The role of initiating NTP concentrations in the regulation of influenza virus replication and transcription

Sir William Dunn School of Pathology, University of Oxford, Oxford, United Kingdom

	Abstract

The mechanisms regulating the synthesis of mRNA, cRNA and vRNA by the influenza A virus RNA-dependent RNA polymerase are not fully understood. Previous studies in our laboratory have shown that virion-derived viral ribonucleoprotein complexes synthesize both mRNA and cRNA, in vitro and early in the infection cycle in vivo. Our further studies showed that de novo synthesis of cRNA in vitro is more sensitive than capped primer-dependent synthesis of mRNA to the concentration of ATP, CTP and GTP. Using rescued recombinant influenza A/WSN/33 viruses, we now demonstrate that the 3' terminal sequence of the vRNA promoter dictates the requirement for a high NTP concentration during de novo initiated replication to cRNA, whereas this is not the case for extension of capped primers during transcription to mRNA. In contrast to some other viral polymerases for which only the initiating NTP is required at high concentrations, influenza virus polymerase requires high concentrations of the first three NTPs. In addition, we show that base-pair mutations in the vRNA promoter can lead to non-templated dead-end mutations during replication to cRNA in vivo. Based on our observations, we propose a new model for the de novo initiation of influenza virus replication.