(A) Effects of K18-targeting siRNAs on K18 expression. MCF10A cells were transfected with control or K18-targeting siRNAs at the indicated concentrations of siRNAs and cultured for 48 h. Cell lysates were analyzed by immunoblotting with an anti-K18 antibody. (B) Ventral images of endogenous β4, their binary images, and bright field images of control and K18 knockdown MCF10A cells. Cells were seeded as shown in Fig 2, transfected with control or K18-targeting siRNAs, and cultured for 48 h. The red dotted lines indicate the total adhesion area defined by bright field images. Scale bar, 20 μm. (C) Quantitative analysis of the effect of K18 knockdown on HD formation. The ratio of HD area to total adhesion area was calculated, as in Fig 2. Data represent the means ± SD of 3 or 4 independent experiments (at least 7 images per experiment). ****P < 0.0001 (one-way ANOVA followed by Dunnett's test).