MEK-1 inhibition sensitizes leukemic cells to ATO-induced apoptosis. (A) NB4 and K562 cell lines were seeded at 1 × 105 in the presence of DMSO (vehicle) or PD98059 for 3 hours and then incubated for 72 hours with the indicated concentration of ATO. Apoptosis was then measured as percentage of cells with hypodiploid DNA content. Results are representative of one of 3 independent experiments. (B) Total RNAs extracted from leukemic cells treated for 24 hours were reverse transcribed using the specific primers for ΔNp73 and TAp73. NB4 (C) and K562 (D) cell lines were seeded at 1 × 105 in the presence of DMSO (vehicle), PD98059 (40 μM), or PD184352 (1 μM) for 3 hours, and then incubated for 24 hours with the indicated concentrations of ATO. Endogenous p73α and ΔNp73 proteins were revealed by immunoblotting analysis using a mouse monoclonal anti-p73 (clone 1288; Imgenex, San Diego, CA), or a mouse monoclonal anti-ΔNp73 (clone 38C674; Imgenex). Antiactin (Santa Cruz Biotechnology, Santa Cruz, CA) immunoblotting was performed as loading control. β-actin, ΔN-p73, and TA-p73 bands were subjected to densitometric scanning using TINA 2 software (Raytest Isotopenmessgerate, Germany) and the ΔNp73/β-actin or TAp73/β-actin ratio was calculated. AS = ATO.
