Specific packaging of nodaviral RNA2 requires the N-terminus of the capsidprotein

Flock house virus (FHV), a member of the family Nodaviridae, is a nonenvelo ped, icosahedral insect virus whose capsids are assembled from 180 copies o f a single type of coat protein. The viral genome is split between two segm ents of single-stranded positive-sense RNA, RNA1 and RNA2, which are packag ed into a single virion. We previously demonstrated that synthesis of FHV c oat protein in the baculovirus expression system results in assembly of vir us-like particles whose capsids are indistinguishable from those of native virions, although the encapsidated RNA represents primarily cellular RNA. I n contrast, expression of a deletion mutant lacking N-terminal residues 2-3 1 results in formation of multiple types of particles which differ in size, shape, and RNA contents. We postulated that the polymorphism was imposed b y the type of RNA that the coat protein selected for packaging. In the curr ent study we tested this hypothesis by analyzing the assembly of the mutant coat protein in Drosophila cells in the presence of replicating FHV RNAs. As anticipated, the resulting particles had the same shape and dimensions a s wt virions. Surprisingly. however, they contained little RNA2 while packa ging of RNA1 was not affected. Small amounts of defective interfering RNAs, which emerged rapidly in the presence of the mutant coat protein, were als o detected. Taken together, these observations confirm our earlier hypothes is that selection of nonviral RNAs for packaging can significantly alter th e assembly process. In addition, they demonstrate that the N-terminus of th e FHV coat protein contains important determinants for recognition and pack aging of RNA2. Our results provide the first evidence that encapsidation of the two genomic RNAs occurs independently and that the coat protein uses d ifferent regions for the recognition of RNA1 and RNA2. (C) 2001 Academic Pr ess.