Analysis of the Contribution of Hemocytes and Autophagy to Drosophila Antiviral Immunity

Abstract

Antiviral immunity in the model organism Drosophila melanogaster involves the broadly active intrinsic mechanism of RNA interference (RNAi) and virus-specific inducible responses. Here, using a panel of six viruses, we investigated the role of hemocytes and autophagy in the control of viral infections. Injection of latex beads to saturate phagocytosis, or genetic depletion of hemocytes, resulted in decreased survival and increased viral titers following infection with Cricket paralysis virus (CrPV), Flock House virus (FHV), and vesicular stomatitis virus (VSV) but had no impact on Drosophila C virus (DCV), Sindbis virus (SINV), and Invertebrate iridescent virus 6 (IIV6) infection. In the cases of CrPV and FHV, apoptosis was induced in infected cells, which were phagocytosed by hemocytes. In contrast, VSV did not trigger any significant apoptosis but we confirmed that the autophagy gene Atg7 was required for full virus resistance, suggesting that hemocytes use autophagy to recognize the virus. However, this recognition does not depend on the Toll-7 receptor. Autophagy had no impact on DCV, CrPV, SINV, or IIV6 infection and was required for replication of the sixth virus, FHV. Even in the case of VSV, the increases in titers were modest in Atg7 mutant flies, suggesting that autophagy does not play a major role in antiviral immunity in Drosophila. Altogether, our results indicate that, while autophagy plays a minor role, phagocytosis contributes to virus-specific immune responses in insects. IMPORTANCEPhagocytosis and autophagy are two cellular processes that involve lysosomal degradation and participate in Drosophila immunity. Using a panel of RNA and DNA viruses, we have addressed the contribution of phagocytosis and autophagy in the control of viral infections in this model organism. We show that, while autophagy plays a minor role, phagocytosis contributes to virusspecific immune responses in Drosophila. This work brings to the front a novel facet of antiviral host defense in insects, which may have relevance in the control of virus transmission by vector insects or in the resistance of beneficial insects to viral pathogens.

Defense Mechanisms against Viral Infection in <em>Drosophila</em>: RNAi versus Non-RNAi

“…As already mentioned (section 3.2.2), RNAi screens have identified a role for autophagy and Toll-7 in the antiviral defense against viruses with a negative strand RNA genome such as VSV and RVFV [24,99,128]. Another study focused on the analysis of fly mutants of the autophagy gene Atg7 and confirmed a role for autophagy in the control of VSV although the effects were considered mild [179]. By contrast, VSV infection levels were not affected in Toll-7 mutants, in contrast to other studies [24,128].…”

“…As already indicated above, comparison of transcriptome data has also revealed their poor reproducibility, even if infections were carried out with the same virus. As discussed by Marques and Imler (2016), this could be caused by (among others) differences in the experimental system (cell lines versus adult flies) and infection routes (systemic versus oral), shortcomings of experimental procedures (for instance, off-target effects in RNAi), heterogeneity in genetic background of Drosophila strains, polymorphisms in host restriction factors and the unknown occurrence of persistent infections (other viruses, Wolbachia). Infection by natural (Drosophilaspecific viruses) and non-natural pathogens (arboviruses) are expected to give different responses because of (absence of) co-evolution of the pathogen with the host.…”

“…The majority of blood cells constitute the macrophage-like plasmatocytes (90-95% of hemocytes in Drosophila) that are specialized in the engulfment and degradation of cellular debris, debris and invading pathogens [178]. The involvement of cellular immunity in antiviral defense was dramatically demonstrated in experiments of genetic ablation of hemocytes or inhibition of their phagocytotic capacity by injection of polystyrene or latex beads [142,179]. Virus-specific effects were observed since cellular immunity was required for resistance against CrPV, VSV and FHV but not against SINV or IIV-6.…”

“…Virus-specific effects were observed since cellular immunity was required for resistance against CrPV, VSV and FHV but not against SINV or IIV-6. In the case of DCV, phagocytosis was required to control infection at high doses [180] but was dispensable at low doses [179]. During CrPV infection, hemocytes become depleted which is associated with the viral dose and progression of infection [142].…”

“…The requirement for phagocytosis seems to be correlated with the induction of apoptosis during viral infection. CrPV, DCV and FHV trigger apoptosis in the S2 cell line and apoptotic bodies can be subsequently phagocytosed by plasmatocytes from adult flies [179] or l(2)mbn cells, a Drosophila larval hemocyte-derived cell line [180]. In the latter study of DCV infection, it was shown that phagocytosis preferentially targets apoptotic cells and that it involved the recognition of specific features of apoptotic cells (e.g.…”