GJA1-20k does not decorate the sides of actin filaments and inhibits actin depolymerization. (A) F-actin (2 μM, 5% pyrene labeled) was depolymerized with the addition of 10 μM latrunculin A (LatA) in the presence (cross marks) or absence (circles) of 0.9 μM GJA1-20k. F-actin without LatA does not depolymerize (triangles). (B) In high-speed pelleting assays with F-actin (0–30 μM), the amount of GJA1-20k (0.7 μM) in supernatant fractions does not change as the concentration of F-actin is increased, which indicates the lack of side binding to actin filaments. Co-sedimentation results are shown as a gel (top) and densities which were quantified and plotted (bottom). The intensity of GJA1-20k band in the supernatant lacking actin was set as 1 (100%). Source data are available for this figure: SourceData FS1.