Interactions in vivo between the proteins of infectious bursal disease virus: capsid protein VP3 interacts with the RNA-dependent RNA polymerase, VP1

Institute for Animal Science and Health (ID-Lelystad), Department of Avian Virology, PO Box 65, NL-8200 AB Lelystad, The Netherlands 1 University of Utrecht, Department of Infection and Immunity, Utrecht, The Netherlands 2 Author for correspondence: Mirriam Tacken. Fax +31 320 238 668. e-mail m.g.j.tacken{at}id.wag-ur.nl Little is known about the intermolecular interactions between the viral proteins of infectious bursal disease virus (IBDV). By using the yeast two-hybrid system, which allows the detection of protein–protein interactions in vivo , all possible interactions were tested by fusing the viral proteins to the LexA DNA-binding domain and the B42 transactivation domain. A heterologous interaction between VP1 and VP3, and homologous interactions of pVP2, VP3, VP5 and possibly VP1, were found by co-expression of the fusion proteins in Saccharomyces cerevisiae . The presence of the VP1–VP3 complex in IBDV-infected cells was confirmed by co-immunoprecipitation studies. Kinetic analyses showed that the complex of VP1 and VP3 is formed in the cytoplasm and eventually is released into the cell-culture medium, indicating that VP1–VP3 complexes are present in mature virions. In IBDV-infected cells, VP1 was present in two forms of 90 and 95 kDa. Whereas VP3 initially interacted with both the 90 and 95 kDa proteins, later it interacted exclusively with the 95 kDa protein both in infected cells and in the culture supernatant. These results suggest that the VP1–VP3 complex is involved in replication and packaging of the IBDV genome.