Recombinogenic properties of herpes simplex virus type 1 DNA sequences resident in simian virus 40 minichromosomes

Abstract

In a previous work, it was demonstrated that the bacterial transposon TnS is capable of undergoing sequence inversion via recombination between its duplicated IS50 elements when replicated by the herpes simplex virus type 1 (HSV-1) origin oris but not by the simian virus 40 (SV40) origin oris8. Further analysis of the latter phenomenon indicated that this lack of recombination was the result of topological constraints imposed by the SV40 minichromosome, such that recombination events could be readily detected in TnS derivatives in which the IS50 elements were arranged in a direct rather than inverted orientation. With this information, a second set of experiments were carried out to examine how the highly recombinogenic sequences which mediate the inversion of the long (L) and short (S) components of the HSV-1 genome behave in an SV40 minichromosome. Tandem copies of the L-S junction of the HSV-1 genome were observed to promote deletions in an SV40 shuttle plasmid at a frequency that was considerably greater than that of duplicated bacterial plasmid vector DNA. However, the presence of superinfecting HSV-1 did not enhance the frequency of these recombination events. These results support our previous findings that HSV-1 genome isomerization is mediated by a homologous recombination mechanism which is intimately associated with the act of viral DNA synthesis. Moreover, they demonstrate that the sequences which comprise the L-S junction appear to be inherently recombinogenic and, therefore, do not contain specific signals required for HSV-1 genome isomerization.

Replication of simian virus 40 origin-containing DNA during infection with a recombinant Autographa californica multiple nuclear polyhedrosis virus expressing large T antigen

“…4C, lane 6). The observation that an SV40 plasmid previously shown to be incapable of recombination during replication in mammalian cells (42) can now undergo levels of sequence inversion comparable to that in AcMNPV-replicated DNA (Fig. 4B and 4C) was further evi- Restriction enzyme sites for SalI (S) and XbaI (X) are included in the maps, as are the kanamycin phosphotransferase gene of Tn5 (Km r ) and the SV40 origin of DNA replication.…”

“…4). The ability of the inverted IS50 repeats of Tn5 to promote sequence inversion events as a result of DNA replication-mediated recombination has been shown to be associated with AcMNPV DNA replication (23) but not SV40 DNA replication occurring in mammalian cells (42).…”

“…A unique feature of AcMNPV-replicated DNA is its ability to undergo high-frequency homologous recombination between inverted repeat sequences (23). In contrast, such inversion events are prohibited in SV40 minichromosomes in mammalian cells, presumably because of topological constraints conferred by the supercoiled nature of their template DNA (42). Thus, if an AcMNPV-like mode of replication was indeed responsible for the observed amplification of plasmids carrying SV40 origins, these DNA molecules would now be expected to have the capacity to undergo high-frequency recombination.…”

“…An analogous construct, pTn5⌬1 SV , which was identical to pTn5⌬1 BV except that it contained an SV40 origin of replication, was used in this study. The Tn5 transposon of this construct was previously shown to be incapable of undergoing sequence inversion in mammalian cells (42). If these plasmids are digested with an enzyme(s) that cuts once outside of Tn5 and once asymmetrically within the unique sequence of Tn5, two fragments should be generated (Fig.…”

Herpes Simplex Virus Genome Isomerization: Origins of Adjacent Long Segments in Concatemeric Viral DNA

“…L and S segments freely invert relative to each other (4,11), generating four equimolar genomic isomers. Cleavage of concatemers at alternative L-S junctions can generate two isomers from a single genomic length replication template (15), but formation of the remaining isomers requires homologous recombination between sequences in the large repeated regions of DNA flanking L-S junctions (17,19,24,27,(29)(30)(31)(32). Inversion of adjacent L segments in HSV concatemers has been observed 3 to 4 h after infection, when replicating DNA is still devoid of free genomic termini (34).…”