Figure 5.

Nuclear SOX7 induces granulosa cell apoptosis by binding to the Bax promoter

(A) Flow cytometric analysis of the proportion of apoptotic KGN cells (n = 6).

(B) Western blotting of BAX and BCL2 and their quantification (n = 3).

(C) Representative immunofluorescence images of SOX7 expression in KGN cells. Green represents SOX7 while blue represents DAPI.

(D) Western blotting was performed to quantify SOX7 expression in the nucleus and cytoplasm (n = 3).

(E) The proportion of apoptotic KGN cells was determined using Annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI) staining and flow cytometry (n = 4).

(F) Cell Counting Kit-8 was used to measure the relative cell viability (n = 5).

(G) Representative western blot analysis of BAX and BCL2 from the lysis solution of KGN cells (n = 3).

(H) Chromatin from KGN cells treated with or without cisplatin was collected for immunoprecipitation with SOX7 antibody. The Integrative Genomics Viewer (IGV) track of the input-subtracted SOX7 chromatin immunoprecipitation sequencing (ChIP-seq) signal at the promoter of the Bax gene (200–1,800 bp) is displayed.

(I) The effect of Sox7 overexpression on Bax transcriptional activity. KGN cells were co-transfected with pGL3-basic vector or pGL3-Bax, pUC-NC, or pUC-Sox7 along with the pRL-TK plasmid and cultured for 48 h. Luciferase activity was detected and normalized to Renilla (RL) activity (n = 4).

(J) KGN cells were transfected with pGL3-Bax and pUC-Sox7 for 48 h. KGN cells were then treated with or without β-RA for 6 h. Relative luciferase activity in each group (n = 4). The data are all presented as the mean ± SEM. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001; one-way ANOVA following the Dunnett’s multiple comparisons test in (A, B, D, I, and J); two-tailed Student’s t test in (E–G).