title: A free VP3 C-terminus is essential for the replication of infectious bursal disease virus.
abstract: Green fluorescent protein (GFP) has been successfully incorporated into the viral-like particles of infectious bursal disease virus (IBDV) with a linker at the C-terminus of VP3 in a baculovirus system. However, when the same locus in segment A was used to express GFP by a reverse genetic (RG) system, no viable GFP-expressing IBDV was recovered. To elucidate the underlying mechanism, cDNA construct of segment A with only the linker sequence (9 amino acids) was applied to generate RG IBDV virus (rIBDV). Similarly, no rIBDV was recovered. Moreover, when the incubation after transfection was extended, wildtype rIBDV without the linker was recovered suggesting a free C-terminus of VP3 might be necessary for IBDV replication. On the other hand, rIBDV could be recovered when additional sequence (up to 40 nucleotides) were inserted at the 3' noncoding region (NCR) adjacent to the stop codon of VP3, suggesting that the burden of the linker sequence was not in the stretched genome size but the disruption of the VP3 function. Finally, when the stop codon of VP3 was deleted in segment A to extend the translation into the 3' NCR without introducing additional genomic sequence, no rIBDV was recovered. Our data suggest that a free VP3 C-terminus is essential for IBDV replication.
authors.affiliation: Department of Comparative Pathobiology, College of Veterinary Medicine, Purdue University, West Lafayette, IN 47907, USA.
authors.forename: Yung-Yi C
authors.initials: YC
authors.lastname: Mosley
authors.affiliation: School of Veterinary Medicine, National Taiwan University, Taipei 10617, Taiwan, ROC.
authors.forename: Ching Ching
authors.initials: CC
authors.lastname: Wu
authors.affiliation: Department of Comparative Pathobiology, College of Veterinary Medicine, Purdue University, West Lafayette, IN 47907, USA. Electronic address: tllin@purdue.edu.
authors.forename: Tsang Long
authors.initials: TL
authors.lastname: Lin
mesh_terms: D000818:Animals; D016367:Baculoviridae / Q000235:genetics / Q000378:metabolism; D002461:Cell Line, Transformed; D002645:Chickens; D003001:Cloning, Molecular; D005347:Fibroblasts / Q000821:virology; D015967:Gene Expression Regulation, Viral; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D007243:Infectious bursal disease virus / Q000235:genetics* / Q000378:metabolism; D000072417:Protein Domains; D015202:Protein Engineering / Q000379:methods; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D059386:Reverse Genetics / Q000379:methods*; D013329:Structure-Activity Relationship; D014162:Transfection; D015678:Viral Structural Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D014779:Virus Replication
chemical_list: D011994:Recombinant Proteins; C478244:VP3 protein, infectious bursal disease virus; D015678:Viral Structural Proteins; D049452:Green Fluorescent Proteins
keywords: GFP; IBDV replication; Infectious bursal disease virus; VP3 C-terminus