Tipping the balance between necrosis and apoptosis in human and murine cells treated with interferon and dsRNA

Abstract

Interferons enhance the cellular antiviral response by inducing expression of protective proteins. Many of these proteins are activated by dsRNA, a typical by-product of viral infection. Here we show that type-I and type-II interferons can sensitize cells to dsRNA-induced cytotoxicity. In caspase-8-or FADD-deficient Jurkat cells dsRNA induces necrosis, instead of apoptosis. In L929sA cells dsRNA-induced necrosis involves high reactive oxygen species production. The antioxidant butylated hydroxyanisole protects cells from necrosis, but shifts the response to apoptosis. Treatment with the caspase inhibitor benzyloxycarbonyl-Val-Ala-DLAsp(OMe)-fluoromethylketone or overexpression of Bcl-2 prevent this shift and promote necrosis. Our results suggest that a single stimulus can initiate different deathsignaling pathways, leading to either necrotic or apoptotic cell death. Inhibition of key events in these signaling pathways, such as caspase activation, cytochrome c release or mitochondrial reactive oxygen species production, tips the balance between necrosis and apoptosis, leading to dominance of one of these death programs.

Butylated hydroxyanisole is more than a reactive oxygen species scavenger

“…1,2 The same rationale led to the conclusion that ROS were also involved in dsRNA-induced necrosis of L929 cells. 3 Nevertheless, here we show that although BHA and BHT differ only slightly in their ability to reduce the levels of ROS induced by TNF stimulation of L929 cells, BHA is clearly more antinecrotic than BHT in this necrotic cell death system (Figure 1b). Both BHA and BHT reduce ROS levels and necrosis of L929 cells after stimulation with dsRNA ( Figure 1c).…”

“…However, BHA apparently does not block dsRNA-induced cytotoxicity, but shifts cell death from necrosis to apoptosis, whereas BHT does not modulate cell death type (Figure 1d). 3 Together, these data imply that BHA possesses additional properties that make it a more potent antinecrotic agent than BHT.…”

“…Interestingly, it had been shown that treatment of L929 cells with TNF leads to the activation of PLA 2 , whereas overexpression of Ca 2 þ -dependent PLA 2 (cPLA 2 ) sensitizes TNF-resistant L929 variants to TNF-induced cytotoxicity, 14,15 confirming that cPLA 2 contributes to this necrotic signaling. 3 followed by medium (white bars), 100 mM BHA (black bars) or 100 mM BHT (gray bars) for 30 min. Finally 2.5 mg/ml dsRNA (pIC) was added for the indicated durations.…”

The caspase-generated fragments of PKR cooperate to activate full-length PKR and inhibit translation

“…12 When the treated cells were deficient for FADD or caspase-8, or when treatment was carried out in the presence of the pancaspase inhibitor benzyloxycarbonyl-Val-Ala-DL-Asp(OMe)-fluoromethylketone (zVAD-fmk) cells responded by rapid necrosis. 12 Treatment of Jurkat E (JE) and JB6 Jurkat cells with dsRNA (100 mg/ml poly I:poly C), rapidly induced cell death (Figure 1). The morphology of dying JE cells treated with either anti-Fas or dsRNA was typically apoptotic (Figure 1a).…”

“…11 We have shown that in addition to increasing the capacity of cells to resist viral infection, type-I and type-II interferons can sensitize cells to external dsRNA-induced apoptosis and necrosis and demonstrated that dsRNA induced apoptosis but not necrosis requires FADD and caspase-8. 12 In addition to eIF2-a phosphorylation and inhibition of translation, PKR is involved in regulation of the activation of several transcription factors such as NF-kB, p53, or STATs.…”