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. 1997 Nov 3;16(21):6603–6612. doi: 10.1093/emboj/16.21.6603

Plus-strand strong-stop DNA transfer in yeast Ty retrotransposons.

V Lauermann 1, J D Boeke 1
PMCID: PMC1170264  PMID: 9351840

Abstract

The yeast Ty1 LTR retrotransposon replicates by reverse transcription and integration; the process shows many similarities to the retroviral life cycle. However, we show that plus strand strong-stop DNA transfer in yeast Ty1 elements differs from the analogous retroviral process. By analysis of the native structure of the Ty1 primer binding site and by a series of manipulations of this region and assessment of the effects on retrotransposition, we show that primer binding site inheritance is not from the tRNA primer, which is inconsistent with classical retroviral models. This unusual inheritance pattern holds even when the Ty1 primer binding site is lengthened in order to be more retrovirus-like. Finally, the distantly related Ty3 element has an inheritance pattern like Ty1, indicating evolutionary conservation of the alternative pathway used by Ty1. Based on these results we arrive at a plus strand primer recycling model that explains Ty1 plus strand strong-stop DNA transfer and inheritance patterns in the primer binding site.

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Selected References

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

  1. Berwin B., Barklis E. Retrovirus-mediated insertion of expressed and non-expressed genes at identical chromosomal locations. Nucleic Acids Res. 1993 May 25;21(10):2399–2407. doi: 10.1093/nar/21.10.2399. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bishop D. K., Andersen J., Kolodner R. D. Specificity of mismatch repair following transformation of Saccharomyces cerevisiae with heteroduplex plasmid DNA. Proc Natl Acad Sci U S A. 1989 May;86(10):3713–3717. doi: 10.1073/pnas.86.10.3713. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Blain S. W., Goff S. P. Effects on DNA synthesis and translocation caused by mutations in the RNase H domain of Moloney murine leukemia virus reverse transcriptase. J Virol. 1995 Jul;69(7):4440–4452. doi: 10.1128/jvi.69.7.4440-4452.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Boeke J. D., Eichinger D., Castrillon D., Fink G. R. The Saccharomyces cerevisiae genome contains functional and nonfunctional copies of transposon Ty1. Mol Cell Biol. 1988 Apr;8(4):1432–1442. doi: 10.1128/mcb.8.4.1432. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Boeke J. D., Garfinkel D. J., Styles C. A., Fink G. R. Ty elements transpose through an RNA intermediate. Cell. 1985 Mar;40(3):491–500. doi: 10.1016/0092-8674(85)90197-7. [DOI] [PubMed] [Google Scholar]
  6. Boeke J. D., LaCroute F., Fink G. R. A positive selection for mutants lacking orotidine-5'-phosphate decarboxylase activity in yeast: 5-fluoro-orotic acid resistance. Mol Gen Genet. 1984;197(2):345–346. doi: 10.1007/BF00330984. [DOI] [PubMed] [Google Scholar]
  7. Boeke J. D., Styles C. A., Fink G. R. Saccharomyces cerevisiae SPT3 gene is required for transposition and transpositional recombination of chromosomal Ty elements. Mol Cell Biol. 1986 Nov;6(11):3575–3581. doi: 10.1128/mcb.6.11.3575. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Boeke J. D., Xu H., Fink G. R. A general method for the chromosomal amplification of genes in yeast. Science. 1988 Jan 15;239(4837):280–282. doi: 10.1126/science.2827308. [DOI] [PubMed] [Google Scholar]
  9. Chalker D. L., Sandmeyer S. B. Transfer RNA genes are genomic targets for de Novo transposition of the yeast retrotransposon Ty3. Genetics. 1990 Dec;126(4):837–850. doi: 10.1093/genetics/126.4.837. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Chapman K. B., Byström A. S., Boeke J. D. Initiator methionine tRNA is essential for Ty1 transposition in yeast. Proc Natl Acad Sci U S A. 1992 Apr 15;89(8):3236–3240. doi: 10.1073/pnas.89.8.3236. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Charneau P., Alizon M., Clavel F. A second origin of DNA plus-strand synthesis is required for optimal human immunodeficiency virus replication. J Virol. 1992 May;66(5):2814–2820. doi: 10.1128/jvi.66.5.2814-2820.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Curcio M. J., Garfinkel D. J. Heterogeneous functional Ty1 elements are abundant in the Saccharomyces cerevisiae genome. Genetics. 1994 Apr;136(4):1245–1259. doi: 10.1093/genetics/136.4.1245. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. 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]
  14. Friant S., Heyman T., Poch O., Wilhelm M., Wilhelm F. X. Sequence comparison of the Ty1 and Ty2 elements of the yeast genome supports the structural model of the tRNAiMet-Ty1 RNA reverse transcription initiation complex. Yeast. 1997 Jun 15;13(7):639–645. doi: 10.1002/(SICI)1097-0061(19970615)13:7<639::AID-YEA143>3.0.CO;2-Y. [DOI] [PubMed] [Google Scholar]
  15. Friant S., Heyman T., Wilhelm M. L., Wilhelm F. X. Extended interactions between the primer tRNAi(Met) and genomic RNA of the yeast Ty1 retrotransposon. Nucleic Acids Res. 1996 Feb 1;24(3):441–449. doi: 10.1093/nar/24.3.441. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Gabriel A., Willems M., Mules E. H., Boeke J. D. Replication infidelity during a single cycle of Ty1 retrotransposition. Proc Natl Acad Sci U S A. 1996 Jul 23;93(15):7767–7771. doi: 10.1073/pnas.93.15.7767. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Garfinkel D. J., Boeke J. D., Fink G. R. Ty element transposition: reverse transcriptase and virus-like particles. Cell. 1985 Sep;42(2):507–517. doi: 10.1016/0092-8674(85)90108-4. [DOI] [PubMed] [Google Scholar]
  18. Hansen L. J., Sandmeyer S. B. Characterization of a transpositionally active Ty3 element and identification of the Ty3 integrase protein. J Virol. 1990 Jun;64(6):2599–2607. doi: 10.1128/jvi.64.6.2599-2607.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Heyman T., Agoutin B., Friant S., Wilhelm F. X., Wilhelm M. L. Plus-strand DNA synthesis of the yeast retrotransposon Ty1 is initiated at two sites, PPT1 next to the 3' LTR and PPT2 within the pol gene. PPT1 is sufficient for Ty1 transposition. J Mol Biol. 1995 Oct 20;253(2):291–303. doi: 10.1006/jmbi.1995.0553. [DOI] [PubMed] [Google Scholar]
  20. Hu W. S., Temin H. M. Retroviral recombination and reverse transcription. Science. 1990 Nov 30;250(4985):1227–1233. doi: 10.1126/science.1700865. [DOI] [PubMed] [Google Scholar]
  21. Huber H. E., Richardson C. C. Processing of the primer for plus strand DNA synthesis by human immunodeficiency virus 1 reverse transcriptase. J Biol Chem. 1990 Jun 25;265(18):10565–10573. [PubMed] [Google Scholar]
  22. Keeney J. B., Chapman K. B., Lauermann V., Voytas D. F., Aström S. U., von Pawel-Rammingen U., Byström A., Boeke J. D. Multiple molecular determinants for retrotransposition in a primer tRNA. Mol Cell Biol. 1995 Jan;15(1):217–226. doi: 10.1128/mcb.15.1.217. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Kinsey P. T., Sandmeyer S. B. Ty3 transposes in mating populations of yeast: a novel transposition assay for Ty3. Genetics. 1995 Jan;139(1):81–94. doi: 10.1093/genetics/139.1.81. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Kulpa D., Topping R., Telesnitsky A. Determination of the site of first strand transfer during Moloney murine leukemia virus reverse transcription and identification of strand transfer-associated reverse transcriptase errors. EMBO J. 1997 Feb 17;16(4):856–865. doi: 10.1093/emboj/16.4.856. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Lauermann V., Boeke J. D. The primer tRNA sequence is not inherited during Ty1 retrotransposition. Proc Natl Acad Sci U S A. 1994 Oct 11;91(21):9847–9851. doi: 10.1073/pnas.91.21.9847. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Lauermann V., Hermankova M., Boeke J. D. Increased length of long terminal repeats inhibits Ty1 transposition and leads to the formation of tandem multimers. Genetics. 1997 Apr;145(4):911–922. doi: 10.1093/genetics/145.4.911. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Lauermann V., Nam K., Trambley J., Boeke J. D. Plus-strand strong-stop DNA synthesis in retrotransposon Ty1. J Virol. 1995 Dec;69(12):7845–7850. doi: 10.1128/jvi.69.12.7845-7850.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Mellor J., Malim M. H., Gull K., Tuite M. F., McCready S., Dibbayawan T., Kingsman S. M., Kingsman A. J. Reverse transcriptase activity and Ty RNA are associated with virus-like particles in yeast. Nature. 1985 Dec 12;318(6046):583–586. doi: 10.1038/318583a0. [DOI] [PubMed] [Google Scholar]
  29. Moore S. P., Powers M., Garfinkel D. J. Substrate specificity of Ty1 integrase. J Virol. 1995 Aug;69(8):4683–4692. doi: 10.1128/jvi.69.8.4683-4692.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Omer C. A., Faras A. J. Mechanism of release of the avian rotavirus tRNATrp primer molecule from viral DNA by ribonuclease H during reverse transcription. Cell. 1982 Oct;30(3):797–805. doi: 10.1016/0092-8674(82)90284-7. [DOI] [PubMed] [Google Scholar]
  31. 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]
  32. Paquin C. E., Williamson V. M. Temperature effects on the rate of ty transposition. Science. 1984 Oct 5;226(4670):53–55. doi: 10.1126/science.226.4670.53. [DOI] [PubMed] [Google Scholar]
  33. Pochart P., Agoutin B., Rousset S., Chanet R., Doroszkiewicz V., Heyman T. Biochemical and electron microscope analyses of the DNA reverse transcripts present in the virus-like particles of the yeast transposon Ty1. Identification of a second origin of Ty1DNA plus strand synthesis. Nucleic Acids Res. 1993 Jul 25;21(15):3513–3520. doi: 10.1093/nar/21.15.3513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Rattray A. J., Champoux J. J. Plus-strand priming by Moloney murine leukemia virus. The sequence features important for cleavage by RNase H. J Mol Biol. 1989 Aug 5;208(3):445–456. doi: 10.1016/0022-2836(89)90508-1. [DOI] [PubMed] [Google Scholar]
  35. 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]
  36. Scherer S., Mann C., Davis R. W. Reversion of a promoter deletion in yeast. Nature. 1982 Aug 26;298(5877):815–819. doi: 10.1038/298815a0. [DOI] [PubMed] [Google Scholar]
  37. Sharon G., Burkett T. J., Garfinkel D. J. Efficient homologous recombination of Ty1 element cDNA when integration is blocked. Mol Cell Biol. 1994 Oct;14(10):6540–6551. doi: 10.1128/mcb.14.10.6540. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Smith C. M., Potts W. B., 3rd, Smith J. S., Roth M. J. RNase H cleavage of tRNAPro mediated by M-MuLV and HIV-1 reverse transcriptases. Virology. 1997 Mar 17;229(2):437–446. doi: 10.1006/viro.1997.8454. [DOI] [PubMed] [Google Scholar]
  39. Struhl K. Nucleotide sequence and transcriptional mapping of the yeast pet56-his3-ded1 gene region. Nucleic Acids Res. 1985 Dec 9;13(23):8587–8601. doi: 10.1093/nar/13.23.8587. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Temin H. M. Retrovirus variation and reverse transcription: abnormal strand transfers result in retrovirus genetic variation. Proc Natl Acad Sci U S A. 1993 Aug 1;90(15):6900–6903. doi: 10.1073/pnas.90.15.6900. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Wakefield J. K., Kang S. M., Morrow C. D. Construction of a type 1 human immunodeficiency virus that maintains a primer binding site complementary to tRNA(His). J Virol. 1996 Feb;70(2):966–975. doi: 10.1128/jvi.70.2.966-975.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Wakefield J. K., Wolf A. G., Morrow C. D. Human immunodeficiency virus type 1 can use different tRNAs as primers for reverse transcription but selectively maintains a primer binding site complementary to tRNA(3Lys). J Virol. 1995 Oct;69(10):6021–6029. doi: 10.1128/jvi.69.10.6021-6029.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Whitcomb J. M., Kumar R., Hughes S. H. Sequence of the circle junction of human immunodeficiency virus type 1: implications for reverse transcription and integration. J Virol. 1990 Oct;64(10):4903–4906. doi: 10.1128/jvi.64.10.4903-4906.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Wilhelm M., Heyman T., Friant S., Wilhelm F. X. Heterogeneous terminal structure of Ty1 and Ty3 reverse transcripts. Nucleic Acids Res. 1997 Jun 1;25(11):2161–2166. doi: 10.1093/nar/25.11.2161. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Xiong Y., Eickbush T. H. Origin and evolution of retroelements based upon their reverse transcriptase sequences. EMBO J. 1990 Oct;9(10):3353–3362. doi: 10.1002/j.1460-2075.1990.tb07536.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Xu H., Boeke J. D. High-frequency deletion between homologous sequences during retrotransposition of Ty elements in Saccharomyces cerevisiae. Proc Natl Acad Sci U S A. 1987 Dec;84(23):8553–8557. doi: 10.1073/pnas.84.23.8553. [DOI] [PMC free article] [PubMed] [Google Scholar]

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