Figure 3.
The PTP inhibitor-induced bypass of the G1/S checkpoint is associated with changes in p27 protein expression and cellular localization. HLF cells were treated with 0, 1 or 3 µM Cr in the presence and absence of 10 µM SOV for 24 h. Total cellular protein was extracted after the treatment. (A) Proteins were separated by SDS-PAGE and total p27 was detected by immunoblotting. The same amount of protein loading was confirmed by immunoblotting for β-actin. Data are the means ± SE of three independent experiments and are expressed as percentage of control, normalized to β-actin. *indicates a statistically significant difference from control (p < 0.05) and +indicates a statistically significant difference between the samples treated with and without SOV (p < 0.05). (B) HLF cells were seeded on chamber slides and, after treatment, were fixed and incubated with mouse p27 antibody and Alexa 488-conjugated goat anti-mouse secondary antibody. DNA was stained with PI to indicate the position of the nucleus. The numbers in each box represent the % of cells that have p27 nucleo-cytoplasmic localization. (C) Immunoblotting of the nuclear and cytoplasmic fractions for p27. α-Tubulin and Lamin A/C are loading controls for the cytoplasmic and nuclear fractions respectively. Data are mean ± SE of four experiments and are expressed as percentage of SOV 10 µM control. *indicates a statistically significant difference from control (p < 0.05).