Human embryonic fibroblasts (HEF) have been transformed by BK virus (BKV) DNA and by u.v.-inactivated or live BKV alone or in association with methyl-cholanthrene (MTC). The transformed cells produced BKV large T and small t antigens as well as the cellular 53 kdal protein, detected by immunofluorescence and immunoprecipitation. After an initial phase of lysis and virus shedding, virus or its coat protein antigen could not be detected in transformed cells. All human transformed cell lines could be superinfected by BKV or BKV DNA, but their susceptibility to superinfection was 20- to 500-fold lower than normal HEF. BKV could be rescued by fusion of transformed cells with normal HEF or Vero cells and by transfection of normal HEF with total DNA and DNA extracted from the Hirt supernatant of transformed cells. Blot hybridization analysis of DNA from transformed cells showed a considerable amount of free BKV DNA in monomeric and polymeric forms. Integrated BKV DNA was absent in most cell lines but present in only small amounts in BKV-transformed cells treated with MTC. Analysis of free BKV DNA with various restriction endonucleases and by blot hybridization showed that monomeric forms were complete BKV genomes, whereas polymers contained both complete and defective or rearranged BKV DNA. Transformation of HEF was also obtained with a 3.7 kilobase (kb) fragment of the BKV genome, produced by sequential digestion of BKV with the restriction endonucleases HhaI and EcoRI. This fragment extends clockwise on the virus genome from 0 to 72.2 map units and contains the entire early region. Blot hybridization analysis of cells transformed by the HhaI/EcoRI 3.7 kb fragment showed two separate integrations of BKV sequences without free virus DNA.
AaronsonS. A.,
TodaroG. J.1970; Infectious SV40 and SV40 DNA: rapid fluorescent focus assay. Proceedings of the Society for Experimental Biology and Medicine 134:103–106
BeardP.,
MorrowJ. F.,
BergP.1973; Cleavage of circular, superhelical simian virus 40 DNA to a linear duplex by SI nuclease. Journal of Virology 12:1303–1313
CarpR. I.,
GildenR. V.1966; A comparison of the replication cycles of simian virus 40 in human diploid and African green monkey kidney cells. Virology 28:150–162
ChencinerN.,
GrossiM. P.,
MeneguzziG.,
CoralliniA.,
ManservigiR.,
Barbanti-BrodanoG.,
MilanesiG.1980; State of viral DNA in BK virus-transformed rabbit cells. Virology 103:138–148
ChiaW.,
RigbyP. W. J.1981; Fate of viral DNA in nonpermissive cells infected with simian virus 40. Proceedings of the National Academy of Sciences of the United States of America 78:6638–6642
ChowdhuryK.,
GrussP.,
WaldeckW.,
SauerG.1975; Action of SI nuclease on nicked circular simian virus 40 DNA. Biochemical and Biophysical Research Communications 64:709–716
CoralliniA.,
GrossiM. P.,
Carra’L.,
CaputoA.,
BalboniP. G.,
NegriniM.,
MilanesiG.,
FederspilG.,
Barbanti-BrodanoG.1983; Episomal DNA of a BK virus variant in a human insulinoma. Journal of Medical Virology (in press)
DulbeccoR.,
VogtM.1960; Significance of continued virus production in tissue culture rendered neoplastic by polyoma virus. Proceedings of the National Academy of Sciences of the United States of America 46:1617–1632
FernandesM.,
MoorheadP.1965; Transformation of African green monkey kidney cultures infected with simian vacuolating virus 40 (SV40). Texas Reports on Biology and Medicine 23:242–258
GermondJ. E.,
VogtV. M.,
HirtB.1974; Characterization of the single-strand-specific nuclease SI activity on double-stranded supercoiled polyoma DNA. European Journal of Biochemistry 43:591–600
GirardiA. J.,
JensenF. C.,
KoprowskiH.1965; SV40-induced transformation of human diploid cells: crisis and recovery. Journal of Cellular and Comparative Physiology 65:69–84
HowleyP. M.1980; Molecular biology of SV40 and the human polyomaviruses BK and JC. In Viral Oncology pp. 489–550 Edited by
KleinG.
New York: Raven Press;
KoprowskiH.,
PontenJ. A.,
JensenF.,
RavdinR. G.,
MoorheadP.,
SakselaE.1962; Transformation of cultures of human tissue infected with simian virus 40. Journal of Cellular and Comparative Physiology 59:281–292
KressM.,
MayE.,
CassingenaR.,
MayP.1979; Simian virus 40-transformed cells express new species of proteins precipitable by anti-simian virus 40 tumor serum. Journal of Virology 31:472–483
KriegP.,
AmtmannE.,
JonasD.,
FischerH.,
ZangK.,
SauerG.1981; Episomal simian virus 40 genomes in human brain tumors. Proceedings of the National Academy of Sciences of the United States of America 78:6446–6450
LancasterW. D.,
OlsonC.1980; State of bovine papilloma virus DNA in connective-tissue tumors. In Viruses in Naturally Occurring Cancers pp. 223–232 Edited by
EssexM.,
TodaroG.,
Zur HausenH.
New York: Cold Spring Harbor Laboratory;
LaviS.1981; Carcinogen-mediated amplification of viral DNA sequences in simian virus 40-transformed Chinese hamster embryo cells. Proceedings of the National Academy of Sciences of the United States of America 78:6144–6148
LawM. F.,
LowryD. R.,
DvoretzkyI.,
HowleyP.1981; Mouse cells transformed by bovine papillomavirus contain only extrachromosomal viral DNA sequences. Proceedings of the National Academy of Sciences of the United States of America 78:2727–2731
McIntoshP. K.,
DunkerR.,
MulderC.,
BrownN. C.1978; DNA of Bacillus subtilis bacteriophage SPPI; physical mapping and localization of the origin of replication. Journal of Virology 28:865–876
MaraldiN. M.,
Barbanti-BrodanoG.,
PortolaniM.,
La PlacaM.1975; Ultrastructural aspects of BK virus uptake and replication in human fibroblasts. Journal of General Virology 27:71–80
MartinM. A.,
HowleyP. M.,
ByrneJ. C.,
GaronC. F.1976; Characterization of supercoiled oligomeric SV40 DNA molecules in productively infected cells. Virology 71:28–40
MoarM. H.,
CampoM. S.,
LairdH.,
JarretW. F. H.1981; Persistence of non-integrated viral DNA in bovine cells transformed in vitro by bovine papillomavirus type 2. Nature, London 293:749–751
PadgettB.1980; Human papovaviruses. In Molecular Biology of Tumor Viruses II. DNA Tumor Viruses pp. 339–370 Edited by
ToozeJ.
New York: Cold Spring Harbor Laboratory;
PaterM. M.,
PaterA.,
FioriM.,
SlotaJ.,
Di MayorcaG.1980; BK virus DNA sequences in human tumors and normal tissues and cell lines. In Viruses in Naturally Occurring Cancers pp. 329–341 Edited by
EssexM.,
TodaroG., and zur HansenH.
New York: Cold Spring Harbor Laboratory;
PortolaniM.,
BorgattiM.,
CoralliniA.,
CassaiE.,
GrossiM. P.,
Barbanti-BrodanoG.,
PossatiL.1978; Stable transformation of mouse, rabbit and monkey cells and abortive transformation of human cells by BK virus, a human papovavirus. Journal of General Virology 38:369–374
ShahK. V.,
HudsonC.,
ValisJ.,
StrandbergJ. D.1976; Experimental infection of human foreskin cultures with BK virus, a human papovavirus. Proceedings of the Society for Experimental Biology and Medicine 153:180–186
SheinH. M.,
EndersJ. F.1962; Multiplication and cytopathogenicity of simian vacuolating virus 40 in cultures of human tissues. Proceedings of the Society for Experimental Biology and Medicine 109:495–500
TakemotoK. K.,
MullarkeyM. F.1973; Human papovavirus BK strain: biological studies including antigenic relationship to simian virus 40. Journal of Virology 12:625–631
TakemotoK. K.,
LinkeH.,
MiyamuraT.,
FareedG. C.1979; Persistent BK papovavirus infection of transformed human fetal brain cells. Journal of Virology 29:1177–1185
WiglerM.,
PellicerA.,
SilversteinS.,
AxelR.,
UrlaubG.,
ChasinL.1979; DNA-mediated transfer of the adenine phosphoribosyltransferase locus into mammalian cells. Proceedings of the National Academy of Sciences of the United States of America 76:1373–1376