The location, sequence, transcription, and regulation of a baculovirus DNA polymerase gene

Abstract

The Autographa californica nuclear polyhedrosis virus (AcMNPV) DNA polymerase gene was identified with the aid of an oligonucleotide probe corresponding to an amino acid sequence conserved among viral DNA polymerases of other virus families. A 3.6-kb pair region of the AcMNPV DNA, from 39.5 to 42.5 map units (m.u.), was sequenced and an open reading frame (ORF) of 2994 bp was observed. From the first ATG of this ORF, a translation product of 984 amino acids (Mr 114,310) was predicted. Amino acid sequence similarities to other viral DNA polymerases were found. Transcription was analyzed by Northern RNA blot analysis and nuclease protection studies of RNA:DNA hybrids. The ORF is transcribed in the counterclockwise direction as a 3-kb RNA. Transcripts appear to initiate at two differently regulated sites (ca. -120 and -212 bp) upstream of the initiating ATG (+1,+2,+3) and to be polyadenylated at a single site near a signal (A2UA3) which overlaps the translational termination signal (UAA) at +2952. Transcripts were observed only during a narrow window between 2 to 8 hr postinfection (p.i.) with maximum expression between 4 and 6 hr p.i. Polymerase gene transcripts were observed in the presence of the protein synthesis inhibitor cycloheximide which also blocked the shut-off of these early transcripts. Aphidicolin, an inhibitor of both viral and host DNA polymerases, inhibited polymerase gene transcription suggesting a unique regulation involving DNA replication.