Late coding region sequences required for competition by SV40 defectives.
SV40 defectives containing the complete early coding region (E-SV40) or the complete late region (L-SV40) were separately transfected into green monkey cells. They were analyzed for their ability to compete with wtSV40 (introduced by infection) or to undergo replication in the presence of constitutively produced SV40 T-antigen. L-SV40 competed very strongly. It appeared rapidly in infected cells, overgrowing wt genomes by at least 10:1. In addition, it slowed the growth of wt virus and reduced its ability to kill cells. L-SV40 DNA, as expected, replicated continuously in Cosl cells. E-SV40 genomes were poor competitors. They appeared slowly and by themselves did not overgrow wtSV40. When transfected into Cosl cells, E-SV40 genomes replicated efficiently for the first few days and disappeared within a week. Deletion or insertion mutations were introduced into a molecular clone of L-SV40, within the Vp1 gene or the Vp2 gene. All mutants were unable to form infectious virus in two different assays. The mutants were then assayed for competition against wtSV40 and for replication in Cosl cells. The Vp1 mutants competed very poorly with wt genomes and were rapidly lost from coinfected cells. These mutants, like E-SV40, replicated for only a few days in Cosl cells. In contrast, the Vp2 mutant competed with wtSV40 nearly as well as L-SV40. It also replicated continuously rather than transiently in Cosl cells. Next, we determined whether L-SV40 could effectively compete with other evolved SV40 defectives, not containing the late region but containing up to nine SV40 origin regions. We have shown that within five serial passages, L-SV40 became the predominant viral DNA species and the other defectives were lost. Although the Vp1 mutants and E-SV40 were weak competitors, they were shown to recombine with wtSV40 genomes to generate new L-SV40 genomes which again became the predominant species of viral DNA. These results demonstrate that L-SV40 is a potent competitor and that the Vp1 gene or a part of Vp1 plays an important role in this extraordinary competition. We suggest that the Vp1 gene functions to allow L-SV40 genomes to persist rather than generating a product which directly interferes with wtSV40 replication.
O'Neill
F J
FJ
Research Service, V. A. Medical Center, Salt Lake City, Utah.
Miller
T H
TH
Stevens
R
R
D000818:Animals; D002460:Cell Line; D004252:DNA Mutational Analysis; D004261:DNA Replication; D003673:Defective Viruses / Q000235:genetics*; D005814:Genes, Viral; D005816:Genetic Complementation Test; D011995:Recombination, Genetic; D013539:Simian virus 40 / Q000235:genetics*; D014779:Virus Replication