The maturation process of pVP2 requires assembly of infectious bursal disease virus capsids.
Infectious bursal disease virus (IBDV) is a nonenveloped avian virus with a two-segment double-stranded RNA genome. Its T=13 icosahedral capsid is most probably assembled with 780 subunits of VP2 and 600 copies of VP3 and has a diameter of about 60 nm. VP1, the RNA-dependent RNA polymerase, resides inside the viral particle. Using a baculovirus expression system, we first observed that expression of the pVP2-VP4-VP3 polyprotein encoded by the genomic segment IBDA results mainly in the formation of tubules with a diameter of about 50 nm and composed of pVP2, the precursor of VP2. Very few virus-like particles (VLPs) and VP4 tubules with a diameter of about 25 nm were also identified. The inefficiency of VLP assembly was further investigated by expression of additional IBDA-derived constructs. Expression of pVP2 without any other polyprotein components results in the formation of isometric particles with a diameter of about 30 nm. VLPs were observed mainly when a large exogeneous polypeptide sequence (the green fluorescent protein sequence) was fused to the VP3 C-terminal domain. Large numbers of VLPs were visualized by electron microscopy, and single particles were shown to be fluorescent by standard and confocal microscopy analysis. Moreover, the final maturation process converting pVP2 into the VP2 mature form was observed on generated VLPs. We therefore conclude that the correct scaffolding of the VP3 can be artificially induced to promote the formation of VLPs and that the final processing of pVP2 to VP2 is controlled by this particular assembly. To our knowledge, this is the first report of the engineering of a morphogenesis switch to control a particular type of capsid protein assembly.
Chevalier
Christophe
C
Unité de Virologie et Immunologie Moléculaires, Institut National de la Recherche Agronomique, F-78350 Jouy-en-Josas, France.
Lepault
Jean
J
Erk
Inge
I
Da Costa
Bruno
B
Delmas
Bernard
B
D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D002213:Capsid / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D049452:Green Fluorescent Proteins; D007243:Infectious bursal disease virus / Q000235:genetics / Q000378:metabolism*; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008856:Microscopy, Fluorescence; D011498:Protein Precursors / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D018411:Spodoptera; D015678:Viral Structural Proteins / Q000235:genetics / Q000378:metabolism*; D014771:Virion / Q000378:metabolism; D019065:Virus Assembly
D008164:Luminescent Proteins; D011498:Protein Precursors; D011993:Recombinant Fusion Proteins; C082513:VP2 protein, infectious bursal disease virus; D015678:Viral Structural Proteins; D049452:Green Fluorescent Proteins