with the Fürth model. (B) Apparent speed of single, vinculin FL and mutant cells. (C) Left: direct fluorescence of vinculin TSMod in a single cell. Right: FRET Index map (see Materials and Methods). Scale bar = 20 µm. (D) FRET Index of TSMod in the FL, TL, T12 and YE constructs in FAs. (E) Fluorescence recovery after photobleaching of FL, TL, T12 and YE constructs in FAs fit with a 2-phases association model. (F) Proposed model: at FAs, the head-tail interaction of vinculin (green) competes with both tail dimerization and actin filaments (red) binding, which provide FA stability and force transmission, respectively, together promoting cell speed. Data of (A,B) from 125 (FL), 129 (TL), 193 (T12) and 110 (YE) cell tracks of 3 independent experiments. Data of (D) from 81 (FL), 102 (TL), 52 (T12) and 84 (YE) cells of 4 (FL) and 3 (TL, T12, YE) independent experiments. Data of (E) from 105 (FL), 112 (TL), 118 (T12) and 65 (YE) cells of 2 independent experiments. Mean ± SEM. Two-tailed Mann-Whitney tests between each mutant and FL (B,D). Extra-sum-of-squares F test with shared immobile fraction between each mutant and FL as the null hypothesis (E).

(A) Left: Schematic representation of the collective cell migration assay. Right: fluorescence image of nucleus-stained cells and their migration tracks in yellow. Scale bar = 100µm. (B) MSD(Δt) of WT cells fit with a straight line in ln-ln scale. (C) Cumulative distributions of the step length R for Δt = 10min. One blue curve per frame pair along a movie. The black line is a global fit of all blue curves with a cumulative Rayleigh distribution function. (D) Angular memory (cos θ(Δt)) between Δt = 10min (yellow dot) and Δt = 250min (blue dot). (E) MSD(Δt) of E-cadherin KO and α-catenin KD cells fit with a straight line in ln-ln scale. (F) Angular memory (cos θ(Δt)) of E-cadherin KO and α-catenin KD cells. Data of (B-D) from 11 Fields Of View (FOV, 500-1000 tracks each) of 3 independent experiments. Data of (E-F) from 3 (E-cadherin KO) and 4 (α-catenin KD) FOV (500-100 tracks each) of 2 (E-cadherin KO) and 3 (α-catenin KD) independent experiments. Mean ± SEM. (G) Migration behavior of Self-Propelled Particles with Vicsek interactions as a function of P0 /τ and number of interacting neighbors n = πρd2. Blue and orange points are simulation coordinates that resulted in persistent and super-diffusive behavior, respectively. Cold and warm color backgrounds are interpolations from points above of P/P0 and α values, respectively.

(A) MSD(Δt) of vinculin FL and mutant cells fit with a straight line in ln-ln scale. (B) Apparent speed of vinculin FL and mutant cells at confluence. (C) Left: direct fluorescence of vinculin TSMod in a confluent monolayer. Right: FRET Index map (see Materials and Methods). Top: in the AJs plane. Bottom: in the FAs plane. Scale bar = 20µm. (D) FRET Index of TSMod in the FL, TL, T12 and YE constructs in AJs at confluence. (E) FRET Index of TSMod in the FL, TL, T12 and YE constructs in FAs at confluence. (F) Fluorescence recovery after photobleaching of FL, TL, T12 and YE constructs in AJs fit with a 2-phases association model. (G) Proposed model: at AJs, α-catenin (orange) provides AJ integrity (connection to the cytoskeleton), which promotes persistence. The head-tail interaction of vinculin (green) competes with both tail dimerization and actin filaments (red) binding. Dimerization provides AJ stability, which slows cell down. Data of (A-B) from 12 (FL), 11 (TL, T12) and 14 (YE) FOV (500-1000 tracks each) of 3 independent experiments. Data of (D) from 169 (FL), 30 (TL) and 64 (T12, YE) contacts of 5 (FL) and 3 (TL, T12, YE) independent experiments, and (E) from 45 (FL), 35 (TL), 104 (T12) and 67 (YE) cells of 4 (FL) and 3 (TL, T12, YE) independent experiments. Data of (F) from 75 (FL), 137 (TL), 78 (T12) and 36 (YE) contacts of 4 (FL, TL, T12) and 3 (YE) independent experiments. Mean ± SEM. Two-tailed Mann-Whitney tests between each mutant and FL (B,D,E). Extra-sum-of-squares F test with shared immobile fraction between each mutant and FL as the null hypothesis (F).

(A) Left: Schematic representation of the guided collective cell migration assay. Right: fluorescence image of nucleus-stained cells and their migration tracks in yellow. Scale bar = 100µm. (B) MSD(Δt) of WT cells fit with a straight line in ln-ln scale. (C) MSD(Δt) of E-cadherin KO and α-catenin KD cells fit with a straight line in ln-ln scale. (D) MSD(Δt) of vinculin FL and mutant cells fit with a straight line in ln-ln scale. (E) Angular distribution of apparent velocities of vinculin FL cells at 6hrs post-wound fitted with a Von Mises distribution giving a concentration κ ∼ 1.3 (solid red line). (F) Directionality (cos ϕ(t), see Materials and Methods) of vinculin FL and α-catenin KD cells. All data from 4 (all but YE) and 3 (YE) FOV (500-1000 tracks each) of 4 (all but YE and α-catenin KD) and 3 (YE, α-catenin KD) independent experiments. Mean ± SEM.

(A) Wound healing rate (see Materials and Methods) as a function of cell speed at confluence for WT, vinculin FL and mutant cells. Pearson coefficients r and p-values in black for all data, in blue minus YE. (B) Left: direct fluorescence of vinculin TSMod in the AJs plane of cells at the wound front (top) and back (bottom). Right: corresponding FRET Index maps. (C) FRET Index difference between front and back cells at AJs of TSMod in vinculin FL and mutants. (D) Left: direct fluorescence of vinculin TSMod in the FAs plane of cells at the wound front (top) and back (bottom). Right: corresponding FRET Index maps. (E) FRET Index difference between front and back cells at FAs of TSMod in vinculin FL and mutants. (F) Proposed model: at AJs, vinculin dimerization and actin binding provide mechanosensitivity to stable AJs, which promotes wound healing. Data of (A) from 4 (all but YE) and 3 (YE) FOV of 4 (all but YE) and 3 (YE) independent experiments. Data of (C) from 55 (FL), 16 (TL), 56 (T12) and 26 (YE) front cells and 122 (FL), 30 (TL), 71 (T12) and 48 (YE) back cells of 4 (FL, T12) and 3 (TL, YE) independent experiments. Data of (E) from 144 (FL), 63 (TL), 124 (T12) and 46 (YE) front cells and 155 (FL), 84 (TL), 128 (T12) and 51 (YE) back cells of 7 (FL, T12), 4 (TL) and 3 (YE) independent experiments. Mean ± SEM. Two-tailed Mann-Whitney test between front and back cells for each construct (C,E). Scale bars = 50µm.