Location of Homologous DNA Sequences Interspersed at Five Regions in the Baculovirus AcMNPV Genome

Abstract

An examination of Autographa californica nuclear polyhedrosis virus DNA revealed the presence of five interspersed regions, rich in EcoRI restriction sites, which shared homologous sequences. These homologous regions (hr), designated hr1 to hr5, occur at or near the following EcoRI firagment junctions: hr1 EcoRI-B-EcoRI-I (0.0 map units); hr2, EcoRI-A-EcoRI-J (19.8 map units); hr3, EcoRI-C-EcoRI-G (52.9 map units); hr4, EcoRI-Q-EcoRI-L (69.8 map units); and hr5, EcoRI-S-EcoRI-X (88.0 map units). Four of these regions were identified, by cross-blot hybridization of HindIII-restricted A. californica nuclear polyhedrosis virus DNA, to be within the HindIII-AIB,-F,-L, and-Q fragments. The location of these regions and the identification of a fifth homologous region were confirmed, and their characterization was facilitated, by using two plasmids with HindIII-L or-Q fragment insertions, which contained the homologous regions hr2 and hr5, respectively. The sizes of the homologous regions were about 800 base pairs for hr2, 500 base pairs for hr5, and less than 500 base pairs for hr1, hr3, and hr4. A set of small EcoRI fragments (EcoRI minifragments) which ranged in size from 225 to 73 base pairs were detected in A. californica nuclear polyhedrosis virus DNA and HindIII-L and-Q fragments by polyacrylamide gel analysis. Some of the minifragments in viral DNA were present in extramolar amounts and corresponded in size to some of the minifragments present in HindIII-L and-Q. Clones of some of the EcoRI minifragments were used as probes in hybridizations to digests of viral DNA and of HindIII-L and-Q. The hybridization data, obtained under various levels of stringency, suggested that there was a degree of mismatching between the sequences which were responsible for the homology.

The hr5 transcriptional enhancer stimulates early expression from the Autographa californica nuclear polyhedrosis virus genome but is not required for virus replication

“…The hrs contain from two to eight imperfect palindromic repeats bisected by an EcoRI site (13,16,27). Because of their genomic distribution and repetitive nature, the hrs have been postulated to serve as origins of viral DNA replication (6). Consistent with this role is the observation that a single hr confers AcMNPV-dependent replication to plasmids in transfected cells (23,32).…”

“…Early and late viral genes are dispersed throughout the genome, and their proper expression results in the production of two morphologically and temporally distinct forms of AcMNPV progeny, budded virus (BV) and occluded virus (for recent reviews, see references 1 and 31). Five homologous regions (designated hrl through hr5), varying in size from 400 to 800 bp, are also interspersed throughout the genome (6). The hrs contain from two to eight imperfect palindromic repeats bisected by an EcoRI site (13,16,27).…”

The lef-3 gene of Autographa californica nuclear polyhedrosis virus encodes a single-stranded DNA-binding protein

“…The genome of Autographa californica nuclear polyhedrosis virus (AcNPV) is a double-stranded, supercoiled DNA molecule of 134 kb and potentially encodes 150 proteins (1). The viral genome consists of unique sequences, with the exception of five regions of homologous sequences (hrs 1 to 5) that are interspersed along the length of the genome (5). Each hr contains from two to eight 24-bp imperfect palindromes with naturally occurring EcoRI sites at their core.…”

Identification and localization of reiterated sequences in the Choristoneura fumiferana MNPV genome.

“…Kelly (1977) has shown by measurements of reassociation kinetics that 2-307o of the baculovirus genome are rapidly reannealing fractions, indicative of repeated DNA sequences. More recently, the genome of Autographa californica nuclear polyhedrosis virus (AcMNPV) has been shown to contain homologous sequences interdispersed at five locations (Cochran and Faulkner, 1983). Moreover, serial passage of AcMNPV in an established cell line derived from Trichoplusia ni, results in the amplification of certain parts of the genome (Burand and Summers, 1982).…”