Fig 1: KO and inhibition of ARF1 abolish ERK1/2 activation by OR51E2. (A) Effect of CRISPR-Cas9-mediated depletion of ARF1 on ERK1/2 activation by ß-ionone in DU145 cells. The cells were transiently transfected with CRISPR-Cas9 KO plasmids targeting human ARF1, starved for 48 h and then stimulated with ß-ionone at 100 µM for 5 min. (B) Effect of ARF1 inhibitors on ERK1/2 activation by ß-ionone in DU145 cells. The cells were starved for 48 h and treated with secinH3 (100 µM), GCA (30 µM) or Exo2 (60 µM) for 30 min before stimulation with ß-ionone. AS-604850 treatment (2.5 µM for 6 h) was used as a positive control. The Western blots shown in each panel are representatives of at least three experiments.
Fig 2: Stimulation with ß-ionone recruits ARF1 to the GA and activates ARF1 via Gß? and PI3K? in prostate cancer cells. (A) Subcellular distribution of ARF1Q71L and ARF1T31N in DU145 and LNCaP cells. The cells were transfected with GFP-tagged ARF1 active or dominant negative mutant for 24 h and the subcellular distribution of ARF1 mutants was revealed by confocal microscopy. (B) ARF1 translocation to the GA after ß-ionone stimulation. The cells were starved for 48 h, stimulated with ß-ionone at 100 µM for 5 min and then stained with antibodies against ARF1 and p230. (C) Quantification of ARF1 expression at the GA by using p230 as a GA marker. (D) Quantification of Pearson’s coefficient between ARF1 and p230. (E) ARF1 activation in G?3, G?9, and p110? KO cells in response to ß-ionone stimulation. DU145 KO cells were starved for 48 h before stimulation with ß-ionone at 100 µM for 5 min. ARF1 activation was measured in GST fusion protein pulldown assays. The quantitative data are presented as means ± SE (n = 29–43 cells in C and 18–25 cells in D in three individual experiments). ***, p < 0.001 versus respective control. Scale bar: 10 µm. The Western blots shown are representatives of three experiments.
Supplier Page from Abcam for Anti-ARF1 antibody