Infectious bursal disease virus (IBDV) is a double-stranded RNA (dsRNA) virus. Segment A contains two overlapping open reading frames (ORFs), which encode viral proteins VP2, VP3, VP4, and VP5. Segment B contains one ORF and encodes the viral RNA-dependent RNA polymerase, VP1. IBDV ribonucleoprotein complexes are composed of VP1, VP3, and dsRNA and play a critical role in mediating viral replication and transcription during the virus life cycle. In the present study, we identified a cellular factor, VDAC1, which was upregulated during IBDV infection and found to mediate IBDV polymerase activity. VDAC1 senses IBDV infection by interacting with viral proteins VP1 and VP3. This association is caused by RNA bridging, and all three proteins colocalize in the cytoplasm. Furthermore, small interfering RNA (siRNA)-mediated downregulation of VDAC1 resulted in a reduction in viral polymerase activity and a subsequent decrease in viral yield. Moreover, overexpression of VDAC1 enhanced IBDV polymerase activity. We also found that the viral protein VP3 can replace segment A to execute polymerase activity. A previous study showed that mutations in the C terminus of VP3 directly influence the formation of VP1-VP3 complexes. Our immunoprecipitation experiments demonstrated that protein-protein interactions between VDAC1 and VP3 and between VDAC1 and VP1 play a role in stabilizing the interaction between VP3 and VP1, further promoting IBDV polymerase activity.IMPORTANCE The cellular factor VDAC1 controls the entry and exit of mitochondrial metabolites and plays a pivotal role during intrinsic apoptosis by mediating the release of many apoptogenic molecules. Here we identify a novel role of VDAC1, showing that VDAC1 interacts with IBDV ribonucleoproteins (RNPs) and facilitates IBDV replication by enhancing IBDV polymerase activity through its ability to stabilize interactions in RNP complexes. To our knowledge, this is the first report that VDAC1 is specifically involved in regulating IBDV RNA polymerase activity, providing novel insight into virus-host interactions.
Autophagy functions as an intrinsic antiviral defense. However, some viruses can subvert or even enhance host autophagic machinery to increase viral replication and pathogenesis. The role of autophagy during avibirnavirus infection, especially late stage infection, remains unclear. In this study, infectious bursal disease virus (IBDV) was used to investigate the role of autophagy in avibirnavirus replication. We demonstrated IBDV induction of autophagy as a significant increase in puncta of LC3 ϩ autophagosomes, endogenous levels of LC3-II, and ultrastructural characteristics typical of autophagosomes during the late stage of infection. Induction of autophagy enhances IBDV replication, whereas inhibition of autophagy impairs viral replication. We also demonstrated that IBDV infection induced autophagosome-lysosome fusion, but without active degradation of their contents. Moreover, inhibition of fusion or of lysosomal hydrolysis activity significantly reduced viral replication, indicating that virions utilized the low-pH environment of acidic organelles to facilitate viral maturation. Using immuno-transmission electron microscopy (TEM), we observed that a large number of intact IBDV virions were arranged in a lattice surrounded by p62 proteins, some of which lay between virions. Additionally, many virions were encapsulated within the vesicular membranes, with an obvious release stage observed by TEM. The autophagic endosomal pathway facilitates low-pH-mediated maturation of viral proteins and membrane-mediated release of progeny virions.IMPORTANCE IBDV is the most extensively studied virus in terms of molecular characteristics and pathogenesis; however, mechanisms underlying the IBDV life cycle require further exploration. The present study demonstrated that autophagy enhances viral replication at the late stage of infection, and the autophagy pathway facilitates IBDV replication complex function and virus assembly, which is critical to completion of the virus life cycle. Moreover, the virus hijacks the autophagic vacuoles to mature in an acidic environment and release progeny virions in a membrane-mediated cellto-cell manner. This autophagic endosomal pathway is proposed as a new mechanism that facilitates IBDV maturation, release, and reinternalization. This report presents a concordance in exit strategies among some RNA and DNA viruses, which exploit autophagy pathway for their release from cells.
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