Interaction of structural core protein of classical swine fever virus with endoplasmic reticulum-associated degradation pathway protein OS9

• We identified cellular OS9 as a specific binding partner for the CSFV core protein.
• OS9 specifically binds only full length unprocessed CSFV Core protein.
• The OS9 binding site in the core protein was identified.
• Mutation of the binding site, caused decreased replication in swine macrophages.

Classical swine fever virus (CSFV) Core protein is involved in virus RNA protection, transcription regulation and virus virulence. To discover additional Core protein functions a yeast two-hybrid system was used to identify host proteins that interact with Core. Among the identified host proteins, the osteosarcoma amplified 9 protein (OS9) was further studied. Using alanine scanning mutagenesis, the OS9 binding site in the CSFV Core protein was identified, between Core residues 90IAIM93, near a putative cleavage site. Truncated versions of Core were used to show that OS9 binds a polypeptide representing the 12 C-terminal Core residues. Cells transfected with a double-fluorescent labeled Core construct demonstrated that co-localization of OS9 and Core occurred only on unprocessed forms of Core protein. A recombinant CSFV containing Core protein where residues 90IAIM93 were substituted by alanines showed no altered virulence in swine, but a significant decreased ability to replicate in cell cultures.