FIG. 8.
E7 diminishes TGF-β-induced p27Kip1 nuclear localization. (A) tet-E7 Mv1Lu cells were treated for 24 h with 2 μg of doxycycline (Dox) or vehicle per ml. Cell lysates were analyzed by Western blotting with antibody specific for HPV-16 E7. (B) tet-E7 Mv1Lu cells were treated for 24 h with 3 ng of TGF-β/ml in the presence or absence of 2 μg of doxycycline/ml and subsequently analyzed for p27Kip1 expression via Western blot. (C and D) tet-E7 Mv1Lu cells were treated as described for panel B, fixed with 3.7% paraformaldehyde, and stained with p27Kip1 specific antibody as described in Materials and Methods. (C) Representative fluorescent (left) and phase-contrast (right) micrographs of TGF-β-treated cells in the absence (top) or presence (bottom) of doxycycline. (D) Quantitation of cells exhibiting nuclear localization of p27Kip1. The number of cells with nuclear p27Kip1 accumulation is represented as a percentage of total cells counted. The average and deviation values shown are from two independent experiments with at least 200 cells counted per experiment.