Fig. 3

TEX19 has nuclear and cytoplasmic localization in cancer cells: a Sub-confluent SW480 cells show TEX19 (red) localized to both nucleus and cytoplasm. Distinct cells show distinct proportions of cellular TEX19 within the nucleus. DNA is stained with DAPI (blue); bar = 40 μm. b Western blot analysis of nuclear and cytoplasmic extracts of sub-confluent SW480 cells indicates the majority of TEX19 is cytoplasmic, but a clear nuclear fraction is detected. Lamin (nucleus) and tubulin (cytoplasm) are used as controls (controls show clean faction signals, indicating no inter-fraction contamination). c Sub-confluent SW480 cells treated with the nuclear export blocking agent LMB demonstrates that TEX19 (red) can accumulate in the nucleus. DNA is stained by DAPI (blue); bar = 20 μm. d Western blot analysis of chromatin associated TEX19 demonstrates that TEX19 does not have a tight association with chromatin, even when cells are blocked in M phase with colcemid. Histone H3 is a control for tight chromatin association (only dissociated with 1.0 M NaCl); tubulin is mostly cytoplasmic (see b) and does not associate with chromatin during the chromatin preparation. e TEX19 is predominantly cytoplasmic at higher cell densities. Staining of sub-confluent HCT116 cells (top panels) indicates that TEX19 (red; blue = DAPI) is nuclear and cytoplasmic at this cell density, as for SW480 cells. When cells reach confluence (lower panels) the majority of TEX19 is cytoplasmic. Bar = 20 μm. f When SW480 cells are densely associated in spheres TEX19 (red; DAPI = blue) is mostly cytoplasmic. Bar = 100 μm. NTERA2 cells show strong nuclear staining with anti-TEX19 antibodies (green) irrespective of level of confluence. Red = anti-tubulin; Blue = DAPI; bar = 20 μm. g NTERA2 cells exhibit predominantly nuclear TEX19. Green = anti-TEX19; Blue = DAPI; Red = anti-tubulin; bar = 20 μm