Tween-20 treatment causes Ran to accumulate in the cytoplasm of HeLa cells and a block in nuclear export of proteins but not nuclear import of proteins.

<p>(A) Tween-20 causes cytoplasmic retention of Ran in HeLa cells. HeLa cells were incubated in fresh serum-free DMEM (Untreated) or in serum-free DMEM containing 150 µM Tween-20 for 4 h. After the 4 h incubation, the cells were left in Tween-20 containing medium (Tween-20), washed and placed in fresh serum-free media (Wash), or had the serum-free DMEM media containing Tween-20 supplemented with 10% FBS (Serum Add) and incubated at 37°C for 1 h. The distribution of Ran was monitored by immunofluorescence microscopy as described in materials and methods. (B) Tween-20 causes nuclear accumulation of Importin-α. HeLa cells were incubated in serum-free DMEM without (Untreated) or with 150 µM Tween-20 in serum-free DMEM for 4 h (Tween-20). Following the 4 h incubation, the cells were washed and incubated in serum-free DMEM for 1 h. The distribution of Importin-α was monitored by immunofluorescence microscopy. (C) Tween-20 causes nuclear accumulation of an NES-EGFP. HeLa cells were transfected with NES-EGFP and allowed to express for 24 h. Post-transfection cells were washed and placed in fresh serum-free DMEM with (Tween-20) or without (Untreated) 150 µM Tween-20 for 4 h. The distribution of NES-EGFP was monitored by direct fluorescent microscopy. (D) Tween-20 does not block nuclear import of the NLS containing Histone H2A. HeLa cells were incubated in serum-free DMEM without (Untreated) or with 150 µM Tween-20 for 4 h. The distribution of Histone H2A was monitored by immunofluorescence microscopy. (E) Tween-20 does not affect TNF-α stimulated nuclear import of NF-κB. HeLa cells were incubated in serum-free DMEM without (Untreated) or with 10 ng/ml TNF-α for 30 min, or with 150 µM Tween-20 for 4 h and then for 30 min with 10 ng/ml TNF-α. The distribution of NF-κB was monitored by immunofluorescence microscopy. The cells were DAPI stained to visualize the nucleus. Scale bar represents 10 µm.