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. 2019 Aug 15;21(10):945–962. doi: 10.1016/j.neo.2019.07.002

Figure 1.

Figure 1

SMAR1 regulates calnexin gene expression. (a) Semi-quantitative PCR to check the expression of calnexin, PDIA6 in HCT116 p53+/+ cells transfected with SMAR1 overexpression and sh-SMAR1 constructs. GAPDH was used as the loading control. (b) HEK293T cells were transfected with GFP-SMAR1 and sh-SMAR1 in a dose dependent manner. 2 μg, 4 μg and 6 μg of DNA was used for transfection. β-Actin was used as loading control (c) Western blotting of different cell lines transfected with GFP-vector, GFP-SMAR1 and sh-SMAR1 to check the expression profile of calnexin. Cell lines used were HCT116 p53+/+, HCT116 p53−/− and NIH3T3 cells. (d) Real-time PCR to check the gene regulation of calnexin by SMAR1. HCT116 p53+/+ cells were transfected with GFP-SMAR1 and treated with either 100 nM trichostatin A (TSA) or 1 mM sodium butyrate (NaB), the general inhibitors of histone deacytylase enzyme. (e) Luciferase assay was done in HEK293T cells transfected with pGL3-enhancer-calnexin promoter, Flag-SMAR1, and Flag-HDAC1. Cells were harvested after 48 h. Protein was extracted and luciferase assay was performed using the standard protocol from promega. (Bars represent standard deviation from three independent experiments. */** indicate statistically significant difference at P ≤ .05/0.005, respectively).