PROTEIN-PROTEIN INTERACTIONS AND GLYCEROPHOSPHOLIPIDS IN BROMOVIRUS AND NODAVIRUS RNA REPLICATION

The plant bromoviruses and animal nodaviruses are distinct groups of p ositive strand RNA viruses that have proven to be useful models for RN A replication studies. Bromoviruses encode two large proteins required for RNA replication: 1a contains domains implicated in helicase and c apping functions, and 2a contains a central polymerase-like domain. Us ing immunoprecipitation and far-western blotting, we have now shown th at 1a and 2a form a specific complex in vitro and have mapped the inte racting domains. Molecular genetic data implicate the 1a-2a complex in RNA replication and suggest that it supports coordinate action of the putative helicase, polymerase, and capping domains. The locations of the interacting 1a and 2a domains have implications for replication mo dels and the evolution of virus genomes bearing homologous replication genes in fused vs. divided forms. For the nodavirus Flock house virus (FHV), a true RNA replicase has been isolated that carries out comple te, highly active replication of added FHV RNA, producing newly synthe sized positive strand RNA in predominantly ssRNA form. Positive strand RNA synthesis in this FHV cell-free system is strongly dependent on t he addition of any of several glycerophospholipids. Positive strand RN A synthesis depends on the complete glycerophospholipid structure, inc luding the polar head group and diacyl glycerol lipid portion, and is strongly influenced by acyl chain length.