Non-Lytic Egression of Infectious Bursal Disease Virus (IBDV) Particles from Infected Cells

Abstract

Infectious bursal disease virus (IBDV), a member of the Birnaviridae family, is responsible for a devastating immunosuppressive disease affecting juvenile domestic chickens. IBDV particles are naked icosahedrons enclosing a bipartite double-stranded RNA genome harboring three open reading frames (ORF). One of these ORFs codes for VP5, a non-structural polypeptide dispensable for virus replication in tissue culture but essential for IBDV pathogenesis. Using two previously described recombinant viruses, whose genomes differ in a single nucleotide, expressing or not the VP5 polypeptide, we have analyzed the role of this polypeptide during the IBDV replication process. Here, we show that VP5 is not involved in house-keeping steps of the virus replication cycle; i.e. genome transcription/replication, protein translation and virus assembly. Although infection with the VP5 expressing and non-expressing viruses rendered similar intracellular infective progeny yields, striking differences were detected on the ability of their progenies to exiting infected cells. Experimental data shows that the bulk of the VP5-expressing virus progeny efficiently egresses infected cells during the early phase of the infection, when viral metabolism is peaking and virus-induced cell death rates are as yet minimal, as determined by qPCR, radioactive protein labeling and quantitative real-time cell death analyses. In contrast, the release of the VP5-deficient virus progeny is significantly abridged and associated to cell death. Taken together, data presented in this report show that IBDV uses a previously undescribed VP5-dependent non-lytic egress mechanism significantly enhancing the virus dissemination speed. Ultrastructural analyses revealed that newly assembled IBDV virions associate to a vesicular network apparently facilitating their trafficking from virus assembly factories to the extracellular milieu, and that this association requires the expression of the VP5 polypeptide.

Identification and assessment of virulence of a natural reassortant of infectious bursal disease virus

“…In turn, the VP4 protein is a viral protease [ 14 ] that effects post-translational cleavage of the polyprotein (VP2–VP4–VP3) and takes part in the maturing of the VP2 peptide [ 15 ]. VP5 is a non-structural protein that cooperates in releasing of virus progeny from infected cells both through the non-lytic mechanism during the early phase of infection [ 16 ] and the activation of cell apoptosis in later stages by interacting with mitochondrial ionic channels (VDAC2) [ 17 ].…”

Ex vivo rescue of recombinant very virulent IBDV using a RNA polymerase II driven system and primary chicken bursal cells

“…Although we observed expression of VP3 protein, we anticipated that rescue of rCu-1 would be easier than rvv. As long as a few infectious particles of a cell-culture adapted IBDV such as rCu-1 are produced upon transfection, they can further replicate in DF-1 cells, increasing the infectious titer in the well 4,25 . On the contrary, in the case of rvv, a first artificial replication cycle may take place in DF-1 cells after transfection of the rescue vectors.…”

“…Typically, rescue of recombinants strains adapted to cell culture is performed by transfecting cell lines such as QM7 4 , Vero 25 or primary CEF 26 . Rescue of non-cell culture adapted recombinant viruses has been achieved by the transfer of supernatants and/or lysates from transfected cell lines to specific pathogen-free (SPF) chickens embryonated eggs—via the chorioallantoic membrane route 1 , 20 - or SPF chickens, which are inoculated by different routes such as intrabursal 27 , intramuscular routes 28 or by eye-drop 15 .…”

“…Segment A harbors two partially overlapping open reading frames (ORFs). The first one codes for VP5, a non-structural protein dispensable for virus replication which has been suggested to possess a crucial role in the cell-to-cell transmission of the virus 4 . The second one encodes a polyprotein, which is self-processed upon co-translational cleavage, yielding the capsid precursor protein pVP2, the multifunctional nucleoprotein VP3 and the viral protease VP4.…”

Chicken eEF1α is a Critical Factor for the Polymerase Complex Activity of Very Virulent Infectious Bursal Disease Virus

“…The larger segment A has two overlapping open reading frames (ORFs). The small ORF encodes the non-structural virus protein 5 (VP5; 17 kDa) involved in viral release and cell apoptosis induced by virus [15][16][17]. The large ORF in segment A encodes a polyprotein (109 kDa) that is processed into pVP2, VP3, and VP4.…”