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. 1979 Apr;30(1):21–31. doi: 10.1128/jvi.30.1.21-31.1979

In vitro translation of adenovirus type 12-specific mRNA isolated from infected and transformed cells.

H Esche, R Schilling, W Doerfler
PMCID: PMC353293  PMID: 480453

Abstract

The early and late gene products of human adenovirus type 12 (Ad12), as well as the viral proteins synthesized in an Ad12-transformed cell line, were identified by translation of viral mRNA in an in vitro protein-synthesizing system. Cytoplasmic RNA was isolated from permissive KB or nonpermissive BHK cells infected with Ad12 and from Ad12-transformed HA12/7 cells. Virus-specific RNA was selected by hybridization to Ad12 DNA covalently bound to cellulose. Viral RNA was then translated in a fractionated rabbit reticulocyte cell-free system or in wheat germ S-30 extracts. The proteins synthesized were characterized by immunoprecipitation and subsequent electrophoresis on sodium dodecyl sulfate-polyacrylamide gels. RNA prepared from KB cells late after infection with Ad12 elicited the synthesis of most of the structural polypeptides of the virion and at least two presumably nonstructural Ad12 proteins. When viral RNA isolated early after infection of KB cells with Ad12 was translated in vitro, 10 polypeptides were observed: E-68K, E-50K, E-42K, E-39K, E-34K, E-21K, E-19K, E-13K, E-12K, and E-10K. Ad12-specific RNA was also isolated from the Ad12-transformed hamster cell line HA12/7, which contains several copies of the Ad12 genome integrated in the host genome. The RNA codes for at least seven polypeptides with molecular weights very similar to those of the early viral proteins.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Anderson C. W., Baum P. R., Gesteland R. F. Processing of adenovirus 2-induced proteins. J Virol. 1973 Aug;12(2):241–252. doi: 10.1128/jvi.12.2.241-252.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Anderson C. W., Lewis J. B., Atkins J. F., Gesteland R. F. Cell-free synthesis of adenovirus 2 proteins programmed by fractionated messenger RNA: a comparison of polypeptide products and messenger RNA lengths. Proc Natl Acad Sci U S A. 1974 Jul;71(7):2756–2760. doi: 10.1073/pnas.71.7.2756. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. BABLANIAN R., EGGERS H. J., TAMM I. STUDIES ON THE MECHANISM OF POLIOVIRUS-INDUCED CELL DAMAGE. I. THE RELATION BETWEEN POLIOVIRUS,-INDUCED METABOLIC AND MORPHOLOGICAL ALTERATIONS IN CULTURED CELLS. Virology. 1965 May;26:100–113. doi: 10.1016/0042-6822(65)90030-9. [DOI] [PubMed] [Google Scholar]
  4. DULBECCO R., FREEMAN G. Plaque production by the polyoma virus. Virology. 1959 Jul;8(3):396–397. doi: 10.1016/0042-6822(59)90043-1. [DOI] [PubMed] [Google Scholar]
  5. Doerfler W. Integration of the deoxyribonucleic acid of adenovirus type 12 into the deoxyribonucleic acid of baby hamster kidney cells. J Virol. 1970 Nov;6(5):652–666. doi: 10.1128/jvi.6.5.652-666.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Doerfler W., Lundholm U. Absence of replication of the DNA of adenovirus type 12 in BHK21 cells. Virology. 1970 Mar;40(3):754–757. doi: 10.1016/0042-6822(70)90222-9. [DOI] [PubMed] [Google Scholar]
  7. Doerfler W., Lundholm U., Hirsch-Kauffmann M. Intracellular forms of adenovirus deoxyribonucleic acid. I. Evidence for a deoxyribonucleic acid-protein complex in baby hamster kidney cells infected with adenovirus type 12. J Virol. 1972 Feb;9(2):297–308. doi: 10.1128/jvi.9.2.297-308.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Doerfler W. Nonproductive infection of baby hamster kidney cells (BHK21) with adenovirus type 12. Virology. 1969 Aug;38(4):587–606. doi: 10.1016/0042-6822(69)90179-2. [DOI] [PubMed] [Google Scholar]
  9. Duesberg P. H., Vogt P. K. Gel electrophoresis of avian leukosis and sarcoma viral RNA in formamide: comparison with other viral and cellular RNA species. J Virol. 1973 Sep;12(3):594–599. doi: 10.1128/jvi.12.3.594-599.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. EAGLE H. Amino acid metabolism in mammalian cell cultures. Science. 1959 Aug 21;130(3373):432–437. doi: 10.1126/science.130.3373.432. [DOI] [PubMed] [Google Scholar]
  11. Eron L., Wesphal H., Callahan R. In vitro synthesis of adenovirus core proteins. J Virol. 1974 Aug;14(2):375–383. doi: 10.1128/jvi.14.2.375-383.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Falvey A. K., Staehelin T. Structure and function of mammalian ribosomes. I. Isolation and characterization of active liver ribosomal subunits. J Mol Biol. 1970 Oct 14;53(1):1–19. doi: 10.1016/0022-2836(70)90042-2. [DOI] [PubMed] [Google Scholar]
  13. Fanning E., Doerfler W. Intracellular forms of adenovirus DNA. V. Viral DNA sequences in hamster cells abortively infected and transformed with human adenovirus type 12. J Virol. 1976 Nov;20(2):373–383. doi: 10.1128/jvi.20.2.373-383.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Groneberg J., Chardonnet Y., Doerfler W. Integrated viral sequences in adenovirus type 12-transformed hamster cells. Cell. 1977 Jan;10(1):101–111. doi: 10.1016/0092-8674(77)90144-1. [DOI] [PubMed] [Google Scholar]
  15. HUEBNER R. J., ROWE W. P., TURNER H. C., LANE W. T. SPECIFIC ADENOVIRUS COMPLEMENT-FIXING ANTIGENS IN VIRUS-FREE HAMSTER AND RAT TUMORS. Proc Natl Acad Sci U S A. 1963 Aug;50:379–389. doi: 10.1073/pnas.50.2.379. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  17. Levinson A. D., Levine A. J. The group C adenovirus tumor antigens: identification in infected and transformed cells and a peptide map analysis. Cell. 1977 Aug;11(4):871–879. doi: 10.1016/0092-8674(77)90298-7. [DOI] [PubMed] [Google Scholar]
  18. Levinson A., Levine A. J. The isolation and identification of the adenovirus group C tumor antigens. Virology. 1977 Jan;76(1):1–11. doi: 10.1016/0042-6822(77)90275-6. [DOI] [PubMed] [Google Scholar]
  19. Lewis J. B., Anderson C. W., Atkins J. F., Gesteland R. F. The origin and destiny of adenovirus proteins. Cold Spring Harb Symp Quant Biol. 1975;39(Pt 1):581–590. doi: 10.1101/sqb.1974.039.01.072. [DOI] [PubMed] [Google Scholar]
  20. Lewis J. B., Atkins J. F., Baum P. R., Solem R., Gesteland R. F., Anderson C. W. Location and identification of the genes for adenovirus type 2 early polypeptides. Cell. 1976 Jan;7(1):141–151. doi: 10.1016/0092-8674(76)90264-6. [DOI] [PubMed] [Google Scholar]
  21. Miles L. E., Hales C. N. The preparation and properties of purified 125-I-labelled antibodies to insulin. Biochem J. 1968 Jul;108(4):611–618. doi: 10.1042/bj1080611. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Noyes B. E., Stark G. R. Nucleic acid hybridization using DNA covalently coupled to cellulose. Cell. 1975 Jul;5(3):301–310. doi: 10.1016/0092-8674(75)90105-1. [DOI] [PubMed] [Google Scholar]
  23. Ortin J., Doerfler W. Transcription of the genome of adenovirus type 12. I. Viral mRNA in abortively infected and transformed cells. J Virol. 1975 Jan;15(1):27–35. doi: 10.1128/jvi.15.1.27-35.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Ortin J., Scheidtmann K. H., Greenberg R., Westphal M., Doerfler W. Transcription of the genome of adenovirus type 12. III. Maps of stable RNA from productively infected human cells and abortively infected and transformed hamster cells. J Virol. 1976 Nov;20(2):355–372. doi: 10.1128/jvi.20.2.355-372.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Preston C. M. The cell-free synthesis of herpesvirus-induced polypeptides. Virology. 1977 May 1;78(1):349–353. doi: 10.1016/0042-6822(77)90109-x. [DOI] [PubMed] [Google Scholar]
  26. Raska K., Jr, Strohl W. A. The response of BHK21 cells to infection with type 12 adenovirus. VI. Synthesis of virus-specific RNA. Virology. 1972 Mar;47(3):734–742. doi: 10.1016/0042-6822(72)90563-6. [DOI] [PubMed] [Google Scholar]
  27. Roberts B. E., Paterson B. M. Efficient translation of tobacco mosaic virus RNA and rabbit globin 9S RNA in a cell-free system from commercial wheat germ. Proc Natl Acad Sci U S A. 1973 Aug;70(8):2330–2334. doi: 10.1073/pnas.70.8.2330. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Roberts B. E., Payne P. I., Osborne D. J. Protein synthesis and the viability of rye grains. Loss of activity of protein-synthesizing systems in vitro associated with a loss of viability. Biochem J. 1973 Feb;131(2):275–286. doi: 10.1042/bj1310275. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Rosenwirth B., Anderson C., Levine A. J. Tryptic fingerprint analysis of adenovirus types 2, 5 and 12 DNA-Binding proteins. Virology. 1976 Feb;69(2):617–625. doi: 10.1016/0042-6822(76)90490-6. [DOI] [PubMed] [Google Scholar]
  30. Rosenwirth B., Shiroki K., Levine A. J., Shimojo H. Isolation and characterization of adenovirus type 12 DNA binding proteins. Virology. 1975 Sep;67(1):14–23. doi: 10.1016/0042-6822(75)90399-2. [DOI] [PubMed] [Google Scholar]
  31. Scheidtmann K. H., Ortin J., Doerfler W. Transcription of the genome of adenovirus type 12. Viral mRNA in productively infected KB cells. Eur J Biochem. 1975 Oct 15;58(2):283–290. doi: 10.1111/j.1432-1033.1975.tb02374.x. [DOI] [PubMed] [Google Scholar]
  32. Schreier M. H., Staehelin T. Initiation of mammalian protein synthesis: the importance of ribosome and initiation factor quality for the efficiency of in vitro systems. J Mol Biol. 1973 Feb 19;73(3):329–349. doi: 10.1016/0022-2836(73)90346-x. [DOI] [PubMed] [Google Scholar]
  33. Strohl W. A. The response of BHK21 cells to infection with type 12 adenovirus. 1. Cell killing and T antigen synthesis as correlated viral genome functions. Virology. 1969 Dec;39(4):642–652. doi: 10.1016/0042-6822(69)90003-8. [DOI] [PubMed] [Google Scholar]
  34. Studier F. W. Analysis of bacteriophage T7 early RNAs and proteins on slab gels. J Mol Biol. 1973 Sep 15;79(2):237–248. doi: 10.1016/0022-2836(73)90003-x. [DOI] [PubMed] [Google Scholar]
  35. Sutter D., Westphal M., Doerfler W. Patterns of integration of viral DNA sequences in the genomes of adenovirus type 12-transformed hamster cells. Cell. 1978 Jul;14(3):569–585. doi: 10.1016/0092-8674(78)90243-x. [DOI] [PubMed] [Google Scholar]
  36. zur Hausen H. Interactions of adenovirus type 12 with host cell chromosomes. Prog Exp Tumor Res. 1973;18:240–259. doi: 10.1159/000393169. [DOI] [PubMed] [Google Scholar]

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