Release of bluetongue virus-like particles from insect cells is mediated by BTV nonstructural protein NS3/NS3A.

Recombinant baculoviruses and immunoelectron microscopy have been used to demonstrate the association of virus-like particles (VLPs) of bluetongue virus (i.e., particles composed of BTV, VP2, VP3, VP5, and VP7 proteins) and the cell cytoskeleton, as well as the release of such particles from infected cells in the presence of BTV NS3/NS3A, but not when BTV NS1 protein was coexpressed. Examination of cells infected with recombinants that express core-like particles (CLPs) composed of VP3 and VP7 showed that CLPs were present within the soluble fraction of the cells and not associated with the cell cytoskeleton. The simultaneous expression of the nonstructural NS1 or NS3/NS3A proteins with CLPs did not lead to the association of such particles with the cytoskeleton, nor to their release from cells. The failure of CLPs synthesized in the presence of VP2 or VP5 to attach to the cytoskeleton indicated that both outer coat proteins are required for a stable virus-cytoskeleton interaction. The ultrastructural, immunoelectron microscopical, and biochemical examinations, showed conclusively that the presence of NS3/NS3A is required for the budding and subsequent release of VLPs from infected cells.