title: Host proteolytic activity is necessary for infectious bursal disease virus capsid protein assembly.
abstract: In many viruses, a precursor particle, or procapsid, is assembled and undergoes massive chemical and physical modification to produce the infectious capsid. Capsid assembly and maturation are finely tuned processes in which viral and host factors participate. We show that the precursor of the VP2 capsid protein (pVP2) of the infectious bursal disease virus (IBDV), a double-stranded RNA virus, is processed at the C-terminal domain (CTD) by a host protease, the puromycin-sensitive aminopeptidase (PurSA). The pVP2 CTD (71 residues) has an important role in determining the various conformations of VP2 (441 residues) that build the T = 13 complex capsid. pVP2 CTD activity is controlled by co- and posttranslational proteolytic modifications of different targets by the VP4 viral protease and by VP2 itself to yield the mature VP2-441 species. Puromycin-sensitive aminopeptidase is responsible for the peptidase activity that cleaves the Arg-452-Arg-453 bond to generate the intermediate pVP2-452 polypeptide. A pVP2 R453A substitution abrogates PurSA activity. We used a baculovirus-based system to express the IBDV polyprotein in insect cells and found inefficient formation of virus-like particles similar to IBDV virions, which correlates with the absence of puromycin-sensitive aminopeptidase in these cells. Virus-like particle assembly was nonetheless rescued efficiently by coexpression of chicken PurSA or pVP2-452 protein. Silencing or pharmacological inhibition of puromycin-sensitive aminopeptidase activity in cell lines permissive for IBDV replication caused a major blockade in assembly and/or maturation of infectious IBDV particles, as virus yields were reduced markedly. PurSA activity is thus essential for IBDV replication.
authors.affiliation: Department of Molecular and Cellular Biology, Centro Nacional de Biotecnología/CSIC, Cantoblanco, 28049 Madrid, Spain.
authors.forename: Nerea
authors.initials: N
authors.lastname: Irigoyen
authors.forename: José R
authors.initials: JR
authors.lastname: Castón
authors.forename: José F
authors.initials: JF
authors.lastname: Rodríguez
mesh_terms: D000626:Aminopeptidases / Q000187:drug effects / Q000378:metabolism*; D000818:Animals; D036022:Capsid Proteins / Q000187:drug effects / Q000378:metabolism*; D002460:Cell Line; D004285:Dogs; D007243:Infectious bursal disease virus / Q000187:drug effects / Q000502:physiology*; D010447:Peptide Hydrolases / Q000187:drug effects / Q000378:metabolism*; D011691:Puromycin / Q000494:pharmacology; D012328:RNA Viruses / Q000187:drug effects / Q000502:physiology*; D012330:RNA, Double-Stranded / Q000235:genetics; D019065:Virus Assembly / Q000187:drug effects / Q000502:physiology*; D014779:Virus Replication / Q000187:drug effects / Q000502:physiology*
chemical_list: D036022:Capsid Proteins; D012330:RNA, Double-Stranded; D011691:Puromycin; D010447:Peptide Hydrolases; D000626:Aminopeptidases