A Virus-Encoded RNA Polymerase Purified from Baculovirus-Infected Cells

Abstract

A DNA-dependent RNA polymerase was purified to homogeneity, starting from insect cells infected with the baculovirus Autographa californica nuclear polyhedrosis virus (AcNPV). The purified polymerase supported accurate and specific transcription from late and very late promoters but was not active on viral early promoters. Thus, promoter recognition is an integral function of the purified enzyme. The purified RNA polymerase was composed of only four equimolar subunits, which makes it the simplest DNA-directed RNA polymerase from a eukaryotic source described so far. Amino-terminal protein sequencing, peptide fingerprinting, and immunochemical analyses were used to identify the four subunits, all of which are virus encoded. Overexpression of the four viral proteins (LEF-8, LEF-4, LEF-9, and p47) in baculovirus-infected cells resulted in a significant increase in the levels of RNA polymerase produced in the infected cells. Thus, the overexpression data are consistent with our identification of the RNA polymerase subunits.