Construction of a vaccinia virus deficient in the essential DNA repair enzyme uracil DNA glycosylase by a complementing cell line.
The vaccinia virus D4R open reading frame, encoding the essential DNA repair enzyme uracil DNA glycosylase, was expressed in two permanent cell lines, the rabbit kidney cell line RK13 and the human fibroblast cell line 293. The temperature-sensitive vaccinia virus mutant ts4149, which maps within D4R, was able to grow under restrictive conditions in both of these transformed cell lines. Cell clones complemented D4R function to various degrees, demonstrating complementation of an essential vaccinia virus gene by a cell line constitutively expressing the essential function. Thus, the complementing host cells allowed the rescue of a virus defective in the D4R gene, demonstrating that this system may be used for the propagation of defective cytoplasmic DNA viruses. The defective virus grew to high yields only in the engineered cell lines. The data support the hypothesis that early gene products, such as uracil DNA glycosylase, supplied in trans can fully complement essential viral functions.
Holzer
G W
GW
IMMUNO AG, Biomedical Research Center, Orth/Donau, Austria.
Falkner
F G
FG
D000818:Animals; D002460:Cell Line; D002461:Cell Line, Transformed; D002642:Chick Embryo; D003001:Cloning, Molecular; D045647:DNA Glycosylases; D004260:DNA Repair; D003673:Defective Viruses / Q000235:genetics / Q000254:growth & development* / Q000378:metabolism / Q000648:ultrastructure; D005816:Genetic Complementation Test; D006801:Humans; D009154:Mutation; D009699:N-Glycosyl Hydrolases / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D016366:Open Reading Frames; D011817:Rabbits; D014407:Tumor Cells, Cultured; D051981:Uracil-DNA Glycosidase; D014616:Vaccinia virus / Q000235:genetics / Q000254:growth & development* / Q000378:metabolism / Q000648:ultrastructure; D014764:Viral Proteins / Q000096:biosynthesis
D014764:Viral Proteins; D045647:DNA Glycosylases; D009699:N-Glycosyl Hydrolases; D051981:Uracil-DNA Glycosidase