MASTL regulates cell spreading independently of focal adhesion size, integrin activity, or cell cycle. (A) Representative images of F-actin (Phalloidin-Atto), paxillin, and DAPI staining in siControl and siMASTL (48 h silencing) MCF10A cells plated on collagen for 2 h. Images were acquired on a 3i CSU-W1 spinning disk confocal. (B–D) Quantification of cell area based on F-actin staining, 45 cells/condition in total (B), focal adhesion size based on paxillin staining (C), and focal adhesion count based on paxillin relative to individual cells or total cell area (D), with 47 (siControl) or 44 (siMASTL) cells in total (three independent experiments) from A. (E and F) Representative flow cytometry histograms of total β1-integrin (P5D2; E) or active β1-integrin (12G10; F) in siControl and siMASTL MDA-MB-231 cells. (G) Quantification of active β1 integrin (12G10) levels relative to total β1 integrin (P5D2) from E and F. (H) Quantification of integrin binding to labeled fibronectin (FN-647) relative to total β1 integrin (P5D2) analyzed with flow cytometry. (I) Cell cycle profiles of siControl and siMASTL MDA-MB-231 cells. Data in all graphs are from n = 3 biologically independent experiments (unpaired t test, mean ± SD). See also Fig. S2 and Table S1.
Figure 2.

MASTL regulates cell spreading independently of focal adhesion size, integrin activity, or cell cycle. (A) Representative images of F-actin (Phalloidin-Atto), paxillin, and DAPI staining in siControl and siMASTL (48 h silencing) MCF10A cells plated on collagen for 2 h. Images were acquired on a 3i CSU-W1 spinning disk confocal. (B–D) Quantification of cell area based on F-actin staining, 45 cells/condition in total (B), focal adhesion size based on paxillin staining (C), and focal adhesion count based on paxillin relative to individual cells or total cell area (D), with 47 (siControl) or 44 (siMASTL) cells in total (three independent experiments) from A. (E and F) Representative flow cytometry histograms of total β1-integrin (P5D2; E) or active β1-integrin (12G10; F) in siControl and siMASTL MDA-MB-231 cells. (G) Quantification of active β1 integrin (12G10) levels relative to total β1 integrin (P5D2) from E and F. (H) Quantification of integrin binding to labeled fibronectin (FN-647) relative to total β1 integrin (P5D2) analyzed with flow cytometry. (I) Cell cycle profiles of siControl and siMASTL MDA-MB-231 cells. Data in all graphs are from n = 3 biologically independent experiments (unpaired t test, mean ± SD). See also Fig. S2 and Table S1.

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