title: Overlap of interaction domains indicates a central role of the P protein in assembly and regulation of the Borna disease virus polymerase complex.
abstract: The active polymerase complex of Borna disease virus is composed of the viral proteins N, P, and L. The viral X (negative regulatory factor) protein acts as a regulator of polymerase activity. Interactions of P with N and X were previously studied, but interactions with L were poorly defined. Using a mammalian two-hybrid system, we observed that L specifically interacts with P but not with N, X, or itself. Mapping of the L-binding domain in the P molecule revealed that it overlaps with two adjacent domains required for multimerization and interaction with N. Competition experiments showed that the interaction between L and P was inefficient when N was present, indicating that L may preferentially interact with free P in infected cells. Interestingly, a multimerization-defective P mutant maintained the ability to interact with L, N, and X but failed to support reporter gene expression from an artificial Borna disease virus minigenome. Furthermore, dominant negative effects on minigenome activity were only observed when P mutants with an intact multimerization domain were used, suggesting that P multimers, rather than monomers, exhibit biological activity. P mutants lacking functional interaction domains for L or N still formed complexes with these viral proteins when wild-type P was available as a bridging molecule, indicating that P multimers have the potential to act as scaffolds on which the RNA polymerase complex is assembled.
authors.affiliation: Department of Virology, University of Freiburg, Hermann-Herder-Strasse 11, D-79104 Freiburg, Germany.
authors.forename: Urs
authors.initials: U
authors.lastname: Schneider
authors.forename: Kareen
authors.initials: K
authors.lastname: Blechschmidt
authors.forename: Martin
authors.initials: M
authors.lastname: Schwemmle
authors.forename: Peter
authors.initials: P
authors.lastname: Staeheli
mesh_terms: D001667:Binding, Competitive; D001891:Borna disease virus / Q000201:enzymology*; D002460:Cell Line; D002850:Chromatography, Gel; D018076:DNA, Complementary / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000737:chemistry*; D005799:Genes, Dominant; D006801:Humans; D009154:Mutation; D010750:Phosphoproteins; D010957:Plasmids / Q000378:metabolism; D017354:Point Mutation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D020798:Two-Hybrid System Techniques; D014764:Viral Proteins / Q000737:chemistry*; D015678:Viral Structural Proteins / Q000737:chemistry* / Q000502:physiology
chemical_list: D018076:DNA, Complementary; C496537:P protein, Borna disease virus; D010750:Phosphoproteins; D014764:Viral Proteins; D015678:Viral Structural Proteins; D012321:DNA-Directed RNA Polymerases