Skip to main content
Journal of Virology logoLink to Journal of Virology
. 1989 Jan;63(1):242–249. doi: 10.1128/jvi.63.1.242-249.1989

Asp-286----Asn-286 in polyomavirus large T antigen relaxes the specificity of binding to the polyomavirus origin.

W J Tang 1, W R Folk 1
PMCID: PMC247678  PMID: 2535730

Abstract

We isolated revertants of a polyomavirus whose origin of DNA replication contains a point mutation in the palindrome to which large T antigen binds. Four independent second-site revertants contain an Asp-286----Asn-286 substitution in large T antigen. This mutant large T antigen activates replication of DNAs containing the mutant polyomavirus origin as well as replication of DNAs containing the wild-type origin; however, replication of DNAs with enhancer mutations is not activated by this large T antigen. The Asn-286 mutation occurs in a positively charge region of large T antigen near the location of several mutations which inactivate DNA replication. We suggest that this region of large T antigen is responsible for recognition of specific DNA sequences at the origin and that ionic forces are important for this interaction.

Full text

PDF
244

Images in this article

Selected References

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

  1. Baker T. A., Sekimizu K., Funnell B. E., Kornberg A. Extensive unwinding of the plasmid template during staged enzymatic initiation of DNA replication from the origin of the Escherichia coli chromosome. Cell. 1986 Apr 11;45(1):53–64. doi: 10.1016/0092-8674(86)90537-4. [DOI] [PubMed] [Google Scholar]
  2. Bendig M. M., Thomas T., Folk W. R. Regulatory mutants of polyoma virus defective in DNA replication and the synthesis of early proteins. Cell. 1980 Jun;20(2):401–409. doi: 10.1016/0092-8674(80)90626-1. [DOI] [PubMed] [Google Scholar]
  3. Berg J. M. Potential metal-binding domains in nucleic acid binding proteins. Science. 1986 Apr 25;232(4749):485–487. doi: 10.1126/science.2421409. [DOI] [PubMed] [Google Scholar]
  4. Bockus B. J., Schaffhausen B. Localization of the phosphorylations of polyomavirus large T antigen. J Virol. 1987 Apr;61(4):1155–1163. doi: 10.1128/jvi.61.4.1155-1163.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Böhnlein E., Gruss P. Interaction of distinct nuclear proteins with sequences controlling the expression of polyomavirus early genes. Mol Cell Biol. 1986 May;6(5):1401–1411. doi: 10.1128/mcb.6.5.1401. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Cowie A., Kamen R. Guanine nucleotide contacts within viral DNA sequences bound by polyomavirus large T antigen. J Virol. 1986 Feb;57(2):505–514. doi: 10.1128/jvi.57.2.505-514.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Cowie A., Kamen R. Multiple binding sites for polyomavirus large T antigen within regulatory sequences of polyomavirus DNA. J Virol. 1984 Dec;52(3):750–760. doi: 10.1128/jvi.52.3.750-760.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Cowie A., de Villiers J., Kamen R. Immortalization of rat embryo fibroblasts by mutant polyomavirus large T antigens deficient in DNA binding. Mol Cell Biol. 1986 Dec;6(12):4344–4352. doi: 10.1128/mcb.6.12.4344. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Dailey L., Basilico C. Sequences in the polyomavirus DNA regulatory region involved in viral DNA replication and early gene expression. J Virol. 1985 Jun;54(3):739–749. doi: 10.1128/jvi.54.3.739-749.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. DePamphilis M. L. Transcriptional elements as components of eukaryotic origins of DNA replication. Cell. 1988 Mar 11;52(5):635–638. doi: 10.1016/0092-8674(88)90398-4. [DOI] [PubMed] [Google Scholar]
  11. Dean F. B., Dodson M., Echols H., Hurwitz J. ATP-dependent formation of a specialized nucleoprotein structure by simian virus 40 (SV40) large tumor antigen at the SV40 replication origin. Proc Natl Acad Sci U S A. 1987 Dec;84(24):8981–8985. doi: 10.1073/pnas.84.24.8981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Deb S., Tsui S., Koff A., DeLucia A. L., Parsons R., Tegtmeyer P. The T-antigen-binding domain of the simian virus 40 core origin of replication. J Virol. 1987 Jul;61(7):2143–2149. doi: 10.1128/jvi.61.7.2143-2149.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Deninger P. L., Esty A., LaPorte P., Hsu H., Friedmann T. The nucleotide sequence and restriction enzyme sites of the polyoma genome. Nucleic Acids Res. 1980 Feb 25;8(4):855–860. [PMC free article] [PubMed] [Google Scholar]
  14. Dodson M., Dean F. B., Bullock P., Echols H., Hurwitz J. Unwinding of duplex DNA from the SV40 origin of replication by T antigen. Science. 1987 Nov 13;238(4829):964–967. doi: 10.1126/science.2823389. [DOI] [PubMed] [Google Scholar]
  15. Dodson M., Echols H., Wickner S., Alfano C., Mensa-Wilmot K., Gomes B., LeBowitz J., Roberts J. D., McMacken R. Specialized nucleoprotein structures at the origin of replication of bacteriophage lambda: localized unwinding of duplex DNA by a six-protein reaction. Proc Natl Acad Sci U S A. 1986 Oct;83(20):7638–7642. doi: 10.1073/pnas.83.20.7638. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Ebright R. H. Evidence for a contact between glutamine-18 of lac repressor and base pair 7 of lac operator. Proc Natl Acad Sci U S A. 1986 Jan;83(2):303–307. doi: 10.1073/pnas.83.2.303. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Ebright R. H., Kolb A., Buc H., Kunkel T. A., Krakow J. S., Beckwith J. Role of glutamic acid-181 in DNA-sequence recognition by the catabolite gene activator protein (CAP) of Escherichia coli: altered DNA-sequence-recognition properties of [Val181]CAP and [Leu181]CAP. Proc Natl Acad Sci U S A. 1987 Sep;84(17):6083–6087. doi: 10.1073/pnas.84.17.6083. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Evans R. M., Hollenberg S. M. Zinc fingers: gilt by association. Cell. 1988 Jan 15;52(1):1–3. doi: 10.1016/0092-8674(88)90522-3. [DOI] [PubMed] [Google Scholar]
  19. Fowler R. G., Degnen G. E., Cox E. C. Mutational specificity of a conditional Escherichia coli mutator, mutD5. Mol Gen Genet. 1974;133(3):179–191. doi: 10.1007/BF00267667. [DOI] [PubMed] [Google Scholar]
  20. Fujimura F. K. Nuclear activity from F9 embryonal carcinoma cells binding specifically to the enhancers of wild-type polyoma virus and PyEC mutant DNAs. Nucleic Acids Res. 1986 Apr 11;14(7):2845–2861. doi: 10.1093/nar/14.7.2845. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Gaudray P., Clertant P., Cuzin F. ATP phosphohydrolase (ATPase) activity of a polyoma virus T antigen. Eur J Biochem. 1980 Aug;109(2):553–560. doi: 10.1111/j.1432-1033.1980.tb04827.x. [DOI] [PubMed] [Google Scholar]
  22. Hassauer M., Scheidtmann K. H., Walter G. Mapping of phosphorylation sites in polyomavirus large T antigen. J Virol. 1986 Jun;58(3):805–816. doi: 10.1128/jvi.58.3.805-816.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Hendrickson E. A., Fritze C. E., Folk W. R., DePamphilis M. L. The origin of bidirectional DNA replication in polyoma virus. EMBO J. 1987 Jul;6(7):2011–2018. doi: 10.1002/j.1460-2075.1987.tb02465.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. 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]
  25. Jones N. C., Rigby P. W., Ziff E. B. Trans-acting protein factors and the regulation of eukaryotic transcription: lessons from studies on DNA tumor viruses. Genes Dev. 1988 Mar;2(3):267–281. doi: 10.1101/gad.2.3.267. [DOI] [PubMed] [Google Scholar]
  26. Kalderon D., Richardson W. D., Markham A. F., Smith A. E. Sequence requirements for nuclear location of simian virus 40 large-T antigen. Nature. 1984 Sep 6;311(5981):33–38. doi: 10.1038/311033a0. [DOI] [PubMed] [Google Scholar]
  27. Kalderon D., Smith A. E. In vitro mutagenesis of a putative DNA binding domain of SV40 large-T. Virology. 1984 Nov;139(1):109–137. doi: 10.1016/0042-6822(84)90334-9. [DOI] [PubMed] [Google Scholar]
  28. Katinka M., Yaniv M. DNA replication origin of polyoma virus: early proximal boundary. J Virol. 1983 Jul;47(1):244–248. doi: 10.1128/jvi.47.1.244-248.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Kovesdi I., Satake M., Furukawa K., Reichel R., Ito Y., Nevins J. R. A factor discriminating between the wild-type and a mutant polyomavirus enhancer. Nature. 1987 Jul 2;328(6125):87–89. doi: 10.1038/328087a0. [DOI] [PubMed] [Google Scholar]
  30. Kunkel T. A., Roberts J. D., Zakour R. A. Rapid and efficient site-specific mutagenesis without phenotypic selection. Methods Enzymol. 1987;154:367–382. doi: 10.1016/0076-6879(87)54085-x. [DOI] [PubMed] [Google Scholar]
  31. Lehming N., Sartorius J., Niemöller M., Genenger G., v Wilcken-Bergmann B., Müller-Hill B. The interaction of the recognition helix of lac repressor with lac operator. EMBO J. 1987 Oct;6(10):3145–3153. doi: 10.1002/j.1460-2075.1987.tb02625.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Luthman H., Nilsson M. G., Magnusson G. Non-contiguous segments of the polyoma genome required in cis for DNA replication. J Mol Biol. 1982 Nov 15;161(4):533–550. doi: 10.1016/0022-2836(82)90406-5. [DOI] [PubMed] [Google Scholar]
  33. Luthman H., Osterlund M., Magnusson G. Inhibition of polyoma DNA synthesis by base pair substitutions at the replication origin. Nucleic Acids Res. 1984 Oct 11;12(19):7503–7515. doi: 10.1093/nar/12.19.7503. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Margolskee R. F., Nathans D. Simian virus 40 mutant T antigens with relaxed specificity for the nucleotide sequence at the viral DNA origin of replication. J Virol. 1984 Feb;49(2):386–393. doi: 10.1128/jvi.49.2.386-393.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Martin M. E., Piette J., Yaniv M., Tang W. J., Folk W. R. Activation of the polyomavirus enhancer by a murine activator protein 1 (AP1) homolog and two contiguous proteins. Proc Natl Acad Sci U S A. 1988 Aug;85(16):5839–5843. doi: 10.1073/pnas.85.16.5839. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. McClarin J. A., Frederick C. A., Wang B. C., Greene P., Boyer H. W., Grable J., Rosenberg J. M. Structure of the DNA-Eco RI endonuclease recognition complex at 3 A resolution. Science. 1986 Dec 19;234(4783):1526–1541. doi: 10.1126/science.3024321. [DOI] [PubMed] [Google Scholar]
  37. 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]
  38. Muller W. J., Mueller C. R., Mes A. M., Hassell J. A. Polyomavirus origin for DNA replication comprises multiple genetic elements. J Virol. 1983 Sep;47(3):586–599. doi: 10.1128/jvi.47.3.586-599.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Murakami Y., Eki T., Yamada M., Prives C., Hurwitz J. Species-specific in vitro synthesis of DNA containing the polyoma virus origin of replication. Proc Natl Acad Sci U S A. 1986 Sep;83(17):6347–6351. doi: 10.1073/pnas.83.17.6347. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Murakami Y., Wobbe C. R., Weissbach L., Dean F. B., Hurwitz J. Role of DNA polymerase alpha and DNA primase in simian virus 40 DNA replication in vitro. Proc Natl Acad Sci U S A. 1986 May;83(9):2869–2873. doi: 10.1073/pnas.83.9.2869. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Ohlendorf D. H., Matthews B. W. Structural studies of protein-nucleic acid interactions. Annu Rev Biophys Bioeng. 1983;12:259–284. doi: 10.1146/annurev.bb.12.060183.001355. [DOI] [PubMed] [Google Scholar]
  42. Ollis D. L., Brick P., Hamlin R., Xuong N. G., Steitz T. A. Structure of large fragment of Escherichia coli DNA polymerase I complexed with dTMP. 1985 Feb 28-Mar 6Nature. 313(6005):762–766. doi: 10.1038/313762a0. [DOI] [PubMed] [Google Scholar]
  43. Pabo C. O., Sauer R. T. Protein-DNA recognition. Annu Rev Biochem. 1984;53:293–321. doi: 10.1146/annurev.bi.53.070184.001453. [DOI] [PubMed] [Google Scholar]
  44. Pallas D. C., Schley C., Mahoney M., Harlow E., Schaffhausen B. S., Roberts T. M. Polyomavirus small t antigen: overproduction in bacteria, purification, and utilization for monoclonal and polyclonal antibody production. J Virol. 1986 Dec;60(3):1075–1084. doi: 10.1128/jvi.60.3.1075-1084.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Parker B. A., Stark G. R. Regulation of simian virus 40 transcription: sensitive analysis of the RNA species present early in infections by virus or viral DNA. J Virol. 1979 Aug;31(2):360–369. doi: 10.1128/jvi.31.2.360-369.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Paucha E., Kalderon D., Harvey R. W., Smith A. E. Simian virus 40 origin DNA-binding domain on large T antigen. J Virol. 1986 Jan;57(1):50–64. doi: 10.1128/jvi.57.1.50-64.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Piette J., Kryszke M. H., Yaniv M. Specific interaction of cellular factors with the B enhancer of polyoma virus. EMBO J. 1985 Oct;4(10):2675–2685. doi: 10.1002/j.1460-2075.1985.tb03987.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Piette J., Yaniv M. Two different factors bind to the alpha-domain of the polyoma virus enhancer, one of which also interacts with the SV40 and c-fos enhancers. EMBO J. 1987 May;6(5):1331–1337. doi: 10.1002/j.1460-2075.1987.tb02372.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Prives C., Covey L., Scheller A., Gluzman Y. DNA-binding properties of simian virus 40 T-antigen mutants defective in viral DNA replication. Mol Cell Biol. 1983 Nov;3(11):1958–1966. doi: 10.1128/mcb.3.11.1958. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Prives C., Murakami Y., Kern F. G., Folk W., Basilico C., Hurwitz J. DNA sequence requirements for replication of polyomavirus DNA in vivo and in vitro. Mol Cell Biol. 1987 Oct;7(10):3694–3704. doi: 10.1128/mcb.7.10.3694. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Richardson W. D., Roberts B. L., Smith A. E. Nuclear location signals in polyoma virus large-T. Cell. 1986 Jan 17;44(1):77–85. doi: 10.1016/0092-8674(86)90486-1. [DOI] [PubMed] [Google Scholar]
  52. Scheidtmann K. H., Echle B., Walter G. Simian virus 40 large T antigen is phosphorylated at multiple sites clustered in two separate regions. J Virol. 1982 Oct;44(1):116–133. doi: 10.1128/jvi.44.1.116-133.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Scheller A., Prives C. Simian virus 40 and polyomavirus large tumor antigens have different requirements for high-affinity sequence-specific DNA binding. J Virol. 1985 May;54(2):532–545. doi: 10.1128/jvi.54.2.532-545.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  54. Shortle D. R., Margolskee R. F., Nathans D. Mutational analysis of the simian virus 40 replicon: pseudorevertants of mutants with a defective replication origin. Proc Natl Acad Sci U S A. 1979 Dec;76(12):6128–6131. doi: 10.1073/pnas.76.12.6128. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Smale S. T., Tjian R. T-antigen-DNA polymerase alpha complex implicated in simian virus 40 DNA replication. Mol Cell Biol. 1986 Nov;6(11):4077–4087. doi: 10.1128/mcb.6.11.4077. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. Stahl H., Dröge P., Knippers R. DNA helicase activity of SV40 large tumor antigen. EMBO J. 1986 Aug;5(8):1939–1944. doi: 10.1002/j.1460-2075.1986.tb04447.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  57. Strauss M., Argani P., Mohr I. J., Gluzman Y. Studies on the origin-specific DNA-binding domain of simian virus 40 large T antigen. J Virol. 1987 Oct;61(10):3326–3330. doi: 10.1128/jvi.61.10.3326-3330.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  58. Tanaka I., Appelt K., Dijk J., White S. W., Wilson K. S. 3-A resolution structure of a protein with histone-like properties in prokaryotes. Nature. 1984 Aug 2;310(5976):376–381. doi: 10.1038/310376a0. [DOI] [PubMed] [Google Scholar]
  59. Tang W. J., Berger S. L., Triezenberg S. J., Folk W. R. Nucleotides in the polyomavirus enhancer that control viral transcription and DNA replication. Mol Cell Biol. 1987 May;7(5):1681–1690. doi: 10.1128/mcb.7.5.1681. [DOI] [PMC free article] [PubMed] [Google Scholar]
  60. Triezenberg S. J., Folk W. R. Essential nucleotides in the polyomavirus origin region. J Virol. 1984 Aug;51(2):437–444. doi: 10.1128/jvi.51.2.437-444.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  61. 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]
  62. Wallace R. B., Johnson M. J., Suggs S. V., Miyoshi K., Bhatt R., Itakura K. A set of synthetic oligodeoxyribonucleotide primers for DNA sequencing in the plasmid vector pBR322. Gene. 1981 Dec;16(1-3):21–26. doi: 10.1016/0378-1119(81)90057-3. [DOI] [PubMed] [Google Scholar]
  63. Wharton R. P., Ptashne M. A new-specificity mutant of 434 repressor that defines an amino acid-base pair contact. 1987 Apr 30-May 6Nature. 326(6116):888–891. doi: 10.1038/326888a0. [DOI] [PubMed] [Google Scholar]
  64. de Villiers J., Schaffner W., Tyndall C., Lupton S., Kamen R. Polyoma virus DNA replication requires an enhancer. Nature. 1984 Nov 15;312(5991):242–246. doi: 10.1038/312242a0. [DOI] [PubMed] [Google Scholar]
  65. van der Eb A. J., Graham F. L. Assay of transforming activity of tumor virus DNA. Methods Enzymol. 1980;65(1):826–839. doi: 10.1016/s0076-6879(80)65077-0. [DOI] [PubMed] [Google Scholar]
  66. von Hippel P. H., Berg O. G. On the specificity of DNA-protein interactions. Proc Natl Acad Sci U S A. 1986 Mar;83(6):1608–1612. doi: 10.1073/pnas.83.6.1608. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Virology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES