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. 2012 Apr 5;3(4):e292. doi: 10.1038/cddis.2012.32

Figure 5.

Figure 5

Figure 5

rNef-mediated cytoplasmic accumulation of eEF1A in BFA-treated MDMs inhibits caspase activation and decreases the cytoplasmic release of cytochrome c. (a) Inhibition of caspase-3 activation in BFA-stimulated MDMs treated with rNef (1000 ng/ml). (b) Inhibition of caspase-3 activation in BFA-stimulated MDMs treated with rNef is dose-dependent and positively correlates with cytoplasmic accumulation of eEF1A. (c) Knockdown of Exp-1 and Exp-5 proteins by siRNA in MDMs. MDM cultures were transfected with scrambled control, Exp-1 siRNA or Exp-5 siRNA. Total cellular extracts were prepared 48 h post-transfection. Protein expression was analyzed by western blot. β-Actin was used as a loading control. (d) Inhibition of caspase-3 activation in BFA-stimulated MDMs treated with rNef is dependent on Exp-t. (e) Mitochondrial cytochrome c release in BFA-treated MDMs is blocked by rNef. (f) Mitochondrial cytochrome c release in BFA-treated MDMs is blocked by rNef in a dose-dependent manner and positively correlates with cytoplasmic accumulation of eEF1A. (g) Inhibition of caspase-9 activation in BFA-stimulated MDMs treated with rNef (1000 ng/ml). Protein levels of caspase-9 were quantified by densitometry using ImageJ 1.40 software (the level of caspase-9 in mock cells was arbitrarily established at 1). (h) Inhibition of caspase-9 activation in BFA-stimulated MDMs treated with rNef is dose-dependent. Protein levels of caspase-9 were quantified by densitometry using ImageJ 1.40 software (the level of caspase-9 in mock cells was arbitrarily established at 1). (i) Inhibition of caspase activation in BFA-stimulated MDMs treated with rNef, but neither in TM-stimulated MDMs nor in TG-stimulated MDMs treated with rNef. Left panel, Time course of caspase activation in BFA-stimulated MDMs, TM-stimulated MDMs, or TG-stimulated MDMs. Right panel, Inhibition of caspase activation in BFA-stimulated MDMs treated with rNef, but neither in TM-stimulated MDMs nor in TG-stimulated MDMs treated with rNef. MDM cultures were treated with BFA (10 μg/ml), TM (10 μg/ml) or TG (10 μg/ml) for 12h in the presence of rNef (1 μg/ml), and caspase-3 and caspase-9 activation was measured in total cellular lysates. Results are representative of data obtained in three independent experiments