Fig. 3: Ligand density influences molecular-clutch kinetics and myosin II organization.

a,b, Cell traction measured on MEF WT cells (a) and MEF WT and MEF Talin 1 KO, Talin 2 shRNA cells (b) plated on substrates coated with increasing concentrations of fibronectin¹³. The experimental data were fitted to the talin WT model (dotted curves) and the linear WT and KD models (solid curves) by varying the rate of molecular-clutch formation (\({k}_{{\rm{on}}}^{0}\)), the characteristic size of adhesion sites of molecular clutches (r) and the density of myosin II motors (σ_m). Other model parameters were held fixed (Supplementary Table 1). c, The best-fitting parameter values as a function of fibronectin density show that enriching the ligand density accelerates molecular-clutch formation, reduces the characteristic size of adhesion sites of molecular clutches and increases the density of myosin II, thereby enhancing actin–substrate force transmission. d, Structured illumination microscopy image of an NIH-3T3 fibroblast expressing myosin RLC-GFP. Inset: close-up of the boxed region. e,f, Quantification of myosin II filament stacks in NIH-3T3 and HeLa cells. Increasing the fibronectin density did not alter the density of myosin II stacks (e) but did alter their projected area in a cell-type-dependent manner (f). The response of NIH-3T3 cells is consistent with model predictions for MEF WT cells. In e from left to right, N = 13, 11, 14, 13, 12 and 16 cells were processed under the corresponding experimental conditions. In f from left to right, N = 2,534, 846, 3,836, 834, 2,305 and 928 stacks of myosin II filaments were collected and processed under the corresponding experimental conditions from the same number of cells as indicated for e. Boxes and error bars in e represent mean ± standard errors of the mean (s.e.m.) and standard deviation, respectively. The circles in f show the median, and boxes represent the mean ± s.e.m. The pairwise statistical difference between the corresponding datasets was calculated using one-way ANOVA with post-hoc Tukey’s honestly significant difference test (no adjustments).