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. 1991 May;65(5):2415–2421. doi: 10.1128/jvi.65.5.2415-2421.1991

A single-stranded gap in human immunodeficiency virus unintegrated linear DNA defined by a central copy of the polypurine tract.

P Charneau 1, F Clavel 1
PMCID: PMC240594  PMID: 2016765

Abstract

The structure of unintegrated human immunodeficiency virus type 1 (HIV-1) DNA from acutely infected human lymphoid cells was analyzed by nuclease S1 cleavage. We observed a unique, discrete single-stranded gap in unintegrated linear DNA molecules, located near the center of the genome. Oligonucleotide primer extension experiments determined that the downstream limit of this gap coincides with the last nucleotide of a central copy of the polypurine tract found in all sequenced lentivirus genomes. Other retroviruses have only one copy of the polypurine tract at the 5' boundary of the 3' long terminal repeat, which has been shown to determine initiation of retroviral DNA plus-strand synthesis. We conclude from our observations that the central repeat of the polypurine tract can create an additional site for plus-strand synthesis initiation in lentiviruses. The central single-stranded gap was not found in circular DNA molecules, the vast majority of them carrying only one long terminal repeat. This finding suggests that the generation of such circular molecules is associated with early DNA ligation events.

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Selected References

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

  1. Adachi A., Gendelman H. E., Koenig S., Folks T., Willey R., Rabson A., Martin M. A. Production of acquired immunodeficiency syndrome-associated retrovirus in human and nonhuman cells transfected with an infectious molecular clone. J Virol. 1986 Aug;59(2):284–291. doi: 10.1128/jvi.59.2.284-291.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Barré-Sinoussi F., Chermann J. C., Rey F., Nugeyre M. T., Chamaret S., Gruest J., Dauguet C., Axler-Blin C., Vézinet-Brun F., Rouzioux C. Isolation of a T-lymphotropic retrovirus from a patient at risk for acquired immune deficiency syndrome (AIDS). Science. 1983 May 20;220(4599):868–871. doi: 10.1126/science.6189183. [DOI] [PubMed] [Google Scholar]
  3. Blum H. E., Harris J. D., Ventura P., Walker D., Staskus K., Retzel E., Haase A. T. Synthesis in cell culture of the gapped linear duplex DNA of the slow virus visna. Virology. 1985 Apr 30;142(2):270–277. doi: 10.1016/0042-6822(85)90335-6. [DOI] [PubMed] [Google Scholar]
  4. Boone L. R., Skalka A. M. Viral DNA synthesized in vitro by avian retrovirus particles permeabilized with melittin. II. Evidence for a strand displacement mechanism in plus-strand synthesis. J Virol. 1981 Jan;37(1):117–126. doi: 10.1128/jvi.37.1.117-126.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Chakrabarti L., Guyader M., Alizon M., Daniel M. D., Desrosiers R. C., Tiollais P., Sonigo P. Sequence of simian immunodeficiency virus from macaque and its relationship to other human and simian retroviruses. Nature. 1987 Aug 6;328(6130):543–547. doi: 10.1038/328543a0. [DOI] [PubMed] [Google Scholar]
  6. Feinberg A. P., Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. doi: 10.1016/0003-2697(83)90418-9. [DOI] [PubMed] [Google Scholar]
  7. Finston W. I., Champoux J. J. RNA-primed initiation of Moloney murine leukemia virus plus strands by reverse transcriptase in vitro. J Virol. 1984 Jul;51(1):26–33. doi: 10.1128/jvi.51.1.26-33.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Friedrich R., Moelling K. Effect of viral RNase H on the avian sarcoma viral genome during early transcription in vitro. J Virol. 1979 Sep;31(3):630–638. doi: 10.1128/jvi.31.3.630-638.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Fritsch E. F., Temin H. M. Inhibition of viral DNA synthesis in stationary chicken embryo fibroblasts infected with avian retroviruses. J Virol. 1977 Nov;24(2):461–469. doi: 10.1128/jvi.24.2.461-469.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Gilboa E., Mitra S. W., Goff S., Baltimore D. A detailed model of reverse transcription and tests of crucial aspects. Cell. 1979 Sep;18(1):93–100. doi: 10.1016/0092-8674(79)90357-x. [DOI] [PubMed] [Google Scholar]
  11. Grandgenett D. P., Gerard G. F., Green M. Ribonuclease H: a ubiquitous activity in virions of ribonucleic acid tumor viruses. J Virol. 1972 Dec;10(6):1136–1142. doi: 10.1128/jvi.10.6.1136-1142.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Guyader M., Emerman M., Sonigo P., Clavel F., Montagnier L., Alizon M. Genome organization and transactivation of the human immunodeficiency virus type 2. Nature. 1987 Apr 16;326(6114):662–669. doi: 10.1038/326662a0. [DOI] [PubMed] [Google Scholar]
  13. Harada S., Koyanagi Y., Yamamoto N. Infection of HTLV-III/LAV in HTLV-I-carrying cells MT-2 and MT-4 and application in a plaque assay. Science. 1985 Aug 9;229(4713):563–566. doi: 10.1126/science.2992081. [DOI] [PubMed] [Google Scholar]
  14. Harris J. D., Scott J. V., Traynor B., Brahic M., Stowring L., Ventura P., Haase A. T., Peluso R. Visna virus DNA: discovery of a novel gapped structure. Virology. 1981 Sep;113(2):573–583. doi: 10.1016/0042-6822(81)90185-9. [DOI] [PubMed] [Google Scholar]
  15. 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]
  16. Hsu T. W., Taylor J. M. Single-stranded regions on unintegrated avian retrovirus DNA. J Virol. 1982 Oct;44(1):47–53. doi: 10.1128/jvi.44.1.47-53.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Kim S. Y., Byrn R., Groopman J., Baltimore D. Temporal aspects of DNA and RNA synthesis during human immunodeficiency virus infection: evidence for differential gene expression. J Virol. 1989 Sep;63(9):3708–3713. doi: 10.1128/jvi.63.9.3708-3713.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Kung H. J., Fung Y. K., Majors J. E., Bishop J. M., Varmus H. E. Synthesis of plus strands of retroviral DNA in cells infected with avian sarcoma virus and mouse mammary tumor virus. J Virol. 1981 Jan;37(1):127–138. doi: 10.1128/jvi.37.1.127-138.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Mitra S. W., Chow M., Champoux J., Baltimore D. Synthesis of murine leukemia virus plus strong stop DNA initiates at a unique site. J Biol Chem. 1982 Jun 10;257(11):5983–5986. [PubMed] [Google Scholar]
  20. Mitra S. W., Goff S., Gilboa E., Baltimore D. Synthesis of a 600-nucleotide-long plus-strand DNA by virions of Moloney murine leukemia virus. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4355–4359. doi: 10.1073/pnas.76.9.4355. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Méchali M., de Recondo A. M., Girard M. Action of the S1 endonuclease from Aspergillus oryzae on simian virus 40 supercoiled component I DNA. Biochem Biophys Res Commun. 1973 Oct 15;54(4):1306–1320. doi: 10.1016/0006-291x(73)91130-3. [DOI] [PubMed] [Google Scholar]
  22. Olsen J. C., Watson K. F. Reverse transcription of avian myeloblastosis virus 35S RNA. Early synthesis of plus strand DNA of discrete size in reconstructed reactions. Nucleic Acids Res. 1982 Feb 11;10(3):1009–1027. doi: 10.1093/nar/10.3.1009. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Omer C. A., Resnick R., Faras A. J. Evidence for involvement of an RNA primer in initiation of strong-stop plus DNA synthesis during reverse transcription in vitro. J Virol. 1984 May;50(2):465–470. doi: 10.1128/jvi.50.2.465-470.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Panganiban A. T., Fiore D. Ordered interstrand and intrastrand DNA transfer during reverse transcription. Science. 1988 Aug 26;241(4869):1064–1069. doi: 10.1126/science.2457948. [DOI] [PubMed] [Google Scholar]
  25. Pfeiffer P., Hohn T. Involvement of reverse transcription in the replication of cauliflower mosaic virus: a detailed model and test of some aspects. Cell. 1983 Jul;33(3):781–789. doi: 10.1016/0092-8674(83)90020-x. [DOI] [PubMed] [Google Scholar]
  26. Rattray A. J., Champoux J. J. The role of Moloney murine leukemia virus RNase H activity in the formation of plus-strand primers. J Virol. 1987 Sep;61(9):2843–2851. doi: 10.1128/jvi.61.9.2843-2851.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Resnick R., Omer C. A., Faras A. J. Involvement of retrovirus reverse transcriptase-associated RNase H in the initiation of strong-stop (+) DNA synthesis and the generation of the long terminal repeat. J Virol. 1984 Sep;51(3):813–821. doi: 10.1128/jvi.51.3.813-821.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Rey M. A., Krust B., Laurent A. G., Montagnier L., Hovanessian A. G. Characterization of human immunodeficiency virus type 2 envelope glycoproteins: dimerization of the glycoprotein precursor during processing. J Virol. 1989 Feb;63(2):647–658. doi: 10.1128/jvi.63.2.647-658.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Shank P. R., Hughes S. H., Kung H. J., Majors J. E., Quintrell N., Guntaka R. V., Bishop J. M., Varmus H. E. Mapping unintegrated avian sarcoma virus DNA: termini of linear DNA bear 300 nucleotides present once or twice in two species of circular DNA. Cell. 1978 Dec;15(4):1383–1395. doi: 10.1016/0092-8674(78)90063-6. [DOI] [PubMed] [Google Scholar]
  31. Shank P. R., Varmus H. E. Virus-specific DNA in the cytoplasm of avian sarcoma virus-infected cells is a precursor to covalently closed circular viral DNA in the nucleus. J Virol. 1978 Jan;25(1):104–104. doi: 10.1128/jvi.25.1.104-104.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Smith J. K., Cywinski A., Taylor J. M. Initiation of plus-strand DNA synthesis during reverse transcription of an avian retrovirus genome. J Virol. 1984 Jan;49(1):200–204. doi: 10.1128/jvi.49.1.200-204.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Sonigo P., Alizon M., Staskus K., Klatzmann D., Cole S., Danos O., Retzel E., Tiollais P., Haase A., Wain-Hobson S. Nucleotide sequence of the visna lentivirus: relationship to the AIDS virus. Cell. 1985 Aug;42(1):369–382. doi: 10.1016/s0092-8674(85)80132-x. [DOI] [PubMed] [Google Scholar]
  34. 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]
  35. Staskus K. A., Collett M. S., Faras A. J. Initiation of DNA synthesis by the avian oncornavirus RNA-directed DNA polymerase: structural and functional localization of the major species of primer RNA on the oncornavirus genome. Virology. 1976 May;71(1):162–168. doi: 10.1016/0042-6822(76)90102-1. [DOI] [PubMed] [Google Scholar]
  36. Taylor J. M., Illmensee R. Site on the RNA of an avian sarcoma virus at which primer is bound. J Virol. 1975 Sep;16(3):553–558. doi: 10.1128/jvi.16.3.553-558.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Temin H. M. Structure, variation and synthesis of retrovirus long terminal repeat. Cell. 1981 Nov;27(1 Pt 2):1–3. doi: 10.1016/0092-8674(81)90353-6. [DOI] [PubMed] [Google Scholar]
  38. Varmus H. E., Heasley S., Kung H. J., Oppermann H., Smith V. C., Bishop J. M., Shank P. R. Kinetics of synthesis, structure and purification of avian sarcoma virus-specific DNA made in the cytoplasm of acutely infected cells. J Mol Biol. 1978 Mar 25;120(1):55–82. doi: 10.1016/0022-2836(78)90295-4. [DOI] [PubMed] [Google Scholar]
  39. Varmus H. E., Padgett T., Heasley S., Simon G., Bishop J. M. Cellular functions are required for the synthesis and integration of avian sarcoma virus-specific DNA. Cell. 1977 Jun;11(2):307–319. doi: 10.1016/0092-8674(77)90047-2. [DOI] [PubMed] [Google Scholar]
  40. Verma I. M. Studies on reverse transcriptase of RNA tumor viruses III. Properties of purified Moloney murine leukemia virus DNA polymerase and associated RNase H. J Virol. 1975 Apr;15(4):843–854. doi: 10.1128/jvi.15.4.843-854.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Volovitch M., Drugeon C., Yot P. Studies on the single-stranded discontinuities of the cauliflower mosaic virus genome. Nucleic Acids Res. 1978 Aug;5(8):2913–2925. doi: 10.1093/nar/5.8.2913. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Wain-Hobson S., Sonigo P., Danos O., Cole S., Alizon M. Nucleotide sequence of the AIDS virus, LAV. Cell. 1985 Jan;40(1):9–17. doi: 10.1016/0092-8674(85)90303-4. [DOI] [PubMed] [Google Scholar]
  43. Zack J. A., Arrigo S. J., Weitsman S. R., Go A. S., Haislip A., Chen I. S. HIV-1 entry into quiescent primary lymphocytes: molecular analysis reveals a labile, latent viral structure. Cell. 1990 Apr 20;61(2):213–222. doi: 10.1016/0092-8674(90)90802-l. [DOI] [PubMed] [Google Scholar]

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