Synthesis of full-length, virus genomic DNA by nuclei of vaccinia-infected HeLa cells.
It is well established that vaccinia virus infection induces the synthesis of virus-specific DNA in cytoplasmic 'factories', which are sites of virus-specific transcription. The present study demonstrates that vaccinia virus-specific DNA is synthesized also in the nuclei of infected cells with a similar time course. Direct observation and radiolabelling confirm the integrity of isolated nuclei. Reconstitution experiments and inhibitor studies demonstrate that virus-induced DNA is synthesized de novo within nuclei and does not result from cytoplasmic contamination. Cell-specific DNA synthesis is inhibited completely after infection and nuclei of infected cells then synthesize DNA which co-sediments with virus genomic DNA in denaturing gradients. Restriction endonuclease cleavage and hybridization with a virus-specific probe indicate that this is full-length, virus genomic DNA. The biological implications of this are discussed.
D002458:Cell Fractionation; D002467:Cell Nucleus / Q000378:metabolism*; D002499:Centrifugation, Density Gradient; D003561:Cytarabine / Q000494:pharmacology; D003593:Cytoplasm / Q000378:metabolism; D004279:DNA, Viral / Q000096:biosynthesis*; D005467:Floxuridine / Q000494:pharmacology; D005814:Genes, Viral; D006367:HeLa Cells; D006801:Humans; D009693:Nucleic Acid Hybridization; D013936:Thymidine / Q000378:metabolism; D014616:Vaccinia virus / Q000235:genetics / Q000378:metabolism*
D004279:DNA, Viral; D005467:Floxuridine; D003561:Cytarabine; D013936:Thymidine