POLIOVIRUS POLYURIDYLIC ACID POLYMERASE AND RNA REPLICASE HAVE THE SAME VIRAL POLYPEPTIDE

A poliovirus-specific poly(U) polymerase that copies a poly(A) template complexed to an oligouridylic acid primer was isolated from the membrane fraction of infected HeLa [human cervical carcinoma] cells and sedimented at 4-5S on a linear 5-20% glycerol gradient. When the poly(U) polymerase was isolated from cells labeled with [35S]methionine and analyzed by glycerol gradient centrifugation and polyacrylamide gel electrophoresis, the position of only 1 viral protein correlated with the location of enzyme activity. This protein had an apparent MW of 62,500 based on its electrophoretic mobility and was designated p63. When the poly(U) polymerase was isolated from the soluble fraction of a cytoplasmic extract, the activity sedimented at about 7S. In this case, p63 and NCVP2 (77,000-dalton precursor of p63) co-sedimented with the 7S activity peak. When the 7S polymerase activity was purified by phosphocellulose chromatography, p63 and NCVP2 co-chromatographed with poly(U) polymerase activity. The poliovirus replicase complexed with its endogenous RNA template was isolated from infected cells labeled with [35S]methionine and was centrifuged through a linear 15-30% glycerol gradient. The major viral polypeptide component in a 26S peak of replicase activity was p63, but small amounts of other poliovirus proteins were also present. When the replicase-template complex was treated with RNAase T1 before centrifugation, a single peak of activity was found that sedimented at 20S and contained only labeled p63. Thus, p63 was the only viral polypeptide in the replicase bound to its endogenous RNA template; p63 may be active as a poly(U) polymerase either as a monomer protein or a 7S complex.