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. 2015 Oct 26;211(2):391–405. doi: 10.1083/jcb.201503081

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© 2015 Chiassson-MacKenzie et al.

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Figure 3. — Cell junctions in Nf2^−/− LDCs are under increased mechanical stress. (A) Confocal images showing Nf2^−/− or Nf2^WT-expressing LDCs at early (left) and late (right) stages of confluence; cells were labeled with an anti–β-catenin antibody (red) and with phalloidin to detect F-actin (green). (B) Junctional localization of β-catenin was quantified by fluorescence intensity analysis of AJs in early and late confluent Nf2^−/− and Nf2^WT-expressing LDCs. (C) Junctional localization of F-actin was quantified by fluorescence intensity analysis of AJs in late confluent Nf2^−/− and Nf2^WT-expressing LDCs. (D) Confocal images showing endogenous vinculin at AJs in Nf2^−/− and Nf2^WT-expressing LDCs. (E) The amount of vinculin at AJs in Nf2^−/− and Nf2^WT-expressing LDCs was quantified by measuring the junctional area stained for vinculin. (F) Confocal images showing endogenous β-catenin (red) and/or F-actin (green) in Nf2^−/− LDCs treated with 100-µM blebbistatin or vehicle (DMSO). (G) Junctional localization of β-catenin in DMSO- and blebbistatin-treated confluent Nf2^−/− LDCs was quantified by fluorescence intensity analysis of AJs. (H) Junctional localization of β-catenin in Nf2^−/− LDCs cultured on stiff (40 kPa) and soft (12 kPa) polyacrylamide hydrogels. (B, C, E, G, and H) Error bars indicate SEM (n = 25 junctions per group). *, P < 0.05; ***, P < 0.001. Data are representative of at least three experiments. Bars, 10 µm. a.u., arbitrary units.