Fig. 2

PPFIA4 is an AR-repressed gene. A. Pearson correlation of mRNA expression between PPFIA4 and AR-targeted gene PSA. Transcriptomic data from TCGA and GSE35988 clinical prostate cancer cohort was used to perform correlation analysis. B. Schematic of three putative AR binding sites on the PPFIA4 gene promoter. According to motif analysis in JASPAR database, primers were designed as P1 (-1838 to -1822), P2 (-1387 to -1371) and P3 (-786 to -770) based on their distances to the transcription start site (TSS) of PPFIA4 gene. C. ChIP analysis was conducted in LNCaP and C4-2B cells to validate AR enrichment on PPFIA4 promoter. Purified rabbit IgG was used as negative control. Primers flanking the AR binding site on the PSA gene promoter were used as positive controls. ****p < 0.0001 based on the Student’s t-test. D-E. Luciferase reporter assays were performed in PC3 cells transfected with empty vector or PPFIA4 overexpression plasmid (D) and in LNCaP or C4-2B cells transfected with negative control (siNC) or AR siRNA (siAR). Luciferase reporters containing either wild-type or AR binding sites deletion in PPFIA4 promoter were constructed. Luciferase activities were measured 48 hours after transfection. **p < 0.01 based on the Student’s t-test. NS, no significance