Polypeptides specified by the influenza virus genome 2. Assignment of protein coding functions to individual genome segments by in vitro translation

Abstract

Cytoplasmic RNA extracted from chick embryo fibroblasts infected with influenza A (fowl plague) virus (FPV) was translated in a wheat germ cell-free protein-synthesizing system. Polypeptides which comigrated during SDS-polyacrylamide gel electrophoresis with marker virus-specific polypeptides P 1, P 2, P3, NP, M, and NS were synthesized in vitro. The NP, M, and NS polypeptides were positively identified by tryptic peptide mapping. The polypeptide component of the virus glycoprotein HA was also synthesized in vitro, and was identified by tryptic peptide mapping. RNA extracted from purified FPV (vRNA) did not direct the synthesis of any recognizable virus-specific polypeptides in vitro, either as a total preparation, or as individual RNA genome segments. The protein coding functions of the vRNA segments were identified by hybridization of individual segments to a preparation of infected cell cytoplasmic RNA. On subsequent translation of the RNA in vitro, synthesis of the virus-specific polypeptide corresponding to the hybridized vRNA segment was specifically reduced. We conclude that, for FPV, virion RNA segments 1–3 code for the three P polypeptides and segments 4, 5, 6, 7, and 8 code for polypeptides HA, NP, NA, M, and NS, respectively.