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. 1981 Apr;38(1):91–103. doi: 10.1128/jvi.38.1.91-103.1981

Two deletions within genes for simian virus 40 structural proteins VP2 and VP3 lead to formation of abnormal transcriptional complexes.

R Llopis, G R Stark
PMCID: PMC171129  PMID: 6264117

Abstract

The procedure developed by R. M. Fernandez-Muñoz et al. (J. Virol. 29:612-623, 1979) for isolating simian virus 40 (SV 40) chromatin free of disrupted previrions was optimized for preparing late transcriptional complexes, and these complexes were partially characterized. Transcriptional complexes derived from wild-type virus and from several deletion and temperature-sensitive mutants could be activated more than five-fold either by the anionic detergent Sarkosyl or by 300 mM ammonium sulfate, in agreement with the properties of SV40 transcriptional complexes prepared by other procedures. In contrast, complexes from cells infected with deletion mutants dl1261 or dl1262 were not activated at all by a high salt concentration, even though the extent of their activation by Sarkosyl was normal. Mutants dl1261 and dl1262 carry deletions of 54 and 36 base pairs, respectively, at an approximate map position of 0.91, which is within the overlapping genes for the virion proteins VP2 and VP3. The effects of these deletions on transcription in vitro indicate that VP2 or VP3 or both are bound to late transcriptional complexes in a way that affects the progress of initiated RNA polymerase. The properties of late transcriptional complexes derived from wild-type SV40 can be explained by the presence of the following two different kinds of complexes: (i) a minority class (about 20%), which is free of VP2 or VP3, active at low concentrations of ammonium sulfate in vitro, and responsible for late transcription in vivo, and (ii) a majority class (about 80%) with VP2 or VP3 bound, which is inactive at low salt concentrations both in vitro and in vivo but capable of being activated by high salt concentrations or by Sarkosyl. We propose that mutant VP2 and VP3 proteins from dl1261 and dl1262 bind to the majority class of late transcriptional complexes in a way that can be reversed by Sarkosyl but not by a high salt concentration.

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Selected References

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  1. Alwine J. C., Reed S. I., Stark G. R. Characterization of the autoregulation of simian virus 40 gene A. J Virol. 1977 Oct;24(1):22–27. doi: 10.1128/jvi.24.1.22-27.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Avila J., Saral R., Martin R. G., Khoury G. The temperature-sensitive defect in SV40 group D mutants. Virology. 1976 Aug;73(1):89–95. doi: 10.1016/0042-6822(76)90063-5. [DOI] [PubMed] [Google Scholar]
  3. Birkenmeier E. H., Chiu N., Radonovich M. F., May E., Salzman N. P. Regulation of simian virus 40 early and late gene transcription without viral DNA replication. J Virol. 1979 Mar;29(3):983–989. doi: 10.1128/jvi.29.3.983-989.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Birkenmeier E. H., May E., Salzman N. P. Characterization of simian virus 40 tsA58 transcriptional intermediates at restrictive temperatures: relationship between DNA replication and transcription. J Virol. 1977 Jun;22(3):702–710. doi: 10.1128/jvi.22.3.702-710.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Birkenmeier E. H., Radonovich M. F., Shani M., Salzman N. P. The SV40 DNA template for transcription of late mRNA in viral nucleoprotein complexes. Cell. 1977 Jul;11(3):495–504. doi: 10.1016/0092-8674(77)90067-8. [DOI] [PubMed] [Google Scholar]
  6. Brooks T. L., Green M. H. The sv40 transcription complex. I. Effect of viral chromatin proteins on endogenous RNA polymerase activity. Nucleic Acids Res. 1977 Dec;4(12):4261–4277. doi: 10.1093/nar/4.12.4261. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Chirgwin J. M., Przybyla A. E., MacDonald R. J., Rutter W. J. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry. 1979 Nov 27;18(24):5294–5299. doi: 10.1021/bi00591a005. [DOI] [PubMed] [Google Scholar]
  8. Chou J. Y., Avila J., Martin R. G. Viral DNA synthesis in cells infected by temperature-sensitive mutants of simian virus 40. J Virol. 1974 Jul;14(1):116–124. doi: 10.1128/jvi.14.1.116-124.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Chou J. Y., Martin R. G. DNA infectivity and the induction of host DNA synthesis with temperature-sensitive mutants of simian virus 40. J Virol. 1975 Jan;15(1):145–150. doi: 10.1128/jvi.15.1.145-150.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Cole C. N., Landers T., Goff S. P., Manteuil-Brutlag S., Berg P. Physical and genetic characterization of deletion mutants of simian virus 40 constructed in vitro. J Virol. 1977 Oct;24(1):277–294. doi: 10.1128/jvi.24.1.277-294.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Contreras R., Cole C., Berg P., Fiers W. Nucleotide sequence analysis of two simian virus 40 mutants with deletions in the late region of the genome. J Virol. 1979 Feb;29(2):789–793. doi: 10.1128/jvi.29.2.789-793.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Danna K. J., Nathans D. Bidirectional replication of Simian Virus 40 DNA. Proc Natl Acad Sci U S A. 1972 Nov;69(11):3097–3100. doi: 10.1073/pnas.69.11.3097. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. 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]
  14. Ferdinand F. J., Brown M., Khoury G. Characterization of early simian virus 40 transcriptional complexes: late transcription in the absence of detectable DNA replication. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5443–5447. doi: 10.1073/pnas.74.12.5443. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Ferdinand F. J., Brown M., Khoury G. Synthesis and characterization of late lytic simian virus 40 RNA from transcriptional complexes. Virology. 1977 May 1;78(1):150–161. doi: 10.1016/0042-6822(77)90087-3. [DOI] [PubMed] [Google Scholar]
  16. Fernandez-Munoz R., Coca-Prados M., Hsu M. T. Intracellular forms of simian virus 40 nucleoprotein complexes. I. Methods of isolation and characterization in CV-1 cells. J Virol. 1979 Feb;29(2):612–623. doi: 10.1128/jvi.29.2.612-623.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Garber E. A., Seidman M. M., Levine A. J. The detection and characterization of multiple forms of SV40 nucleoprotein complexes. Virology. 1978 Oct 15;90(2):305–316. doi: 10.1016/0042-6822(78)90315-x. [DOI] [PubMed] [Google Scholar]
  18. Gariglio P., Llopis R., Oudet P., Chambon P. The template of the isolated native simian virus 40 transcriptional complexes is a minichromosome. J Mol Biol. 1979 Jun 15;131(1):75–105. doi: 10.1016/0022-2836(79)90302-4. [DOI] [PubMed] [Google Scholar]
  19. Gariglio P., Mousset S. Characterization of a soluble simian-virus-40 transcription complex. Eur J Biochem. 1977 Jun 15;76(2):583–590. doi: 10.1111/j.1432-1033.1977.tb11628.x. [DOI] [PubMed] [Google Scholar]
  20. Gariglio P., Mousset S. Isolation and partial characterization of a nuclear RNA polymerase - SV40 DNA complex. FEBS Lett. 1975 Aug 1;56(1):149–155. doi: 10.1016/0014-5793(75)80130-x. [DOI] [PubMed] [Google Scholar]
  21. Ghosh P. K., Reddy V. B., Swinscoe J., Lebowitz P., Weissman S. M. Heterogeneity and 5'-terminal structures of the late RNAs of simian virus 40. J Mol Biol. 1978 Dec 25;126(4):813–846. doi: 10.1016/0022-2836(78)90022-0. [DOI] [PubMed] [Google Scholar]
  22. Green M. H., Brooks T. L. Isolation of two forms of SV40 nucleoprotein containing RNA polymerase from infected monkey cells. Virology. 1976 Jul 1;72(1):110–120. doi: 10.1016/0042-6822(76)90316-0. [DOI] [PubMed] [Google Scholar]
  23. Green M. H., Brooks T. L. The sv40 transcription complex. II. Non-dissociation of protein from SV40 chromatin during transcription. Nucleic Acids Res. 1977 Dec;4(12):4279–4289. doi: 10.1093/nar/4.12.4279. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Griffith J., Dieckmann M., Berg P. Electron microscope localization of a protein bound near the origin of simian virus 40 DNA replication. J Virol. 1975 Jan;15(1):167–172. doi: 10.1128/jvi.15.1.167-172.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. 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]
  26. Jakobovits E. B., Aloni Y. Isolation and characterization of various forms of simian virus 40 DNA-protein complexes. Virology. 1980 Apr 15;102(1):107–118. doi: 10.1016/0042-6822(80)90074-4. [DOI] [PubMed] [Google Scholar]
  27. Jakobovits E. B., Bratosin S., Aloni Y. A nucleosome-free region in SV40 minichromosomes. Nature. 1980 May 22;285(5762):263–265. doi: 10.1038/285263a0. [DOI] [PubMed] [Google Scholar]
  28. Khoury G., May E. Regulation of early and late simian virus 40 transcription: overproduction of early viral RNA in the absence of a functional T-antigen. J Virol. 1977 Jul;23(1):167–176. doi: 10.1128/jvi.23.1.167-176.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. La Bella F., Vesco C. Late modifications of simian virus 40 chromatin during the lytic cycle occur in an immature form of virion. J Virol. 1980 Mar;33(3):1138–1150. doi: 10.1128/jvi.33.3.1138-1150.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Lai C. J., Nathans D. A map of temperature-sensitive mutants of simian virus 40. Virology. 1975 Jul;66(1):70–81. doi: 10.1016/0042-6822(75)90179-8. [DOI] [PubMed] [Google Scholar]
  31. Laub O., Aloni Y. Transcription of simian virus 40. VI. SV 40 DNA-RNA polymerase complex isolated from productively infected cells transcribed in vitro. Virology. 1976 Dec;75(2):346–354. doi: 10.1016/0042-6822(76)90033-7. [DOI] [PubMed] [Google Scholar]
  32. Laub O., Bratosin S., Horowitz M., Aloni Y. The initiation of transcription of SV40 DNA at late time after infection. Virology. 1979 Jan 30;92(2):310–323. doi: 10.1016/0042-6822(79)90136-3. [DOI] [PubMed] [Google Scholar]
  33. Laub O., Jakobovits E. B., Aloni Y. 5,6-dichloro-1-beta-ribofuranosylbenzimidazole enhances premature termination of late transcription of simian virus 40 DNA. Proc Natl Acad Sci U S A. 1980 Jun;77(6):3297–3301. doi: 10.1073/pnas.77.6.3297. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Llopis R., Perrin F., Bellard F., Gariglio P. Quantitation of transcribing native simian virus 40 minichromosomes extracted from CV1 cells late in infection. J Virol. 1981 Apr;38(1):82–90. doi: 10.1128/jvi.38.1.82-90.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Mark D. F., Berg P. A third splice site in SV40 early mRNA. Cold Spring Harb Symp Quant Biol. 1980;44(Pt 1):55–62. doi: 10.1101/sqb.1980.044.01.008. [DOI] [PubMed] [Google Scholar]
  36. Meilhac M., Tysper Z., Chambon P. Animal DNA-dependent RNA polymerases. 4. Studies on inhibition by rifamycin derivatives. Eur J Biochem. 1972 Jul 13;28(2):291–300. doi: 10.1111/j.1432-1033.1972.tb01913.x. [DOI] [PubMed] [Google Scholar]
  37. Mertz J. E., Berg P. Defective simian virus 40 genomes: isolation and growth of individual clones. Virology. 1974 Nov;62(1):112–124. doi: 10.1016/0042-6822(74)90307-9. [DOI] [PubMed] [Google Scholar]
  38. 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]
  39. Ptashne M., Jeffrey A., Johnson A. D., Maurer R., Meyer B. J., Pabo C. O., Roberts T. M., Sauer R. T. How the lambda repressor and cro work. Cell. 1980 Jan;19(1):1–11. doi: 10.1016/0092-8674(80)90383-9. [DOI] [PubMed] [Google Scholar]
  40. Reed S. I., Ferguson J., Davis R. W., Stark G. R. T antigen binds to simian virus 40 DNA at the origin of replication. Proc Natl Acad Sci U S A. 1975 Apr;72(4):1605–1609. doi: 10.1073/pnas.72.4.1605. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Reed S. I., Stark G. R., Alwine J. C. Autoregulation of simian virus 40 gene A by T antigen. Proc Natl Acad Sci U S A. 1976 Sep;73(9):3083–3087. doi: 10.1073/pnas.73.9.3083. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Reiser J., Renart J., Crawford L. V., Stark G. R. Specific association of simian virus 40 tumor antigen with simian virus 40 chromatin. J Virol. 1980 Jan;33(1):78–87. doi: 10.1128/jvi.33.1.78-87.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Renart J., Reiser J., Stark G. R. Transfer of proteins from gels to diazobenzyloxymethyl-paper and detection with antisera: a method for studying antibody specificity and antigen structure. Proc Natl Acad Sci U S A. 1979 Jul;76(7):3116–3120. doi: 10.1073/pnas.76.7.3116. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Robb J. A., Martin R. G. Genetic analysis of simian virus 40. 3. Characterization of a temperature-sensitive mutant blocked at an early stage of productive infection in monkey cells. J Virol. 1972 Jun;9(6):956–968. doi: 10.1128/jvi.9.6.956-968.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Saragosti S., Moyne G., Yaniv M. Absence of nucleosomes in a fraction of SV40 chromatin between the origin of replication and the region coding for the late leader RNA. Cell. 1980 May;20(1):65–73. doi: 10.1016/0092-8674(80)90235-4. [DOI] [PubMed] [Google Scholar]
  46. Segawa M., Sugano S., Yamaguchi N. Association of simian virus 40 T antigen with replicating nucleoprotein complexes of simian virus 40. J Virol. 1980 Aug;35(2):320–330. doi: 10.1128/jvi.35.2.320-330.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Shalloway D., Kleinberger T., Livingston D. M. Mapping of SV40 DNA replication origin region binding sites for the SV40 T antigen by protection against exonuclease III digestion. Cell. 1980 Jun;20(2):411–422. doi: 10.1016/0092-8674(80)90627-3. [DOI] [PubMed] [Google Scholar]
  48. Shani M., Birkenmeier E., May E., Salzman N. P. Properties of simian virus 40 transcriptional intermediates isolated from nuclei of permissive cells. J Virol. 1977 Jul;23(1):20–28. doi: 10.1128/jvi.23.1.20-28.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Takemoto K. K., Kirschstein R. L., Habel K. Mutants of simian virus 40 differing in plaque size, oncogenicity, and heat sensitivity. J Bacteriol. 1966 Oct;92(4):990–994. doi: 10.1128/jb.92.4.990-994.1966. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Tegtmeyer P., Rundell K., Collins J. K. Modification of simian virus 40 protein A. J Virol. 1977 Feb;21(2):647–657. doi: 10.1128/jvi.21.2.647-657.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Tegtmeyer P., Schwartz M., Collins J. K., Rundell K. Regulation of tumor antigen synthesis by simain virus 40 gene A. J Virol. 1975 Jul;16(1):168–178. doi: 10.1128/jvi.16.1.168-178.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Tegtmeyer P. Simian virus 40 deoxyribonucleic acid synthesis: the viral replicon. J Virol. 1972 Oct;10(4):591–598. doi: 10.1128/jvi.10.4.591-598.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Tjian R. The binding site on SV40 DNA for a T antigen-related protein. Cell. 1978 Jan;13(1):165–179. doi: 10.1016/0092-8674(78)90147-2. [DOI] [PubMed] [Google Scholar]
  54. Van Heuverswyn H., Cole C., Berg P., Fiers W. Nucleotide sequence analysis of two simian virus 40 mutants with deletions in the region coding for the carboxyl terminus of the T antigen. J Virol. 1979 Jun;30(3):936–941. doi: 10.1128/jvi.30.3.936-941.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Van Heuverswyn H., Fiers W. Nucleotide sequence of the Hind-C fragment of simian virus 40 DNA. Comparison of the 5'-untranslated region of wild-type virus and of some deletion Mutants. Eur J Biochem. 1979 Oct;100(1):51–60. doi: 10.1111/j.1432-1033.1979.tb02032.x. [DOI] [PubMed] [Google Scholar]
  56. Varshavsky A. J., Sundin O., Bohn M. A stretch of "late" SV40 viral DNA about 400 bp long which includes the origin of replication is specifically exposed in SV40 minichromosomes. Cell. 1979 Feb;16(2):453–466. doi: 10.1016/0092-8674(79)90021-7. [DOI] [PubMed] [Google Scholar]
  57. Volckaert G., Feunteun J., Crawford L. V., Berg P., Fiers W. Nucleotide sequence deletions within the coding region for small-t antigen of simian virus 40. J Virol. 1979 Jun;30(3):674–682. doi: 10.1128/jvi.30.3.674-682.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  58. Wasylyk B., Chambon P. Transcription by eukaryotic RNA polymerases A and B of chromatin assembled in vitro. Eur J Biochem. 1979 Aug 1;98(2):317–327. doi: 10.1111/j.1432-1033.1979.tb13191.x. [DOI] [PubMed] [Google Scholar]

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