Figure 4.

Expression of a SH3 domain-deleted mutant of vinexin increases the population of cells at midbody stage. (A) Illustration of various deletion mutants of vinexin β. (B) Thymidine-arrested cells were transfected with the plasmid expressing EGFP or EGFP-vinexin wild type (WT) or ΔSH3 mutants and then released for 14 h before α-tubulin immunostaining and Hoechst labeling. Quantification of green fluorescence intensity at the midbody stage from 20 cells expressing each EGFP construct. (C, D) Immunostaining of vinculin with (C) α-tubulin or (D) Vinexin at the midbody of HeLa cells. (E) HeLa cells infected with a lentivirus expressing control (siCtrl) or a shRNA against vinculin (siVCL) were harvested for immunoblotting. (F) Thymidine-arrested siCtrl and siVCL cells were released to fresh medium for 12 h, followed by immunostaining of vinexin and α-tubulin. The fluorescence intensity of vinexin at the midbody of HeLa cells was quantified from 20 cells per group. (G) Similar to (B), the images from 3 independent experiments were used to calculate the distribution of cells at the midbody stage and interphase. Arrows denote intercellular bridges. Scales, 10 μm. Data are mean ± SEM. ** P < 0.01, * P < 0.05 by Student t test.