Fig. 2

HPV16 regulation of SMAD4 expression. A, B HPV-positive HNC cell lines were transfected for 72 h with specific siRNA against HPV16 E6/E7 or Luciferase as control. After transfection cells were lysed and analysed by immunoblotting with the indicated antibodies (A) or total RNAs were isolated for RT-qPCR and reported as means ±SD of fold changes of at least three independent experiments (B). C, D HKs were transduced with empty or HPV16 E6/E7 or HPV11 E6/E7 recombinant retroviral vectors. After selection with G418 cells were harvested. C Lysates were collected and analysed by immunoblotting with the indicated antibodies. D Total RNAs were isolated for RT-qPCR. SMAD4 expression was normalized to RpP0. Results from at least three independent experiments are expressed as means ±SD. E HPV-positive cell lines were transfected with specific siRNA against ΔNp63α or Luciferase as control. 72 h after transfection cells were lysed and analysed by immunoblotting with the indicated antibodies. F, G HPV-positive HNC cell lines were transfected with either specific siRNA against p53 (F) or pCMV expressing p53 (G). Cells were the lysed and analysed by immunoblotting with the indicated antibodies