The poliovirus replication complex: Site for synthesis of poliovirus RNA

Abstract

Pulse-labeled poliovirus RNA is found in the cytoplasm of HeLa cells in a structure which has been called the “poliovirus RNA replication complex”. The complex is bound to membranes but is released by treatment with deoxycholate. The replication complex sediments heterogeneously in a sucrose gradient with an average value of 250 s. It is unlikely that ribosomes are responsible for its fast rate of sedimentation because neither puromycin nor EDTA changes this rate. Furthermore, using cells in which the ribosomal RNA is highly radioactive, no association of ribosomes with the complex can be demonstrated. The viral RNA polymerase is found in association with the replication complex, and the in vitro synthesis of RNA occurs in the complex. Treatment of the complex with pronase or sodium dodecyl sulfate decreases the sedimentation rate of its RNA to 30 to 70 s. This suggests that protein is a major constituent of the complex; some or all of this protein must be the viral RNA polymerase. It also suggests that the sedimentation rate of the replicative intermediate may be faster than previously reported. The kinetics of incorporation of uridine into the complex and into finished chains of viral RNA are consistent with a precursor-product relation. These experiments are further evidence for the replicative intermediate as the site of viral RNA synthesis.