Skip to main content
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1998 Mar 15;26(6):1487–1494. doi: 10.1093/nar/26.6.1487

Negative regulation of yeast telomerase activity through an interaction with an upstream region of the DNA primer.

N F Lue 1, Y Peng 1
PMCID: PMC147436  PMID: 9490796

Abstract

A number of published studies indicate that telomerase may interact with oligonucleotide primers in a bipartite manner, with the 3'-end of the primer positioned at the catalytic site of the enzyme and a more 5' region of the primer binding to a second or 'anchor' site of the enzyme. We systematically investigated the effects of mutations in the DNA primer on overall binding and polymerization by yeast telomerase. Our studies indicate that there is sequence-specific interaction between telomerase and a substantial region of the DNA primer. Mutations in the 3'-most positions of the primer reduced polymerization, yet had little effect on overall binding affinity. In contrast, mutations around the -20 position reduced binding affinity but had no effect on polymerization. Most strikingly, mutations centered around the -12 position of the DNA primer reduced overall binding affinity but dramatically enhanced primer extension, as well as primer cleavage. This finding suggests that reduced interaction with the -12 region of the DNA primer can facilitate a step in the catalytic region of yeast telomerase that leads to greater polymerization. A tripartite model of interaction between primer and telomerase is proposed to account for the distinct effects of mutations in different regions of the DNA primer.

Full Text

The Full Text of this article is available as a PDF (480.9 KB).

Selected References

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

  1. Blackburn E. H. Structure and function of telomeres. Nature. 1991 Apr 18;350(6319):569–573. doi: 10.1038/350569a0. [DOI] [PubMed] [Google Scholar]
  2. Blackburn E. H. Telomerases. Annu Rev Biochem. 1992;61:113–129. doi: 10.1146/annurev.bi.61.070192.000553. [DOI] [PubMed] [Google Scholar]
  3. Cohn M., Blackburn E. H. Telomerase in yeast. Science. 1995 Jul 21;269(5222):396–400. doi: 10.1126/science.7618104. [DOI] [PubMed] [Google Scholar]
  4. Collins K., Greider C. W. Tetrahymena telomerase catalyzes nucleolytic cleavage and nonprocessive elongation. Genes Dev. 1993 Jul;7(7B):1364–1376. doi: 10.1101/gad.7.7b.1364. [DOI] [PubMed] [Google Scholar]
  5. Collins K., Kobayashi R., Greider C. W. Purification of Tetrahymena telomerase and cloning of genes encoding the two protein components of the enzyme. Cell. 1995 Jun 2;81(5):677–686. doi: 10.1016/0092-8674(95)90529-4. [DOI] [PubMed] [Google Scholar]
  6. Feng J., Funk W. D., Wang S. S., Weinrich S. L., Avilion A. A., Chiu C. P., Adams R. R., Chang E., Allsopp R. C., Yu J. The RNA component of human telomerase. Science. 1995 Sep 1;269(5228):1236–1241. doi: 10.1126/science.7544491. [DOI] [PubMed] [Google Scholar]
  7. Greider C. W., Blackburn E. H. A telomeric sequence in the RNA of Tetrahymena telomerase required for telomere repeat synthesis. Nature. 1989 Jan 26;337(6205):331–337. doi: 10.1038/337331a0. [DOI] [PubMed] [Google Scholar]
  8. Greider C. W., Blackburn E. H. Identification of a specific telomere terminal transferase activity in Tetrahymena extracts. Cell. 1985 Dec;43(2 Pt 1):405–413. doi: 10.1016/0092-8674(85)90170-9. [DOI] [PubMed] [Google Scholar]
  9. Greider C. W. Telomerase is processive. Mol Cell Biol. 1991 Sep;11(9):4572–4580. doi: 10.1128/mcb.11.9.4572. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Hammond P. W., Lively T. N., Cech T. R. The anchor site of telomerase from Euplotes aediculatus revealed by photo-cross-linking to single- and double-stranded DNA primers. Mol Cell Biol. 1997 Jan;17(1):296–308. doi: 10.1128/mcb.17.1.296. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Harrington L. A., Greider C. W. Telomerase primer specificity and chromosome healing. Nature. 1991 Oct 3;353(6343):451–454. doi: 10.1038/353451a0. [DOI] [PubMed] [Google Scholar]
  12. Harrington L., McPhail T., Mar V., Zhou W., Oulton R., Bass M. B., Arruda I., Robinson M. O. A mammalian telomerase-associated protein. Science. 1997 Feb 14;275(5302):973–977. doi: 10.1126/science.275.5302.973. [DOI] [PubMed] [Google Scholar]
  13. Horikoshi M., Wang C. K., Fujii H., Cromlish J. A., Weil P. A., Roeder R. G. Purification of a yeast TATA box-binding protein that exhibits human transcription factor IID activity. Proc Natl Acad Sci U S A. 1989 Jul;86(13):4843–4847. doi: 10.1073/pnas.86.13.4843. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Lee M. S., Blackburn E. H. Sequence-specific DNA primer effects on telomerase polymerization activity. Mol Cell Biol. 1993 Oct;13(10):6586–6599. doi: 10.1128/mcb.13.10.6586. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Lendvay T. S., Morris D. K., Sah J., Balasubramanian B., Lundblad V. Senescence mutants of Saccharomyces cerevisiae with a defect in telomere replication identify three additional EST genes. Genetics. 1996 Dec;144(4):1399–1412. doi: 10.1093/genetics/144.4.1399. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Lin J. J., Zakian V. A. An in vitro assay for Saccharomyces telomerase requires EST1. Cell. 1995 Jun 30;81(7):1127–1135. doi: 10.1016/s0092-8674(05)80017-0. [DOI] [PubMed] [Google Scholar]
  17. Lingner J., Hughes T. R., Shevchenko A., Mann M., Lundblad V., Cech T. R. Reverse transcriptase motifs in the catalytic subunit of telomerase. Science. 1997 Apr 25;276(5312):561–567. doi: 10.1126/science.276.5312.561. [DOI] [PubMed] [Google Scholar]
  18. Lue N. F., Wang J. C. ATP-dependent processivity of a telomerase activity from Saccharomyces cerevisiae. J Biol Chem. 1995 Sep 15;270(37):21453–21456. doi: 10.1074/jbc.270.37.21453. [DOI] [PubMed] [Google Scholar]
  19. Mantell L. L., Greider C. W. Telomerase activity in germline and embryonic cells of Xenopus. EMBO J. 1994 Jul 1;13(13):3211–3217. doi: 10.1002/j.1460-2075.1994.tb06620.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Melek M., Greene E. C., Shippen D. E. Processing of nontelomeric 3' ends by telomerase: default template alignment and endonucleolytic cleavage. Mol Cell Biol. 1996 Jul;16(7):3437–3445. doi: 10.1128/mcb.16.7.3437. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Morin G. B. Recognition of a chromosome truncation site associated with alpha-thalassaemia by human telomerase. Nature. 1991 Oct 3;353(6343):454–456. doi: 10.1038/353454a0. [DOI] [PubMed] [Google Scholar]
  22. Morin G. B. The human telomere terminal transferase enzyme is a ribonucleoprotein that synthesizes TTAGGG repeats. Cell. 1989 Nov 3;59(3):521–529. doi: 10.1016/0092-8674(89)90035-4. [DOI] [PubMed] [Google Scholar]
  23. Nakayama J., Saito M., Nakamura H., Matsuura A., Ishikawa F. TLP1: a gene encoding a protein component of mammalian telomerase is a novel member of WD repeats family. Cell. 1997 Mar 21;88(6):875–884. doi: 10.1016/s0092-8674(00)81933-9. [DOI] [PubMed] [Google Scholar]
  24. Nugent C. I., Hughes T. R., Lue N. F., Lundblad V. Cdc13p: a single-strand telomeric DNA-binding protein with a dual role in yeast telomere maintenance. Science. 1996 Oct 11;274(5285):249–252. doi: 10.1126/science.274.5285.249. [DOI] [PubMed] [Google Scholar]
  25. Prescott J., Blackburn E. H. Telomerase RNA mutations in Saccharomyces cerevisiae alter telomerase action and reveal nonprocessivity in vivo and in vitro. Genes Dev. 1997 Feb 15;11(4):528–540. doi: 10.1101/gad.11.4.528. [DOI] [PubMed] [Google Scholar]
  26. Prowse K. R., Avilion A. A., Greider C. W. Identification of a nonprocessive telomerase activity from mouse cells. Proc Natl Acad Sci U S A. 1993 Feb 15;90(4):1493–1497. doi: 10.1073/pnas.90.4.1493. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Singer M. S., Gottschling D. E. TLC1: template RNA component of Saccharomyces cerevisiae telomerase. Science. 1994 Oct 21;266(5184):404–409. doi: 10.1126/science.7545955. [DOI] [PubMed] [Google Scholar]
  28. Steiner B. R., Hidaka K., Futcher B. Association of the Est1 protein with telomerase activity in yeast. Proc Natl Acad Sci U S A. 1996 Apr 2;93(7):2817–2821. doi: 10.1073/pnas.93.7.2817. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Virta-Pearlman V., Morris D. K., Lundblad V. Est1 has the properties of a single-stranded telomere end-binding protein. Genes Dev. 1996 Dec 15;10(24):3094–3104. doi: 10.1101/gad.10.24.3094. [DOI] [PubMed] [Google Scholar]
  30. Wang H., Blackburn E. H. De novo telomere addition by Tetrahymena telomerase in vitro. EMBO J. 1997 Feb 17;16(4):866–879. doi: 10.1093/emboj/16.4.866. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Zahler A. M., Williamson J. R., Cech T. R., Prescott D. M. Inhibition of telomerase by G-quartet DNA structures. Nature. 1991 Apr 25;350(6320):718–720. doi: 10.1038/350718a0. [DOI] [PubMed] [Google Scholar]
  32. Zakian V. A. Telomeres: beginning to understand the end. Science. 1995 Dec 8;270(5242):1601–1607. doi: 10.1126/science.270.5242.1601. [DOI] [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES