Infectious bursal disease virus capsid protein VP3 interacts both with VP1, the RNA-dependent RNA polymerase, and with viral double-stranded RNA.
Infectious bursal disease virus (IBDV) is a double-stranded RNA (dsRNA) virus of the Birnaviridae family. Its two genome segments are encapsidated together with multiple copies of the viral RNA-dependent RNA polymerase, VP1, in a single-shell capsid that is composed of VP2 and VP3. In this study we identified the domains responsible for the interaction between VP3 and VP1. Using the yeast two-hybrid system we found that VP1 binds to VP3 through an internal domain, while VP3 interacts with VP1 solely by its carboxy-terminal 10 amino acids. These results were confirmed by using a reverse-genetics system that allowed us to analyze the interaction of carboxy-terminally truncated VP3 molecules with VP1 in infected cells. Coimmunoprecipitations with VP1- and VP3-specific antibodies revealed that the interaction is extremely sensitive to truncation of VP3. The mere deletion of the C-terminal residue reduced coprecipitation almost completely and also fully abolished production of infectious virions. Surprisingly, these experiments additionally revealed that VP3 also binds to RNA. RNase treatments and reverse transcription-PCR analyses of the immunoprecipitates demonstrated that VP3 interacts with dsRNA of both viral genome segments. This interaction is not mediated by the carboxy-terminal domain of VP3 since C-terminal truncations of 1, 5, or 10 residues did not prevent formation of the VP3-dsRNA complexes. VP3 seems to be the key organizer of birnavirus structure, as it maintains critical interactions with all components of the viral particle: itself, VP2, VP1, and the two genomic dsRNAs.
Tacken
Mirriam G J
MG
Division of Infectious Diseases and Food Chain Quality, Institute for Animal Science and Health (ID-Lelystad B.V.), NL-8200 AB Lelystad, The Netherlands. M.G.J.Tacken@id.wag-ur.nl
Peeters
Ben P H
BP
Thomas
Adri A M
AA
Rottier
Peter J M
PJ
Boot
Hein J
HJ
D000818:Animals; D001665:Binding Sites; D002213:Capsid / Q000737:chemistry / Q000378:metabolism*; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D007243:Infectious bursal disease virus / Q000378:metabolism* / Q000472:pathogenicity; D016296:Mutagenesis; D010940:Plant Proteins; D011233:Precipitin Tests; D012330:RNA, Double-Stranded / Q000378:metabolism*; D012324:RNA-Dependent RNA Polymerase / Q000378:metabolism*; D015534:Trans-Activators; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D020798:Two-Hybrid System Techniques
D004268:DNA-Binding Proteins; D010940:Plant Proteins; D012330:RNA, Double-Stranded; D015534:Trans-Activators; D014157:Transcription Factors; D012324:RNA-Dependent RNA Polymerase