Transformation of Human Embryonic Fibroblasts by BK Virus, BK Virus DNA and a Subgenomic BK Virus DNA Fragment Free

Abstract

Summary

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 I and RI. 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 I/RI 3.7 kb fragment showed two separate integrations of BKV sequences without free virus DNA.

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1982-12-01
2024-03-28
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References

  1. Aaronson S. A., Todaro G. J. 1970; Infectious SV40 and SV40 DNA: rapid fluorescent focus assay. Proceedings of the Society for Experimental Biology and Medicine 134:103–106
    [Google Scholar]
  2. Beard P., Morrow J. F., Berg P. 1973; Cleavage of circular, superhelical simian virus 40 DNA to a linear duplex by SI nuclease. Journal of Virology 12:1303–1313
    [Google Scholar]
  3. Carp R. I., Gilden R. 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
    [Google Scholar]
  4. Carroll D., Hansen J. L., Maryon E. B., O’Neill F. J. 1981; SV40 defectives selected during low multiplicity passage on A172 human glioblastoma cells. Virology 112:461–471
    [Google Scholar]
  5. Chenciner N., Grossi M. P., Meneguzzi G., Corallini A., Manservigi R., Barbanti-Brodano G., Milanesi G. 1980; State of viral DNA in BK virus-transformed rabbit cells. Virology 103:138–148
    [Google Scholar]
  6. Chia W., Rigby P. 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
    [Google Scholar]
  7. Chowdhury K., Gruss P., Waldeck W., Sauer G. 1975; Action of SI nuclease on nicked circular simian virus 40 DNA. Biochemical and Biophysical Research Communications 64:709–716
    [Google Scholar]
  8. Corallini A., Grossi M. P., Carra’ L., Caputo A., Balboni P. G., Negrini M., Milanesi G., Federspil G., Barbanti-Brodano G. 1983; Episomal DNA of a BK virus variant in a human insulinoma. Journal of Medical Virology (in press)
    [Google Scholar]
  9. Dulbecco R., Vogt M. 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
    [Google Scholar]
  10. Fernandes M., Moorhead P. 1965; Transformation of African green monkey kidney cultures infected with simian vacuolating virus 40 (SV40). Texas Reports on Biology and Medicine 23:242–258
    [Google Scholar]
  11. Gardner S. D., Field A. M., Coleman D. V., Hulme B. 1971; New human papovavirus (B.K.) isolated from urine after renal transplantation. Lancet 1:1253–1257
    [Google Scholar]
  12. Germond J. E., Vogt V. M., Hirt B. 1974; Characterization of the single-strand-specific nuclease SI activity on double-stranded supercoiled polyoma DNA. European Journal of Biochemistry 43:591–600
    [Google Scholar]
  13. Girardi A. J., Jensen F. C., Koprowski H. 1965; SV40-induced transformation of human diploid cells: crisis and recovery. Journal of Cellular and Comparative Physiology 65:69–84
    [Google Scholar]
  14. Graham F. L., Van Der Eb A. J. 1973; A new technique for the assay of infectivity of human adenovirus 5 DNA. Virology 52:456–467
    [Google Scholar]
  15. Hirt B. 1967; Selective extraction of polyoma DNA from infected mouse cell cultures. Journal of Molecular Biology 26:365–369
    [Google Scholar]
  16. Howley P. M. 1980; Molecular biology of SV40 and the human polyomaviruses BK and JC. In Viral Oncology pp. 489–550 Edited by Klein G. New York: Raven Press;
    [Google Scholar]
  17. Kelly T. J. Jr., Nathans D. 1977; The genome of simian virus 40. Advances in Virus Research 21:85–173
    [Google Scholar]
  18. Koprowski H., Ponten J. A., Jensen F., Ravdin R. G., Moorhead P., Saksela E. 1962; Transformation of cultures of human tissue infected with simian virus 40. Journal of Cellular and Comparative Physiology 59:281–292
    [Google Scholar]
  19. Kress M., May E., Cassingena R., May P. 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
    [Google Scholar]
  20. Krieg P., Amtmann E., Jonas D., Fischer H., Zang K., Sauer G. 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
    [Google Scholar]
  21. Laemmli U. K. 1970; Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature, London 227:680–685
    [Google Scholar]
  22. Lancaster W. D., Olson C. 1980; State of bovine papilloma virus DNA in connective-tissue tumors. In Viruses in Naturally Occurring Cancers pp. 223–232 Edited by Essex M., Todaro G., Zur Hausen H. New York: Cold Spring Harbor Laboratory;
    [Google Scholar]
  23. Lane D. P., Crawford L. V. 1979; T antigen is bound to a host protein in SV40-transformed cells. Nature, London 278:261–263
    [Google Scholar]
  24. Lavi S. 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
    [Google Scholar]
  25. Law M. F., Lowry D. R., Dvoretzky I., Howley P. 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
    [Google Scholar]
  26. McIntosh P. K., Dunker R., Mulder C., Brown N. C. 1978; DNA of Bacillus subtilis bacteriophage SPPI; physical mapping and localization of the origin of replication. Journal of Virology 28:865–876
    [Google Scholar]
  27. Maraldi N. M., Barbanti-Brodano G., Portolani M., La Placa M. 1975; Ultrastructural aspects of BK virus uptake and replication in human fibroblasts. Journal of General Virology 27:71–80
    [Google Scholar]
  28. Martin M. A., Howley P. M., Byrne J. C., Garon C. F. 1976; Characterization of supercoiled oligomeric SV40 DNA molecules in productively infected cells. Virology 71:28–40
    [Google Scholar]
  29. Moar M. H., Campo M. S., Laird H., Jarret W. 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
    [Google Scholar]
  30. O’Malley K. A., Davidson R. L. 1977; A new dimension in suspension fusion techniques with polyethylene glycol. Somatic Cell Genetics 3:441–448
    [Google Scholar]
  31. Padgett B. 1980; Human papovaviruses. In Molecular Biology of Tumor Viruses II. DNA Tumor Viruses pp. 339–370 Edited by Tooze J. New York: Cold Spring Harbor Laboratory;
    [Google Scholar]
  32. Pater M. M., Pater A., Fiori M., Slota J., Di Mayorca G. 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 Essex M., Todaro G., and zur Hansen H. New York: Cold Spring Harbor Laboratory;
    [Google Scholar]
  33. Pfister H., Fink B., Thomas C. 1981; Extrachromosomal bovine papillomavirus type 1 DNA in hamster fibromas and fibrosarcomas. Virology 115:414–418
    [Google Scholar]
  34. Portolani M., Barbanti-Brodano G., Laplaca M. 1975; Malignant transformation of hamster kidney cells by BK virus. Journal of Virology 15:420–422
    [Google Scholar]
  35. Portolani M., Borgatti M., Corallini A., Cassai E., Grossi M. P., Barbanti-Brodano G., Possati L. 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
    [Google Scholar]
  36. Purchio A. F., Fareed G. C. 1979; Transformation of human embryonic kidney cells by human papovavirus BK. Journal of Virology 29:763–769
    [Google Scholar]
  37. Rigby P. W. J., Berg P. 1978; Does simian virus 40 DNA integrate into cellular DNA during productive infection?. Journal of Virology 28:475–489
    [Google Scholar]
  38. Shah K. V., Hudson C., Valis J., Strandberg J. 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
    [Google Scholar]
  39. Shein H. M., Enders J. 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
    [Google Scholar]
  40. Takemoto K. K., Mullarkey M. F. 1973; Human papovavirus BK strain: biological studies including antigenic relationship to simian virus 40. Journal of Virology 12:625–631
    [Google Scholar]
  41. Takemoto K. K., Linke H., Miyamura T., Fareed G. C. 1979; Persistent BK papovavirus infection of transformed human fetal brain cells. Journal of Virology 29:1177–1185
    [Google Scholar]
  42. Wigler M., Pellicer A., Silverstein S., Axel R., Urlaub G., Chasin L. 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
    [Google Scholar]
  43. Winocour E., Keshet I., Nedjar G., Vogel T. 1980; Origins of SV40 genetic variation. Annals of the New York Academy of Sciences 354:43–52
    [Google Scholar]
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