Figure 3
From: The syntaxin-binding protein STXBP5 regulates progerin expression
Influence of STXBP5 overexpression on HA-progerin expression in HEK293 cells. (A) qRT-PCR revealed that the STXBP5 transcript level was increased in HA-progerin-expressing HEK293 cells. The empty vector was transfected into WT HEK293 cells as a control. (B) Western blot analysis revealed that, compared with that in control OE cells, the translation of STXBP5 was increased in HA-progerin-expressing HEK293 cells. The translation of STXBP5 was evaluated with an anti-STXBP5 antibody, with normalized to \(\beta\)-Actin expression. (C) An anti-STXBP5 antibody was used to quantify STXBP5 overexpression in HA-progerin-expressing HEK293 cells. The data were normalized to \(\beta\)-Actin antibody expression. (D) Protein level of SRSF1 were measured by anti-SRSF1 antibody respectively, then graphed. (E) qRT-PCR revealed that the level of STXBP5 was increased in HEK293 cells overexpressing HA-progerin + FLAG-STXBP5, Over-vector-FLAG was transfected into HA-progerin-expressing HEK293 cells as the control OE condition. (F) The level of HA-tagged progerin was measured by western blotting with an anti-HA antibody. (H) The LaminA/C expression level was measured by western blotting with an anti-Lamin A/C antibody, and a graph was generated. (G, I) The mRNA expression levels of LaminA, progerin, LaminB1, LaminC, P53, P21, IL6 and IL8 in HA-progerin + FLAG-STXBP5 OE HEK293 cells were measured via qRT-PCR. (J) Summary of the quantification of SA-\(\beta\)-gal staining in STXBP5-transfected progerin-HA-expressing HEK293 cells from three independent experiments. Scale bars, 50\(\mu\)m. (K) Protein expression was analyse via western blotting with an anti-Lamin B1 antibody. \(\beta\)-Actin was used as a loading control. The orange arrowheads indicate the Lamin A and Lamin C bands. The data are presented as the means ± SEMs. Comparisons were performed with one way ANOVA (D, K) or two-way ANOVA (G, I) with Brown-Forsythe test for multiple comparisons. two-tailed Student’s t test (A, B, E, F, H, J). ns, p > 0.05; *, p < 0.05; **, p < 0.01; ***, p < 0.001; ****, p < 0.0001.
