Fig. 6: miR-663 controls PIN-1-mediated apoptosis and p53 mitochondrial function

a Immunoblot showing BTG2 downmodulation 72 h after siRNA transfection. TUBULIN was used as protein loading control. Normalized BTG2 levels are expressed as fold decrease relative to cntrl treated cells. b Western blotting showing BTG2 modulation in NIH-H460 after the indicated treatments. Double transfection of anti-BTG2 siRNA together with LNA-663 restores BTG2 levels of control cells, counteracting LNA-mediated induction. PARP-1 cleavage is reduced to 60%. TUBULIN was used as protein loading control. Numbers indicate BTG2 levels fold change relative to cntrl treated cells, and cleaved PARP-1 levels fold decrease relative to LNA-663/cntrl siRNA-treated cells. The values were normalized to TUBULIN signal, as measured by densitometric analyses. c Subcellular BTG2 and PIN-1 localization in NIH-H460 cells treated with control LNA or LNA-663 was assessed by immunofluorescence staining. d Mitochondrial membrane potential of NIH-H460 cells upon miR-663 downmodulation was measured by JC-1 staining. An increase of the ratio between green and red signals is indicative of membrane depolarization typical of apoptotic cells. e Western blotting showing PIN-1 modulation in NIH-H460 subcellular fractions 24 h after LNA-663 treatment. Cytoplasmic accumulation and nuclear decrease of PIN-1 are expressed as fold changes relative to cntrl-treated cells. The histogram shows the ratio between cytoplasmic and nuclear signal. f Immunoblot of p53 in NIH-H460 subcellular fractions. Hsp90 and Bcl-2 were used as cytoplasmic and mitochondrial markers, respectively. Mitochondrial p53 modulation is expressed as fold increase relative to cntrl treated cells and normalized to Bcl-2 signal