Baculovirus P35 inhibits NO-induced apoptosis in activated macrophages by inhibiting cytochrome c release

The baculovirus protein P35 inhibits apoptosis in a diverse range of animals such as insects, nematodes and mammals. Evidence suggests that P35 can inhibit members of caspase family proteases that are key mediators of mammalian apoptosis. We demonstrate that p35 inhibits activation-induced nitric oxide (NO)-mediated apoptosis in the RAW 264.7 mouse macrophages. Parent or vector-transfected RAW 264.7 cells underwent apoptosis when treated with a combination of cisplatin and interferon- (IFN-) or LPS and IFN- in a NO-dependent manner. By contrast, RAW 264.7 cells stably expressing P35 did not undergo apoptosis when treated with a combination of cisplatin and IFN- or LPS and IFN-. Activation of parent, vector- or p35-transfected cells with cisplatin and IFN- or LPS and IFN- caused equivalent levels of inducible nitric oxide synthase (iNOS) expression and produced equal amounts of nitrite, which ruled out attenuated iNOS activity during P35-mediated protection. Rather, expression of P35 inhibited translocation of mitochondrial cytochrome c into cytosol, mitochondrial depolarization, activation of caspase-9 and caspase-3, and cleavage of poly(ADP-ribose)polymerase (PARP). These findings indicate that P35 inhibits NO-induced apoptotic cell death of activated macrophages by inhibiting mitochondrial cytochrome c release, which suggests that P35 has targets upstream of the caspase cascade in apoptosis.