Molecular cloning of DNA complementary to mRNA of the baculovirus Autographa californica nuclear polyhedrosis virus: location and gene products of RNA transcripts found late in infection

Abstract

DNAs complementary to late Autographa californica nuclear polyhedrosis virus (AcNPV) mRNA were synthesized by reverse transcription and cloned in Escherichia coli by using pBR322 as a vector. Eleven different cDNAs were distinguished in our screening of 45 AcNPV-homologous clones. Location of the regions of cDNA homology with respect to the AcNPV physical map showed that the 11 cDNAs were dispersed throughout the genome. The most abundant cDNA insertion, representing approximately one-third of the late viral mRNAs, was homologous to the AcNPV HindIII-P,Q and EcoRI-P fragments. The direction of transcription in this region was from left to right on a linearized AcNPV physical map. Hybridization selection followed by in vitro translation showed that this region encoded a 7,200-dalton (7.2K) protein which comigrated with a minor protein found in the extracellular nonoccluded form of the virus (NOV). Similarly, the gene for polyhedrin, the major structural protein of the occluded virus form, was located, at least in part, in the HindIII-V/EcoRI-I region of the AcNPV map. The polyhedrin transcript represented approximately one-quarter of the viral polyadenylic acid-containing RNAs at 27 h postinfection. Another relatively abundant cDNA was homologous to the HindIII-A/EcoRI-C/SstI-G region, and RNA selected by this cDNA directed the synthesis of two proteins (31K and 30K). The protein products of five other cDNA-selected RNAs were identified. The HindIII-D/EcoRI-O, HindIII-C/EcoRI-D, HindIII-B1/EcoRI-E, and HindIII-B2/EcoRI-H regions of the AcNPV L-1 genome were homologous to RNAs which directed the synthesis of a 57K protein, a 25K protein, a 61K protein, and a 37K protein (plus a minor 26K protein), respectively. Late mRNA selected by a cDNA homologous to the HindIII-P/EcoRI-B region of the AcNPV map directed the synthesis of 31K and 30K proteins which comigrated with the 31K and 30K proteins translated from RNA selected by the HindIII-A/EcoRI-C/SstI-G cDNA. Three other cDNAs have not been correlated yet with specific protein products.