Fig. 3.

PBF alters the subcellular localisation of NIS. (A) (i) Immunofluorescent detection of NIS-HA and endogenous PBF in cells transfected with NIS-HA and VO control. (ii) PBF overexpression is associated with an increase in NIS staining within intracellular vesicles. Scale bars: 20 μM. (B) Cell-surface biotinylation assay in COS-7 cells transfected with a VO control or with human PBF. (i) Representative immunoblot analysis of surface-biotinylated polypeptides precipitated with streptavidin-agarose beads and probed with anti-HA antibody, which revealed HA-tagged human NIS detection at around 100 kDa. Flotillin-1 was used as a marker of membrane protein expression to determine loading between samples. (ii) Total NIS-HA protein expression. Graphs indicate mean differences in cell-surface and total expression of NIS (n=3 separate scanning densitometry experiments were performed). ^*P<0.05; ns, not significant.