Fig. 5.

Substratum rigidity regulates the abundance and localization of p120-catenin and YAP. (A) p120-catenin abundance in CDCs cultured on MRS in spheroids, and remaining adherent cells. (B) Immunostaining for p120-catenin and p190RhoGAP in CDCs cultured on glass (left) or on MRS (right). Colocalization analysis is shown in fig. S4E. (C) p120-catenin abundance in CDCs transfected with control scrambled small interfering RNAs (siRNAs) (scr^KD) or siRNAs directed against p190RhoGAP (p190^KD). (D) Time course analysis of p120-catenin abundance in CDCs cultured on MRS. (E) Flow cytometric analysis of CD31 in scr^KD and YAP^KD CDCs. n = 2 cultures. (F) Immunostaining reveals that YAP has a predominantly nuclear localization in CDCs cultured on glass and a cytosolic distribution in CDCs cultured on MRS. (G) Quantification of nuclear localization in (F). n = 3 samples. (H) Analysis of p190RhoGAP, YAP, and p120-catenin abundance in cell monolayers and spheroid cell aggregates cultured on MRS. (I) Two-photon imaging of YAP in a CDC spheroid. (J) Quantification of nuclear localization of YAP. (K) Immunoblotting analysis of YAP in scr^KD and GATA2^KD CDCs. (L) p190RhoGAP abundance in CDCs cultured on MRS for 8 days followed by 2 days on glass, or cells cultured on MRS and glass for 10 days. Scale bars, 20 μm (B and F) and 100 μm (I). P values determined by paired t tests, **P < 0.01; ***P < 0.001. Error bars, SEM.