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. 2021 Aug 12;10:e62592. doi: 10.7554/eLife.62592

Figure 2. SREBP1 mediates induction of 6PGD downstream of the androgen receptor (AR).

Figure 2.

(A) ChIP-seq data showing AR DNA binding near the 6PGD gene in non-malignant and prostate tumour samples (Pomerantz et al., 2015) and the LNCaP (Barfeld et al., 2017) and VCaP (Asangani et al., 2014) cell line models. The grey box indicates a region ±50 kb of the 6PGD transcriptional start site. (B) ChIP-seq data showing SREBP1 DNA binding at the 6PGD promoter in HEPG2 and MCF7 cells. Data is from ENCODE (ENCODE Project Consortium, 2012; HEPG2: ENCFF000XXR; MCF7: ENCFF911YFI). (C) Effect of siSREBP1 on 6PGD protein. LNCaP cells were transfected with siRNA (siSREBP1; 12.5 nM) or control (siCon) for 72 hr after which SREBP1 and 6PGD protein levels were evaluated by immunoblotting. GAPDH was used as loading control. (D) Effect of siSREBP1 on 6PGD induction by DHT. LNCaP (left) or VCaP (right) cells were transfected with siRNA (siSREBP1; 12.5 nM) or control (siCon) in charcoal-stripped FBS media for 72 hr and then treated with 10 nM DHT for another 24 hr. SREBP1 and 6PGD protein levels were evaluated by immunoblotting. GAPDH was used as loading control. (E) RT-qPCR of 6PGD expression in response to DHT and fatostatin in LNCaP cells. Cells were serum starved in charcoal-stripped FBS media for 72 hr and then treated with Veh or 10 nM DHT ±10 µM fatostatin for another 24 hr. Gene expression was normalised to GUSB and L19 and represents the mean + SEM of three biological replicates. Treatment effects were evaluated using ANOVA and Dunnett’s multiple comparison tests (*p<0.05; **p<0.01; ****p<0.0001; NS, not significant).