Skip to main content
PMC home page
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1989 Dec;9(12):5412–5423. doi: 10.1128/mcb.9.12.5412

The adenovirus E1B 19-kilodalton protein stimulates gene expression by increasing DNA levels.

C H Herrmann 1, M B Mathews 1
PMCID: PMC363709  PMID: 2531284

Abstract

In transient expression assays, the adenovirus E1B 19-kilodalton (19K) tumor antigen increases expression from viral promoters and the promoter for the cellular 70-kilodalton heat shock protein (hsp70). To study the mechanism of this effect, we constructed HeLa cell lines that contain stably integrated copies of the 19K gene. Compared with a 19K- control cell line, 19K+ cells produced a significantly higher level of expression from every promoter introduced into the cells by transfection. The 19K protein also increased expression of an RNA polymerase III-transcribed gene but did not affect the level of expression of the endogenous hsp70 gene. The rate of transcription from transfected promoters, as measured by a nuclear run-on assay, was higher in the 19K+ cells than in the 19K- control cells. Furthermore, the level of plasmid DNA remained higher in the 19K+ cell line, suggesting that the 19K protein stabilizes transfected plasmid DNA. The elevated DNA levels seemed to account in full for the increased transcription. The role of the 19K protein in increasing gene expression during viral infection was found to be due to a replication-dependent increase in viral DNA levels. Thus, the 19K protein activates transcription indirectly by producing a higher level of viral or plasmid DNA. The DNA stabilization function of the 19K protein is probably related to the protective role of the 19K protein during viral infection and represents the first example of a viral oncogene product that modulates gene expression by regulating viral and plasmid DNA levels.

Full text

PDF
5412

Images in this article

5415Image
on p.5415
5415Image
on p.5415
5416Image
on p.5416
5417Image
on p.5417
5417Image
on p.5417
5418Image
on p.5418
5419Image
on p.5419
5419Image
on p.5419
5421Image
on p.5421
5421Image
on p.5421

Selected References

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

  1. Alwine J. C. Transient gene expression control: effects of transfected DNA stability and trans-activation by viral early proteins. Mol Cell Biol. 1985 May;5(5):1034–1042. doi: 10.1128/mcb.5.5.1034. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Anderson C. W., Schmitt R. C., Smart J. E., Lewis J. B. Early region 1B of adenovirus 2 encodes two coterminal proteins of 495 and 155 amino acid residues. J Virol. 1984 May;50(2):387–396. doi: 10.1128/jvi.50.2.387-396.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Babiss L. E., Fisher P. B., Ginsberg H. S. Effect on transformation of mutations in the early region 1b-encoded 21- and 55-kilodalton proteins of adenovirus 5. J Virol. 1984 Nov;52(2):389–395. doi: 10.1128/jvi.52.2.389-395.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Barker D. D., Berk A. J. Adenovirus proteins from both E1B reading frames are required for transformation of rodent cells by viral infection and DNA transfection. Virology. 1987 Jan;156(1):107–121. doi: 10.1016/0042-6822(87)90441-7. [DOI] [PubMed] [Google Scholar]
  5. Berk A. J. Functions of adenovirus E1A. Cancer Surv. 1986;5(2):367–387. [PubMed] [Google Scholar]
  6. Bernards R., de Leeuw M. G., Houweling A., van der Eb A. J. Role of the adenovirus early region 1B tumor antigens in transformation and lytic infection. Virology. 1986 Apr 15;150(1):126–139. doi: 10.1016/0042-6822(86)90272-2. [DOI] [PubMed] [Google Scholar]
  7. Bos J. L., Polder L. J., Bernards R., Schrier P. I., van den Elsen P. J., van der Eb A. J., van Ormondt H. The 2.2 kb E1b mRNA of human Ad12 and Ad5 codes for two tumor antigens starting at different AUG triplets. Cell. 1981 Nov;27(1 Pt 2):121–131. doi: 10.1016/0092-8674(81)90366-4. [DOI] [PubMed] [Google Scholar]
  8. Chinnadurai G. Adenovirus 2 Ip+ locus codes for a 19 kd tumor antigen that plays an essential role in cell transformation. Cell. 1983 Jul;33(3):759–766. doi: 10.1016/0092-8674(83)90018-1. [DOI] [PubMed] [Google Scholar]
  9. Chinnadurai G., Chinnadurai S., Brusca J. Physical mapping of a large-plaque mutation of adenovirus type 2. J Virol. 1979 Nov;32(2):623–628. doi: 10.1128/jvi.32.2.623-628.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. D'Halluin J. C., Allart C., Cousin C., Boulanger P. A., Martin G. R. Adenovirus early function required for protection of viral and cellular DNA. J Virol. 1979 Oct;32(1):61–71. doi: 10.1128/jvi.32.1.61-71.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Dery C. V., Herrmann C. H., Mathews M. B. Response of individual adenovirus promoters to the products of the E1A gene. Oncogene. 1987;2(1):15–23. [PubMed] [Google Scholar]
  12. Esche H., Mathews M. B., Lewis J. B. Proteins and messenger RNAs of the transforming region of wild-type and mutant adenoviruses. J Mol Biol. 1980 Sep 25;142(3):399–417. doi: 10.1016/0022-2836(80)90279-x. [DOI] [PubMed] [Google Scholar]
  13. Ezoe H., Fatt R. B., Mak S. Degradation of intracellular DNA in KB cells infected with cyt mutants of human adenovirus type 12. J Virol. 1981 Oct;40(1):20–27. doi: 10.1128/jvi.40.1.20-27.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Gaynor R. B., Feldman L. T., Berk A. J. Transcription of class III genes activated by viral immediate early proteins. Science. 1985 Oct 25;230(4724):447–450. doi: 10.1126/science.2996135. [DOI] [PubMed] [Google Scholar]
  15. Gorman C. M., Moffat L. F., Howard B. H. Recombinant genomes which express chloramphenicol acetyltransferase in mammalian cells. Mol Cell Biol. 1982 Sep;2(9):1044–1051. doi: 10.1128/mcb.2.9.1044. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Graham F. L., Smiley J., Russell W. C., Nairn R. Characteristics of a human cell line transformed by DNA from human adenovirus type 5. J Gen Virol. 1977 Jul;36(1):59–74. doi: 10.1099/0022-1317-36-1-59. [DOI] [PubMed] [Google Scholar]
  17. Greenberg M. E., Ziff E. B. Stimulation of 3T3 cells induces transcription of the c-fos proto-oncogene. Nature. 1984 Oct 4;311(5985):433–438. doi: 10.1038/311433a0. [DOI] [PubMed] [Google Scholar]
  18. Herr W., Clarke J. The SV40 enhancer is composed of multiple functional elements that can compensate for one another. Cell. 1986 May 9;45(3):461–470. doi: 10.1016/0092-8674(86)90332-6. [DOI] [PubMed] [Google Scholar]
  19. Herrmann C. H., Dery C. V., Mathews M. B. Transactivation of host and viral genes by the adenovirus E1B 19K tumor antigen. Oncogene. 1987;2(1):25–35. [PubMed] [Google Scholar]
  20. Hirt B. Selective extraction of polyoma DNA from infected mouse cell cultures. J Mol Biol. 1967 Jun 14;26(2):365–369. doi: 10.1016/0022-2836(67)90307-5. [DOI] [PubMed] [Google Scholar]
  21. Jones N., Shenk T. An adenovirus type 5 early gene function regulates expression of other early viral genes. Proc Natl Acad Sci U S A. 1979 Aug;76(8):3665–3669. doi: 10.1073/pnas.76.8.3665. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. 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]
  23. Lai Fatt R. B., Mak S. Mapping of an adenovirus function involved in the inhibition of DNA degradation. J Virol. 1982 Jun;42(3):969–977. doi: 10.1128/jvi.42.3.969-977.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Lewis J. B., Anderson C. W. Identification of adenovirus type 2 early region 1B proteins that share the same amino terminus as do the 495R and 155R proteins. J Virol. 1987 Dec;61(12):3879–3888. doi: 10.1128/jvi.61.12.3879-3888.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Lucher L. A., Brackmann K. H., Symington J. S., Green M. Antibody directed to a synthetic peptide encoding the NH2-terminal 16 amino acids of the adenovirus type 2 E1B-53K tumor antigen recognizes the E1B-20K tumor antigen. Virology. 1984 Jan 15;132(1):217–221. doi: 10.1016/0042-6822(84)90106-5. [DOI] [PubMed] [Google Scholar]
  26. Matsuo T., Wold W. S., Hashimoto S., Rankin A., Symington J., Green M. Polypeptides encoded by transforming region E 1b of human adenovirus 2: immunoprecipitation from transformed and infected cells and cell-free translation of E 1b-specific mRNA. Virology. 1982 Apr 30;118(2):456–465. doi: 10.1016/0042-6822(82)90366-x. [DOI] [PubMed] [Google Scholar]
  27. McGlade C. J., Tremblay M. L., Yee S. P., Ross R., Branton P. E. Acylation of the 176R (19-kilodalton) early region 1B protein of human adenovirus type 5. J Virol. 1987 Oct;61(10):3227–3234. doi: 10.1128/jvi.61.10.3227-3234.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Mellits K. H., Mathews M. B. Effects of mutations in stem and loop regions on the structure and function of adenovirus VA RNAI. EMBO J. 1988 Sep;7(9):2849–2859. doi: 10.1002/j.1460-2075.1988.tb03141.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Montell C., Fisher E. F., Caruthers M. H., Berk A. J. Control of adenovirus E1B mRNA synthesis by a shift in the activities of RNA splice sites. Mol Cell Biol. 1984 May;4(5):966–972. doi: 10.1128/mcb.4.5.966. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Moran E., Grodzicker T., Roberts R. J., Mathews M. B., Zerler B. Lytic and transforming functions of individual products of the adenovirus E1A gene. J Virol. 1986 Mar;57(3):765–775. doi: 10.1128/jvi.57.3.765-775.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Natarajan V. Adenovirus-2 E1a and E1b gene products regulate enhancer mediated transcription. Nucleic Acids Res. 1986 Dec 9;14(23):9445–9456. doi: 10.1093/nar/14.23.9445. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Nevins J. R. Induction of the synthesis of a 70,000 dalton mammalian heat shock protein by the adenovirus E1A gene product. Cell. 1982 Jul;29(3):913–919. doi: 10.1016/0092-8674(82)90453-6. [DOI] [PubMed] [Google Scholar]
  33. Pater A., Pater M. M. Expression of the control elements of BK and SV40 viruses in human cells exhibiting different transformed phenotypes. Virology. 1988 Apr;163(2):625–628. doi: 10.1016/0042-6822(88)90305-4. [DOI] [PubMed] [Google Scholar]
  34. Pilder S., Logan J., Shenk T. Deletion of the gene encoding the adenovirus 5 early region 1b 21,000-molecular-weight polypeptide leads to degradation of viral and host cell DNA. J Virol. 1984 Nov;52(2):664–671. doi: 10.1128/jvi.52.2.664-671.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Riabowol K. T., Mizzen L. A., Welch W. J. Heat shock is lethal to fibroblasts microinjected with antibodies against hsp70. Science. 1988 Oct 21;242(4877):433–436. doi: 10.1126/science.3175665. [DOI] [PubMed] [Google Scholar]
  36. Rosen C. A., Sodroski J. G., Haseltine W. A. The location of cis-acting regulatory sequences in the human T cell lymphotropic virus type III (HTLV-III/LAV) long terminal repeat. Cell. 1985 Jul;41(3):813–823. doi: 10.1016/s0092-8674(85)80062-3. [DOI] [PubMed] [Google Scholar]
  37. Ruley H. E. Adenovirus early region 1A enables viral and cellular transforming genes to transform primary cells in culture. Nature. 1983 Aug 18;304(5927):602–606. doi: 10.1038/304602a0. [DOI] [PubMed] [Google Scholar]
  38. Senear A. W., Lewis J. B. Morphological transformation of established rodent cell lines by high-level expression of the adenovirus type 2 E1a gene. Mol Cell Biol. 1986 Apr;6(4):1253–1260. doi: 10.1128/mcb.6.4.1253. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Spector D. J., McGrogan M., Raskas H. J. Regulation of the appearance of cytoplasmic RNAs from region 1 of the adenovirus 2 genome. J Mol Biol. 1978 Dec 15;126(3):395–414. doi: 10.1016/0022-2836(78)90048-7. [DOI] [PubMed] [Google Scholar]
  40. Stillman B. W. The replication of adenovirus DNA with purified proteins. Cell. 1983 Nov;35(1):7–9. doi: 10.1016/0092-8674(83)90201-5. [DOI] [PubMed] [Google Scholar]
  41. Stillman B. Functions of the adenovirus E1B tumour antigens. Cancer Surv. 1986;5(2):389–404. [PubMed] [Google Scholar]
  42. Subramanian T., Kuppuswamy M., Gysbers J., Mak S., Chinnadurai G. 19-kDa tumor antigen coded by early region E1b of adenovirus 2 is required for efficient synthesis and for protection of viral DNA. J Biol Chem. 1984 Oct 10;259(19):11777–11783. [PubMed] [Google Scholar]
  43. Subramanian T., Kuppuswamy M., Mak S., Chinnadurai G. Adenovirus cyt+ locus, which controls cell transformation and tumorigenicity, is an allele of lp+ locus, which codes for a 19-kilodalton tumor antigen. J Virol. 1984 Nov;52(2):336–343. doi: 10.1128/jvi.52.2.336-343.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Takemori N., Cladaras C., Bhat B., Conley A. J., Wold W. S. cyt gene of adenoviruses 2 and 5 is an oncogene for transforming function in early region E1B and encodes the E1B 19,000-molecular-weight polypeptide. J Virol. 1984 Dec;52(3):793–805. doi: 10.1128/jvi.52.3.793-805.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Takemori N., Riggs J. L., Aldrich C. D. Genetic studies with tumorigenic adenoviruses. II. Heterogeneity of cyt mutants of adenovirus type 12. Virology. 1969 May;38(1):8–15. doi: 10.1016/0042-6822(69)90122-6. [DOI] [PubMed] [Google Scholar]
  46. Takemori N., Riggs J. L., Aldrich C. Genetic studies with tumorigenic adenoviruses. I. Isolation of cytocidal (cyt) mutants of adenovirus type 12. Virology. 1968 Dec;36(4):575–586. doi: 10.1016/0042-6822(68)90189-x. [DOI] [PubMed] [Google Scholar]
  47. Vales L. D., Darnell J. E., Jr Promoter occlusion prevents transcription of adenovirus polypeptide IX mRNA until after DNA replication. Genes Dev. 1989 Jan;3(1):49–59. doi: 10.1101/gad.3.1.49. [DOI] [PubMed] [Google Scholar]
  48. Virtanen A., Pettersson U. Organization of early region 1B of human adenovirus type 2: identification of four differentially spliced mRNAs. J Virol. 1985 May;54(2):383–391. doi: 10.1128/jvi.54.2.383-391.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. White E., Blose S. H., Stillman B. W. Nuclear envelope localization of an adenovirus tumor antigen maintains the integrity of cellular DNA. Mol Cell Biol. 1984 Dec;4(12):2865–2875. doi: 10.1128/mcb.4.12.2865. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. White E., Denton A., Stillman B. Role of the adenovirus E1B 19,000-dalton tumor antigen in regulating early gene expression. J Virol. 1988 Sep;62(9):3445–3454. doi: 10.1128/jvi.62.9.3445-3454.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. White E., Faha B., Stillman B. Regulation of adenovirus gene expression in human WI38 cells by an E1B-encoded tumor antigen. Mol Cell Biol. 1986 Nov;6(11):3763–3773. doi: 10.1128/mcb.6.11.3763. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. White E., Grodzicker T., Stillman B. W. Mutations in the gene encoding the adenovirus early region 1B 19,000-molecular-weight tumor antigen cause the degradation of chromosomal DNA. J Virol. 1984 Nov;52(2):410–419. doi: 10.1128/jvi.52.2.410-419.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. White E., Stillman B. Expression of adenovirus E1B mutant phenotypes is dependent on the host cell and on synthesis of E1A proteins. J Virol. 1987 Feb;61(2):426–435. doi: 10.1128/jvi.61.2.426-435.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  54. Wilson M. C., Darnell J. E., Jr Control of messenger RNA concentration by differential cytoplasmic half-life. Adenovirus messenger RNAs from transcription units 1A and 1B. J Mol Biol. 1981 May 25;148(3):231–251. doi: 10.1016/0022-2836(81)90537-4. [DOI] [PubMed] [Google Scholar]
  55. Wu B. J., Kingston R. E., Morimoto R. I. Human HSP70 promoter contains at least two distinct regulatory domains. Proc Natl Acad Sci U S A. 1986 Feb;83(3):629–633. doi: 10.1073/pnas.83.3.629. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. Yoshida K., Venkatesh L., Kuppuswamy M., Chinnadurai G. Adenovirus transforming 19-kD T antigen has an enhancer-dependent trans-activation function and relieves enhancer repression mediated by viral and cellular genes. Genes Dev. 1987 Sep;1(7):645–658. doi: 10.1101/gad.1.7.645. [DOI] [PubMed] [Google Scholar]
  57. van den Elsen P. J., Houweling A., van der Eb A. J. Morphological transformation of human adenoviruses is determined to a large extent by gene products of region E1a. Virology. 1983 Nov;131(1):242–246. doi: 10.1016/0042-6822(83)90549-4. [DOI] [PubMed] [Google Scholar]

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

RESOURCES