Acquisition of Host Cell DNA Sequences by Baculoviruses: Relationship Between Host DNA Insertions and FP Mutants of Autographa californica and Galleria mellonella Nuclear Polyhedrosis Viruses

Abstract

Mutants of Autographa californica and Galleria mellonella nuclear polyhedrosis viruses, which produce an altered plaque phenotype as a result of reduced numbers of viral occlusions in infected cells, were isolated after passage in Trichoplusia ni (TN-368) cells. These mutants, termed FP (few-polyhedra) mutants, had acquired cell DNA sequences ranging from 0.8 to 2.8 kilobase pairs in size. The insertions of cell DNA occurred in a specific region between 35.0 and 37.7 map units of the A. californica viral genome. A cloned viral fragment containing one of the host DNA inserts was homologous to host DNA inserts in two other mutant viruses and to dispersed, repetitious sequences in T. ni cell DNA. Most of the homology between the cloned insert and cell DNA was contained within a 1,280-base-pair Alu I fragment. Marker rescue studies and analysis of infected-cell-specific proteins suggested that the insertion of cell DNA into the viral genomes resulted in the FP plaque phenotype, possibly through the inactivation of a 25,000-molecular-weight protein.

TTAA serves as the target site for TFP3 lepidopteran transposon insertions in both nuclear polyhedrosis virus and Trichoplusia ni genomes

“…This study and previous studies have established that TFP3 elements mobilize easily from the T. ni genome into the invading NPV genome (Fraser et a/., 1983(Fraser et a/., , 1985Kumar & Miller, 1987;Wang eta/., 1989). We have demonstrated for the first time that TFP3 transposons are capable of mobilizing within the uninfected T. ni genome in the absence of viral replication as the genomic TFP3/368.2 transposon interrupts a sequence in the TN-368 cellular genome that is contiguous within the larval genome (Fig.…”

“…We were interested in the structure and mobility of TFPB elements present in the TN-368 cellular and T. ni larval genomes (Fraser et a/., 1983;Wang et al, 1989). The primers p l OUT and p20UT (Experimental Procedures) were used for inverse PCR amplifications (see below) to prime synthesis outwardly from the TFP3-inverted repeats toward the flanking lepidopteran host sequences.…”

“…Hybridization analyses establish that the TFP3 elements are present as low repetitive copies in both the T. ni larvae and cellular genomes (Fraser eta 1983; Wang eta/., 1989). Two TFPB elements previously characterized (TFP3/1 and /2) insert within the 500 bp FP locus of the viral genome (AcMNPV map units 36-37), causing a distinct FP (few polyhedral occlusion bodies) plaque morphology phenotype and interrupting a gene encoding a 25 kDa viral-specific protein (Fraser et a/., 1983;Wang et a/., 1985;Beames & Summers, 1988.…”

“…Both elements insert at the same 'TTAA' tetranucleotide in the 25 K gene, and in the same relative orientation. Precise excision of the TFP3/1 element results in the restoration of the 25 K gene and reversion to the wild type plaque phenotype (Fraser et a/., 1983;Wang eta/., 1985).…”

The Baculoviruses Occlusion‐Derived Virus: Virion Structure and Function

“…FP25K studies began with a few polyhedra (FP) mutant baculovirus phenotypes that occurred during high titer, serial passage in cell culture (Fraser and Hink, 1982;Fraser et al, 1983;Stairs et al, 1981). Many FP mutant baculovirus phenotypes resulted from the disruption of the fp25k gene (Beames and Summers, 1989), thus the nomenclature "few polyhedra 25K" or FP25K.…”