[21] Systems to express recombinant RNA molecules by the influenza A virus polymerase in vivo

Publisher Summary Viruses are obligate intracellular parasites that depend on cellular functions for their reproductive cycle. Two of the critical processes for viral reproduction are replication and transcription of the viral genome. The strategies developed by RNA viruses that require an RNA-dependent RNA polymerase activity to replicate and express their genomes are particularly interesting, because such an activity has not been detected in the rest of the living organisms, and therefore it has to be encoded for by the viral genome. The chapter focuses on recently described methodologies to study the strategy developed by influenza A virus to express and replicate its genome. The chapter presents a review in brief about the replication and transcription processes of the influenza virus genome and describes detailed procedures to express synthetic RNA molecules in influenza virus-infected cells and express functional polymerase in vivo from cloned complementary DNAs (cDNAs). Influenza A viruses are members of the Orthomyxoviridae family and contain a genome consisting of eight single-stranded RNA segments of negative polarity. The in vitro reconstitution of active ribonucleoproteins (RNPs) from synthesized viral RNA (vRNA) and the preparations of viral nucleoprotein and polymerase subunits have opened the possibility of analyzing the cis elements involved in influenza virus transcription and replication.