Replication and Transcription of the Rotavirus Genome

Rotaviruses are an important cause of human morbidity and mortality, representing the primary pathogens responsible for acute dehydrating diarrhea in children under the age of 3. The infectious rotavirus particle is made up of three concentric layers of protein, and contains a genome consisting of eleven segments of double-stranded (ds)RNA. Upon infection, RNA polymerases associated with double-layered virus particles are activated, resulting in genome transcription and extrusion of the eleven viral mRNAs from such particles. The mRNAs not only direct protein synthesis, but also serve as templates for minus-strand synthesis to yield dsRNAs. Synthesis of the dsRNAs is an event that occurs following the gene-specific packaging of viral mRNAs into core-like assembly intermediates. Electron-dense cytoplasmic inclusions, termed viroplasms, function as sites of genome packaging and replication in the infected cell. Our understanding of key events in the viral life cycle has been advanced considerably by the development of cell-free systems that support mRNA synthesis from virion-derived double-layered particles and dsRNA synthesis from virion-derived core particles. The recent expression and purification of rotavirus recombinant proteins have also allowed progress to be made in defining the roles of viral proteins in genome replication and viroplasm formation. However, our efforts towards a full description of the viral life cycle, most notably an understanding of the events occurring during gene-specific packaging, remain hampered by the lack of a cell-free packaging system and a reverse genetics systems. The lack of a reverse genetics systems also confounds efforts towards the generation of molecular engineered second-generation vaccines.