Fig. S8.

FHL2 nuclear localization driven by substrate rigidity is dependent on FAK. (A) SDS/PAGE with polyacrylamide containing Mn²⁺ and Phos-tag (Top). SDS/PAGE without Mn²⁺ and Phos-tag (Bottom). Western blots for FHL2-GFP or FHL2_Y93F-GFP from each sample isolated from HFF cells on the soft substrate or the hard substrate. Lane 1, FHL2-GFP from HFF cells on soft substrate; lane 2, FHL2_Y93F-GFP from HFF cells on soft substrate; lane 3, FHL2-GFP from HFF cells on hard substrate; lane 4, FHL2_Y93F-GFP from HFF cells on hard substrate. Black arrowhead, phosphorylated FHL2 protein; white arrowhead, FHL2 protein. (B) FHL2-GFP localization in an HFF cell on soft substrate. (B′) FAK-mCherry localization in an HFF cell on soft substrate. (B′′) Merged image of B and B′. (C) FHL2 localization in an FAK^−/− cell on 8.78-kPa ACA gel coated with fibronectin. (C′) Merged image of C and DAPI. (D) FHL2 localization in an FAK ^−/− cell on 75.3-kPa ACA gel coated with fibronectin. (D′) Merged image of D and DAPI. (E) Graph showing the intensity ratio of FHL2 immunofluorescence staining between the nucleus and whole cell area (nucleus/whole cell area) on fibronectin-coated gels of varying rigidity. n > 20. Error bars represent SEM. All images are projected images from adhesion sections to nuclear sections.