Extended Data Fig. 8: NONO ligands induce accumulation of NONO and the paralogous proteins PSPC1 and SFPQ into nuclear foci.

a, Immunofluorescence data showing time-dependent effects of active compound 5 and inactive control 8 (20 µM) on the localization of NONO in nuclear foci (yellow wedge) in 22Rv1 cells. NONO was imaged with primary antibody (Bethyl Laboratories, A300-587A) and an Alexa-488 secondary antibody. Scale bar: 20 µm. b, c, NONO foci induced by compounds 5 (b) and (R)-SKBG-1 (c) localize proximal to the nucleolar marker fibrillarin (20 µM compound treatment, 24 h). NONO was imaged with primary antibody (BD Biosciences, 611278) and an Alexa-488 secondary antibody. Fibrillarin was imaged with primary antibody (Cell Signaling, 2639 S) and an Alexa-647 secondary. Scale bar: 20 µm. d, e, Zoomed-in merged images of NONO (green) and fibrillarin (red) localization from compounds 5 (d) or (R)-SKBG-1 (e) treated 22Rv1 cells (24 h time point). DAPI stain, blue. Scale bar: 10 µm. f, g, Immunofluorescence data showing effects of active compounds 5 (f) and (R)-SKBG-1 (g) and inactive controls 8 and (S)-SKBG-1 on the localization of SFPQ in nuclear foci (yellow wedge) in 22Rv1 cells (20 µM of each compound, 24 h). NONO was imaged with primary antibody (Bethyl Laboratories, A300-587A) and an Alexa-488 secondary antibody. SFPQ was imaged with primary antibody (Sigma-Aldrich, WH0006421M2) and an Alexa-647 secondary. Scale bar: 20 µm. h, Immunofluorescence data showing effects of active compounds 5 and (R)-SKBG-1 and inactive controls 8 and (S)-SKBG-1 on the localization of PSPC1 in nuclear foci (yellow wedge) in 22Rv1 cells (20 µM of each compound, 24 h). NONO was imaged with primary antibody (Bethyl Laboratories, A300-587A) and an Alexa-488 secondary antibody. PSPC1 was imaged with primary antibody (Sigma-Aldrich, SAB4200503) and an Alexa-647 secondary. Scale bar: 20 µm.