Fig. 2: SMYD2 methylates CHMP2B K6 at the intercellular bridge of dividing cells.
From: Methylation of ESCRT-III components regulates the timing of cytokinetic abscission
A Immunodetection of CHMP2B K6 methylation on GFP_CHMP2B using CHMP2B K6me1 antibody in HeLa cells transfected with Flag_SMYD2 or SMYD2 Y240A and GFP_CHMP2B or CHMP2B K6A mutant as indicated on the figure (N = 3). B Protein extracts from HeLa cells treated with DMSO or 10 uM of BAY-598 (SMYD2 inhibitor) were immunoprecipitated using CHMP2B antibody and the CHMP2B K6 methylation was assessed by western blot using CHMP2B K6me1 antibody (N = 3). C Left panels: Representative cell images of CHMP2B K6me1 (green) and alpha-tubulin (magenta) immunostaining in HeLa cells treated with Ctrl or SMYD2 siRNAs. The white circle shows the midbody. Scale bar = 5 um. Right panel: quantification of the CHMP2B K6me1 signal at the midbody of HeLa cells treated with Ctrl or SMYD2 siRNAs as indicated (N = 3, n = 125 for siCtrl cells and n = 140 siSMYD2 cells midbodies quantified by condition, mean ± SD, Two-sided Mann–Whitney test). D Western blots of protein extracts from HeLa treated with control (siLuciferase referred to siCtrl) or SMYD2 siRNAs, revealed with SMYD2 and tubulin antibodies (N = 3). Loading controls: tubulin and Ponceau red staining. E Left panels: Representative cell images of CHMP2B K6me1 (green) and alpha-tubulin (magenta) immunostaining in HeLa cells treated with DMSO or 10 uM of BAY-598. The white circle shows the midbody. Scale bar = 5 um. Right panel: quantification of the CHMP2B K6me1 signal at the midbody of HeLa cells treated with Ctrl or SMYD2 siRNAs as indicated (N = 3, n = 121 for DMSO and n = 125 for BAY-598 midbodies quantified by condition, mean ± SD, Two-sided Mann–Whitney test).
