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. 2013 Sep 16;202(6):849–859. doi: 10.1083/jcb.201302131

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© 2013 Choi et al.

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Figure 5. — FAK-H58A restores decreased pY397 and impaired cell spreading in FAK−/− MEF cells with NHE1 shRNA. (A) Immunoblot indicates that abundance of pY397 with GFP-tagged FAK WT-HA and H75A-HA but not H58A-HA are markedly less with NHE1 shRNA compared with NT shRNA. Immunoblot with HA antibody shows expression of heterologously expressed FAK. (B) Abundance of pY397 relative to total FAK WT and mutants in cells from A. Data are means ± SEM of three cell preparations. Asterisks indicate significant difference of P < 0.001. (C) Impaired cell spreading and focal adhesion morphology in FAK−/− MEFs with NT shRNA, visualized by vinculin immunolabeling, are restored by expression of GFP-tagged FAK-WT, FAK-H58A, and FAK-H75A, but not GFP alone. (D) Impaired cell spreading and focal adhesion morphology in FAK−/− MEFs with NHE1 shRNA are restored by expression of FAK-H58A but not FAK-WT or FAK-H75A. (E) With NHE1 shRNA only H58A rescues spreading cell morphology scored by roundness. Data are from 75–100 cells analyzed in two cell preparations. Asterisks indicate significant difference of P < 0.001.