Fig. 6: CFP1 promotes ovarian cancer cell proliferation by affecting BST2 transcription.

A Diagram showing the promoter regions of BST2, NOG, and S1PR1 genes. B Western blot results for the changes in the levels of the indicated proteins in ovarian cancer cells (A2780) treated with increasing concentrations of MLN4924 for 24 h. C qRT- PCR results of BST2 mRNA levels after siRNA depletion of ROC1 in A2780 cells. D qRT-PCR results for BST2 siRNA depletion efficiency. Total RNA extracted from A2780 WT, CFP1 KO siCon, and CFP1 KO siBST2 cells was subjected to qRT-PCR analysis for Bst2. Data are the mean ± SD of triplicated experiments. Student’s t-test was applied. **P < 0.01. E Effect of BST2 depletion on A2780 cell growth as assessed by CCK8 assay. Data are the mean ± SD of triplicated experiments. Student’s t-test was applied. *P < 0.05; **P < 0.01; ***P < 0.001. F Immunoblotting for BST2 overexpression (vector+BST2) efficiency on A2780 WT cells. G Effect of BST2 overexpression (V+BST2) on the growth of A2780 cells treated with MLN4924 as assessed by CCK8 assay. Data are the mean ± SD of triplicated experiments. Student’s t-test was applied. ***P < 0.001. H Immunoblotting for BST2 overexpression efficiency. Cell lysates (A2780 WT, CFP1 KO-vector, and A2780 CFP1 KO-BST2) were subjected to immunoblot analysis to verify the expression of BST2 protein. I Soft-agar assay for cell clone formation. The soft-agar assay was performed, and the colonies were stained with crystal violet for quantification, which is shown on the right panel. Data are the mean ± SD of three independent experiments. ***P < 0.001. J Mouse tumor-bearing model assay. A2780 WT, CFP1 KO-vector, and CFP1 KO-BST2 cells were implanted subcutaneously into nude mice. Two-way ANOVA test. *P < 0.05; ***P < 0.001. K CFP1 and H3K4me3 bind to the promoters of BST2. ChIP was performed with an antibody against CFP1 or H3K4me3 or with control IgG. The precipitated DNA was quantitated by qRT-PCR. Gapdh was included as the negative control. Data are the mean ± SD of quadruplicate experiments. L CRL4 E3 ubiquitin ligase affects histone methylation level in ovarian cancer by affecting CFP1 protein expression, thus regulating cell proliferation patterns.