Skip to main content
. 2010 Feb 24;285(19):14671–14680. doi: 10.1074/jbc.M109.066118

FIGURE 5.

FIGURE 5.

Knockdown of p35 affects cytoskeletal organization in senescent SAOS-2 and U2OS cells. A, SAOS-2 cells were seeded on coverslips and cotransfected with pSVE or pSVE-Rb and pLKO-p35shRNA-18 or control pLKO vector. 16 h after transfection, cells were selected with puromycin at a final concentration of 0.5 μg/ml. Samples were collected 10 days post-transfection and fixed with 4% formaldehyde. Immunofluorescence staining was performed using anti-α-tubulin antibody (DM1A) (upper panels) and rhodamine-phalloidin (lower panels), to show the structure of microtubules and F-actin filaments, respectively. Scale bars: 100 μm. B, immunofluorescence staining for F-actin (red) and Ezrin (green) in SAOS-2 cells cotransfected with pSVE-Rb and pLKO-p35shRNA-18 or control vectors following 8 days of selection. Scale bars: 100 μm. C, immunoblotting for actin and Ezrin in SAOS-2 cells transfected with pSVE-Rb and pLKO-p35shRNA-18 or control vectors following 5 days of selection. D, immunofluorescence staining for F-actin (red) and Ezrin (green) in U2OS cells cotransfected with CMV-p16 and pLKO-p35shRNA-17, -18, -20, or control vectors following 8 days of puromycin selection (2 μg/ml). Scale bars: 100 μm.