Skip to main content
Fig. 5 | Cell & Bioscience

Fig. 5

From: Rho family GTPase 1 (RND1), a novel regulator of p53, enhances ferroptosis in glioblastoma

Fig. 5

RND1 physically interacted with p53 and inhibited p53 protein degradation. A The transcript levels of p53 remained unchanged under overexpression and down-regulation of RND1 circumstances. B Overexpression of RND1 promoted the stability of p53. After the cells were treated with CHX (50 μM) for different time periods (0, 2, 4, 6 h), p53 protein levels were analyzed by western blot. CHX, cycloheximide. C From western blot, proteasome inhibitor MG132 (10 μM, 6 h) blocked the regulation of p53 protein by RND1. D Immunofluorescent analysis of the co-localization of exogenous RND1 (labelled with Flag) and p53 (labelled with HA) proteins in U87 cells. Scale bar, 10 μm. E The colocalization between Flag-RND1 and HA-p53 has been quantified using the Colocalization Finder of Image J. Pearson’s Rr = 0.92; Overlap R = 0.93. F Co-IP pull-downs were conducted to investigate the association between RND1 and p53 at the exogenous level. 293T cells were co-transfected with Flag-RND1 and HA-p53 plasmids, which were later subjected to IP using anti-Flag, anti-HA antibodies, followed by immunoblotting with anti-RND1 and anti-p53 antibodies. G Co-IP pull-downs were conducted to investigate the association between RND1 and p53 at the endogenous level. H RND1 inhibited p53 ubiquitination in vivo. Proteins used for IP in vivo were extracted from tumor tissues of mouse xenografts. The loading proteins were precipitated by p53 antibody and immunoblotted with the anti-ub antibody. I RND1 inhibited the K48-linked polyubiquitin of p53 protein and stabilized p53. After 293T cells were transfected with HA-p53 and His-ub-WT, His-ub-K48R or His-ub-K63R, the cell lysates were subjected to IP using an anti-HA antibody, followed by immunoblotting with the anti-ub antibody. ns, no significance

Back to article page