Traction stress is biphasically correlated with F-actin speed in FAs. (A) Traction stress versus F-actin speed for all points throughout the cell front over 25 frames of a time-lapse movie. (B) Subset of traction stress versus F-actin speed from A for data located within segmented FAs. Data were grouped by F-actin speed (greater or less than 10 nm/s) and three values of traction stress (<20, 20–50, and >50 Pa) to obtain six “stress-speed” groups identified by different colors/symbols. (C) Inverted GFP-paxillin image with spatial location of stress-speed data groups plotted in B (Video 4, available). Bar, 3 μm. (D, top) Montage of GFP-paxillin images of a single FA over 15 min (Video 5). For each time point, the integrated area (second row), GFP-paxillin intensity (third row), traction stress (fourth row), and local F-actin speed (bottom) were determined. In the fourth row, the dashed line indicates 75% of maximum stress; the arrow indicates time (t_s) when traction stress exceeds this threshold. (E) F-actin speed at t_s (v(t_s)) as a function of t_s. Mean of v(t_s) = 12.7 ± 3 nm/s. Mean of t_s = 3 ± 2 min. (F) v(t_s) as a function of traction stress at t_s, σ(t_s). Mean of σ(t_s) = 65 ± 28 Pa. Data in E and F are from 26 FAs in four cells.