Fig. 4: SIRT7-mediated PTEN deacetylation is associated with PTEN instability.
A–C PTEN protein level shown by immunoblots and quantification analysis of three independent repeats (n = 3). SIRT7 was knocked down (A), otherwise SIRT7 (B) or SIRT7-WT/H187Y (C) was overexpressed in HEC-1B cells. The relative expression level was analyzed in Image J, calculated by PTEN/Tubulin, and normalized by the control lane. The P value was calculated with two tailed unpaired t test. D The immunoblots showing the half-life of protein PTEN in HEC-1B cells with or without SIRT7 knockdown and the quantitative analysis of three independent repeats (n = 3). HEC-1B cells were treated with CHX (100 μg/ml) for indicated time and subjected for immunoblot analysis. HEC-1B shSIRT7 stable cell line was used and relative PTEN protein level was analyzed in Image J, calculated by PTEN/Tubulin, and normalized by the lane ‘0 h’. E Immunoblots showing the half-life of protein PTEN in HEC-1B cells over-expressing SIRT7-WT or SIRT7-H187Y and quantitative analysis (n = 3). F Immunoblots showing the half-life of protein PTEN-WT or PTEN-K260Q overexpressed in HEC-1B cells and quantitative analysis (n = 3). Two-way ANOVA was used for the P value calculation for (D–F). G Representative images of the mice uterus in immunofluorescence assay. The frozen sections of mice uterus tissue from control mice (Sirt7f/f) and Sirt7 uterus CKO mice (Sirt7f/f Pgr-cre) (n = 3) were stained with SIRT7, PTEN and DAPI. Scale bar = 50 μm. H Representative images of IHC staining of the uterus tissue from control mice (Sirt7f/f) and Sirt7 uterus CKO mice (Sirt7f/f Pgr-cre) (n = 5). The uterus tissue of the indicated mice was stained with SIRT7, PTEN, AcK260-PTEN antibodies. Scale bar = 50 μm. Data are presented as mean values ± s.e.m.
