Figure 4. Force-dependent binding interactions between ternaryΔH1 complexes and F-actin.

(A) Optical trap setup used in constant- and low-force assays. Ternary GFP-E-cadherin cytoplasmic tail (green), β-catenin (yellow), and αE-cateninΔH1 (blue) complexes are immobilized on silica microspheres attached to a microscope coverslip. (Inset) The actin-binding domain (ABD), which confers the catch bond interaction between cadherin–catenin complexes and F-actin, is attached to the M domain of αE-catenin by a flexible linker. The four-helix H2–H5 binds to actin directly in the purported strong state conformation. (B) Representative trace from the constant-force assay. (C) Low-force assay. (Top) A representative force versus time series (gray). Plotted are the forces summed from both traps versus time, decimated from 40 to 4 kHz. Binding lifetimes at low force were defined by the duration during which the positional variance of trapped beads exceeded the baseline variance of control experiments. Traces colored in black are regions used for force baseline determination. When a binding event occurs, stage motion is translated to the trapped beads. (Bottom) The stage oscillates in a high-frequency, low-amplitude sinusoidal waveform to enable binding event detection at low forces.

Figure 4—figure supplement 1. E-cadherin/β-catenin/αE-cateninΔH1 low-force bond lifetimes.

(A) Binding event lifetimes measured below ~2.5 pN from the low-force assay (gray, N = 145) and constant-force assay (blue, N = 117). (B) Survival probability versus binding lifetime for low-force (LF) (N = 145) and constant-force (CF) data (N = 117) between 0 and 2.5 pN. The distribution of binding lifetimes between the two assays is not statistically different (p = 0.30 > 0.01, in a two-sample Kolmogorov–Smirnov [KS]).