Skip to main content
PMC home page
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1982 Aug;2(8):949–965. doi: 10.1128/mcb.2.8.949

Isolation and characterization of human DNA fragments with nucleotide sequence homologies with the simian virus 40 regulatory region.

S E Conrad, M R Botchan
PMCID: PMC369883  PMID: 6290874

Abstract

A recombinant library of human DNA sequences was screened with a segment of simian virus 40 (SV40) DNA that spans the viral origin of replication. One hundred and fifty phage were isolated that hybridized to this probe. Restriction enzyme and hybridization analyses indicated that these sequences were partially homologous to one another. Direct DNA sequencing of two such SV40-hybridizing segments indicated that this was not a highly conserved family of sequences, but rather a set of DNA fragments that contained repetitive regions of high guanine plus cytosine content. These sequences were not members of the previously described Alu family of repeats and hybridized to SV40 DNA more strongly than do Alu family members. Computer analyses showed that the human DNA segments contained multiple homologies with sequences throughout the SV40 origin region, although sequences on the late side of the viral origin contained the strongest cross-hybridizing sequences. Because of the number and complexity of the matches detected, we could not determine unambiguously which of the many possible heteroduplexes between these DNAs was thermodynamically most favored. No hybridization of these human DNA sequences to any other segment of the SV40 genome was detected. In contrast, the human DNA segments isolated cross-hybridized with many sequences within the human genome. We tested for the presence of several functional domains on two of these human DNA fragments. One SV40-hybridizing fragment, SVCR29, contained a sequence which enhanced the efficiency of thymidine kinase transformation in human cells by approximately 20-fold. This effect was seen in an orientation-independent manner when the sequence was present at the 3' end of the chicken thymidine kinase gene. We propose that this segment of DNA contains a sequence analogous to the 72-base-pair repeats of SV40. The existence of such an "activator" element in cellular DNA raises the possibility that families of these sequences may exist in the mammalian genome.

Full text

PDF
954

Images in this article

952Image
on p.952
953Image
on p.953
955Image
on p.955
956Image
on p.956
959Image
on p.959

Selected References

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

  1. Banerji J., Rusconi S., Schaffner W. Expression of a beta-globin gene is enhanced by remote SV40 DNA sequences. Cell. 1981 Dec;27(2 Pt 1):299–308. doi: 10.1016/0092-8674(81)90413-x. [DOI] [PubMed] [Google Scholar]
  2. Benoist C., Chambon P. In vivo sequence requirements of the SV40 early promotor region. Nature. 1981 Mar 26;290(5804):304–310. doi: 10.1038/290304a0. [DOI] [PubMed] [Google Scholar]
  3. Benton W. D., Davis R. W. Screening lambdagt recombinant clones by hybridization to single plaques in situ. Science. 1977 Apr 8;196(4286):180–182. doi: 10.1126/science.322279. [DOI] [PubMed] [Google Scholar]
  4. Capecchi M. R. High efficiency transformation by direct microinjection of DNA into cultured mammalian cells. Cell. 1980 Nov;22(2 Pt 2):479–488. doi: 10.1016/0092-8674(80)90358-x. [DOI] [PubMed] [Google Scholar]
  5. DiMaio D., Nathans D. Cold-sensitive regulatory mutants of simian virus 40. J Mol Biol. 1980 Jun 15;140(1):129–142. doi: 10.1016/0022-2836(80)90359-9. [DOI] [PubMed] [Google Scholar]
  6. Fiers W., Contreras R., Haegemann G., Rogiers R., Van de Voorde A., Van Heuverswyn H., Van Herreweghe J., Volckaert G., Ysebaert M. Complete nucleotide sequence of SV40 DNA. Nature. 1978 May 11;273(5658):113–120. doi: 10.1038/273113a0. [DOI] [PubMed] [Google Scholar]
  7. Fritsch E. F., Lawn R. M., Maniatis T. Molecular cloning and characterization of the human beta-like globin gene cluster. Cell. 1980 Apr;19(4):959–972. doi: 10.1016/0092-8674(80)90087-2. [DOI] [PubMed] [Google Scholar]
  8. Ghosh P. K., Lebowitz P. Simian virus 40 early mRNA's contain multiple 5' termini upstream and downstream from a Hogness-Goldberg sequence; a shift in 5' termini during the lytic cycle is mediated by large T antigen. J Virol. 1981 Oct;40(1):224–240. doi: 10.1128/jvi.40.1.224-240.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Grosschedl R., Birnstiel M. L. Delimitation of far upstream sequences required for maximal in vitro transcription of an H2A histone gene. Proc Natl Acad Sci U S A. 1982 Jan;79(2):297–301. doi: 10.1073/pnas.79.2.297. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Grosschedl R., Birnstiel M. L. Spacer DNA sequences upstream of the T-A-T-A-A-A-T-A sequence are essential for promotion of H2A histone gene transcription in vivo. Proc Natl Acad Sci U S A. 1980 Dec;77(12):7102–7106. doi: 10.1073/pnas.77.12.7102. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Gruss P., Dhar R., Khoury G. Simian virus 40 tandem repeated sequences as an element of the early promoter. Proc Natl Acad Sci U S A. 1981 Feb;78(2):943–947. doi: 10.1073/pnas.78.2.943. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Hansen U., Tenen D. G., Livingston D. M., Sharp P. A. T antigen repression of SV40 early transcription from two promoters. Cell. 1981 Dec;27(3 Pt 2):603–613. doi: 10.1016/0092-8674(81)90402-5. [DOI] [PubMed] [Google Scholar]
  13. Jelinek W. R., Toomey T. P., Leinwand L., Duncan C. H., Biro P. A., Choudary P. V., Weissman S. M., Rubin C. M., Houck C. M., Deininger P. L. Ubiquitous, interspersed repeated sequences in mammalian genomes. Proc Natl Acad Sci U S A. 1980 Mar;77(3):1398–1402. doi: 10.1073/pnas.77.3.1398. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Lebowitz P., Ghosh P. K. Initiation and regulation of simian virus 40 early transcription in vitro. J Virol. 1982 Feb;41(2):449–461. doi: 10.1128/jvi.41.2.449-461.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Lee F., Mulligan R., Berg P., Ringold G. Glucocorticoids regulate expression of dihydrofolate reductase cDNA in mouse mammary tumour virus chimaeric plasmids. Nature. 1981 Nov 19;294(5838):228–232. doi: 10.1038/294228a0. [DOI] [PubMed] [Google Scholar]
  16. Levinson B., Khoury G., Vande Woude G., Gruss P. Activation of SV40 genome by 72-base pair tandem repeats of Moloney sarcoma virus. Nature. 1982 Feb 18;295(5850):568–572. doi: 10.1038/295568a0. [DOI] [PubMed] [Google Scholar]
  17. Lusky M., Botchan M. Inhibition of SV40 replication in simian cells by specific pBR322 DNA sequences. Nature. 1981 Sep 3;293(5827):79–81. doi: 10.1038/293079a0. [DOI] [PubMed] [Google Scholar]
  18. Martin R. G. The transformation of cell growth and transmogrification of DNA synthesis by simian virus 40. Adv Cancer Res. 1981;34:1–68. doi: 10.1016/s0065-230x(08)60238-9. [DOI] [PubMed] [Google Scholar]
  19. Maxam A. M., Gilbert W. Sequencing end-labeled DNA with base-specific chemical cleavages. Methods Enzymol. 1980;65(1):499–560. doi: 10.1016/s0076-6879(80)65059-9. [DOI] [PubMed] [Google Scholar]
  20. McCutchan J. H., Pagano J. S. Enchancement of the infectivity of simian virus 40 deoxyribonucleic acid with diethylaminoethyl-dextran. J Natl Cancer Inst. 1968 Aug;41(2):351–357. [PubMed] [Google Scholar]
  21. McCutchan T. F., Singer M. F. DNA sequences similar to those around the simian virus 40 origin of replication are present in the monkey genome. Proc Natl Acad Sci U S A. 1981 Jan;78(1):95–99. doi: 10.1073/pnas.78.1.95. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. McKay R., DiMaio D. Binding of an SV40 T antigen-related protein to the DNA of SV40 regulatory mutants. Nature. 1981 Feb 26;289(5800):810–813. doi: 10.1038/289810a0. [DOI] [PubMed] [Google Scholar]
  23. Miesfeld R., Krystal M., Arnheim N. A member of a new repeated sequence family which is conserved throughout eucaryotic evolution is found between the human delta and beta globin genes. Nucleic Acids Res. 1981 Nov 25;9(22):5931–5947. doi: 10.1093/nar/9.22.5931. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Moreau P., Hen R., Wasylyk B., Everett R., Gaub M. P., Chambon P. The SV40 72 base repair repeat has a striking effect on gene expression both in SV40 and other chimeric recombinants. Nucleic Acids Res. 1981 Nov 25;9(22):6047–6068. doi: 10.1093/nar/9.22.6047. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Myers R. M., Rio D. C., Robbins A. K., Tjian R. SV40 gene expression is modulated by the cooperative binding of T antigen to DNA. Cell. 1981 Aug;25(2):373–384. doi: 10.1016/0092-8674(81)90056-8. [DOI] [PubMed] [Google Scholar]
  26. Myers R. M., Tjian R. Construction and analysis of simian virus 40 origins defective in tumor antigen binding and DNA replication. Proc Natl Acad Sci U S A. 1980 Nov;77(11):6491–6495. doi: 10.1073/pnas.77.11.6491. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Perucho M., Hanahan D., Lipsich L., Wigler M. Isolation of the chicken thymidine kinase gene by plasmid rescue. Nature. 1980 May 22;285(5762):207–210. doi: 10.1038/285207a0. [DOI] [PubMed] [Google Scholar]
  28. Queen C. L., Korn L. J. Computer analysis of nucleic acids and proteins. Methods Enzymol. 1980;65(1):595–609. doi: 10.1016/s0076-6879(80)65062-9. [DOI] [PubMed] [Google Scholar]
  29. Queen C., Lord S. T., McCutchan T. F., Singer M. F. Three segments from the monkey genome that hybridize to simian virus 40 have common structural elements. Mol Cell Biol. 1981 Dec;1(12):1061–1068. doi: 10.1128/mcb.1.12.1061. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Rio D., Robbins A., Myers R., Tjian R. Regulation of simian virus 40 early transcription in vitro by a purified tumor antigen. Proc Natl Acad Sci U S A. 1980 Oct;77(10):5706–5710. doi: 10.1073/pnas.77.10.5706. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Sakano H., Maki R., Kurosawa Y., Roeder W., Tonegawa S. Two types of somatic recombination are necessary for the generation of complete immunoglobulin heavy-chain genes. Nature. 1980 Aug 14;286(5774):676–683. doi: 10.1038/286676a0. [DOI] [PubMed] [Google Scholar]
  32. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  33. Stinchcomb D. T., Struhl K., Davis R. W. Isolation and characterisation of a yeast chromosomal replicator. Nature. 1979 Nov 1;282(5734):39–43. doi: 10.1038/282039a0. [DOI] [PubMed] [Google Scholar]
  34. Struhl K., Stinchcomb D. T., Scherer S., Davis R. W. High-frequency transformation of yeast: autonomous replication of hybrid DNA molecules. Proc Natl Acad Sci U S A. 1979 Mar;76(3):1035–1039. doi: 10.1073/pnas.76.3.1035. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Tyndall C., La Mantia G., Thacker C. M., Favaloro J., Kamen R. A region of the polyoma virus genome between the replication origin and late protein coding sequences is required in cis for both early gene expression and viral DNA replication. Nucleic Acids Res. 1981 Dec 11;9(23):6231–6250. doi: 10.1093/nar/9.23.6231. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Wigler M., Pellicer A., Silverstein S., Axel R. Biochemical transfer of single-copy eucaryotic genes using total cellular DNA as donor. Cell. 1978 Jul;14(3):725–731. doi: 10.1016/0092-8674(78)90254-4. [DOI] [PubMed] [Google Scholar]
  37. de Villiers J., Schaffner W. A small segment of polyoma virus DNA enhances the expression of a cloned beta-globin gene over a distance of 1400 base pairs. Nucleic Acids Res. 1981 Dec 11;9(23):6251–6264. doi: 10.1093/nar/9.23.6251. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Molecular and Cellular Biology are provided here courtesy of Taylor & Francis

RESOURCES