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In previous studies we showed that a ninth polypeptide with a molecular weight of approximately 11,000 (NS2) found in influenza virus-infected cells was unique, that it could be synthesized in vitro, and that its expression in vivo required early protein synthesis. On the basis of these results we suggested that one of the eight genome RNA segments of influenza virus codes for two polypeptides [Lamb, R.A., Etkind, P.R. &amp; Choppin, P.W. (1978) Virology 91, 60-78]. We describe here differences in the electrophoretic mobility of the NS2 polypeptides of different strains of influenza A virus. These results provided further evidence that NS2 is virus coded and also made possible genetic studies using recombinants between two virus strains (HK and PR8) whose NS2 polypeptides differ. These studies showed that the gene for NS2 reassorts with that of the nonstructural polypeptide NS1, which is coded by genome segment 8. A mRNA for NS2 has been separated from that of NS1 and the other viral polypeptides by centrifugation and has been translated in vitro. Hybridization of genome segment 8 to the total mRNAs from infected cells specifically prevented the synthesis of NS2 and NS1. These results indicate that influenza virus genome segment 8 is transcribed into two separate mRNAs that code for two polypeptides, NS1 and NS2. Possible mechanisms for the transcription of the two mRNAs from either contiguous or overlapping genes are discussed.

Segment 8 of the influenza virus genome is unique in coding for two polypeptides.