Flock House Virus Induces Apoptosis by Depletion of Drosophila Inhibitor-of-Apoptosis Protein DIAP1

Abstract

The molecular mechanisms by which RNA viruses induce apoptosis and apoptosis-associated pathology are not fully understood. Here we show that flock house virus (FHV), one of the simplest RNA viruses (family, Nodaviridae), induces robust apoptosis of permissive Drosophila Line-1 (DL-1) cells. To define the pathway by which FHV triggers apoptosis in this model invertebrate system, we investigated the potential role of Drosophila apoptotic effectors during infection. Suggesting the involvement of host caspases, the pancaspase inhibitor benzyloxycarbonyl-Val-Ala-Asp-fluromethylketone (z-VAD-fmk) prevented FHV-induced cytopathology and prolonged cell survival. RNA interference-mediated ablation of the principal Drosophila effector caspase DrICE or its upstream initiator caspase DRONC prevented FHV-induced apoptosis and demonstrated direct participation of this intrinsic caspase pathway. Prior to the FHV-induced activation of DrICE, the intracellular level of inhibitor-of-apoptosis (IAP) protein DIAP1, the principal caspase regulator in Drosophila melanogaster, was dramatically reduced. DIAP1 was depleted despite z-VAD-fmk-mediated caspase inhibition during infection, suggesting that the loss of DIAP1 was caused by an upstream FHV-induced signal. The RNA interferencemediated knockdown of DIAP1 caused rapid and uniform apoptosis of DL-1 cells and thus indicated that DIAP1 depletion is sufficient to trigger apoptosis. Confirming this conclusion, the elevation of intracellular DIAP1 levels in stable diap1-transfected cells blocked caspase activation and prevented FHV-induced apoptosis. Collectively, our findings suggest that DIAP1 is a critical sensor of virus infection, which upon virus-signaled depletion relieves caspase inhibition, which subsequently executes apoptotic death. Thus, our study supports the hypothesis that altering the level or the activity of cellular IAP proteins is a general mechanism by which RNA viruses trigger apoptosis.

Protection of Insects against Viral Infection by Apoptosis-Dependent Phagocytosis

“…2D). Apoptosis caused by FHV infection is accompanied by a decrease in DIAP1 (51). DIAP1 was detectable as a protein band of 57 kDa in the lysates of uninfected S2 cells, as originally reported (45), and we observed a decrease in this caspase inhibitor after DCV infection (third panel, Fig.…”

“…Target specificity in this reaction is due to the occurrence of apoptosis in cells infected with virus. In general, cells induced to undergo apoptosis become susceptible to phagocytosis by surface presentation of a molecule(s) that is recognized by an engulfment receptor of phagocytes (51)(52)(53). DCV infection was found to induce caspase-mediated apoptosis in Drosophila S2 cells and was accompanied by several biochemical changes often observed in mammalian cells undergoing apoptosis.…”

“…Therefore, a decrease in the level of DIAP1 is an indication of the onset of apoptosis in Drosophila cells. In fact, Settles and Friesen (51) claimed that a decreased level of DIAP1 is solely responsible for the induction of apoptosis in Drosophila Line-1 cells upon infection with FHV. We made a similar observation: DIAP1 decreased in S2 cells after DCV infection.…”

“…A previous article reported the induction of apoptosis in Drosophila Line-1 cells upon infection with flock house virus (FHV), another RNA virus belonging to Nodaviridae (51). Thus, we examined possible biochemical changes in S2 cells, which are often observed during apoptosis.…”

Oocyte destruction is activated during viral infection

“…An attractive mechanism for the induction of egg chamber destruction by FHV and DCV infection is suggested by the finding that FHV infection leads to the downregulation of Drosophila inhibitor of apoptosis 1 (DIAP1), rendering cells susceptible to apoptotic death (Settles and Friesen, 2008). DIAP1 has been shown to have a developmentally regulated role during oogenesis (Foley and Cooley, 1998).…”