Article | Published:

Spliceosomal cleavage generates the 3′ end of telomerase RNA

Nature volume 456, pages 910914 (18 December 2008) | Download Citation

Abstract

Telomeres cap the ends of chromosomes and provide a means to complete replication. The DNA portion of telomeres is synthesized by the enzyme telomerase using part of an RNA subunit as a template for reverse transcription. How the mature 3′ end of telomerase RNA is generated has so far remained elusive. Here we show that in Schizosaccharomyces pombe telomerase RNA transcripts must be processed to generate functional telomerase. Characterization of the maturation pathway uncovered an unexpected role for the spliceosome, which normally catalyses splicing of pre-messenger RNA. The first spliceosomal cleavage reaction generates the mature 3′ end of telomerase RNA (TER1, the functional RNA encoded by the ter1+ gene), releasing the active form of the RNA without exon ligation. Blocking the first step or permitting completion of splicing generates inactive forms of TER1 and causes progressive telomere shortening. We establish that 3′ end processing of TER1 is critical for telomerase function and describe a previously unknown mechanism for RNA maturation that uses the ability of the spliceosome to mediate site-specific cleavage.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

References

  1. 1.

    & Telomere length homeostasis. Chromosoma 115, 413–425 (2006)

  2. 2.

    & Telomeres: cancer to human aging. Annu. Rev. Cell Dev. Biol. 22, 531–557 (2006)

  3. 3.

    , & A telomerase component is defective in the human disease dyskeratosis congenita. Nature 402, 551–555 (1999)

  4. 4.

    et al. The RNA component of telomerase is mutated in autosomal dominant dyskeratosis congenita. Nature 413, 432–435 (2001)

  5. 5.

    , , & Association between aplastic anaemia and mutations in telomerase RNA. Lancet 359, 2168–2170 (2002)

  6. 6.

    et al. Telomerase mutations in families with idiopathic pulmonary fibrosis. N. Engl. J. Med. 356, 1317–1326 (2007)

  7. 7.

    et al. Mutations in TERT, the gene for telomerase reverse transcriptase, in aplastic anemia. N. Engl. J. Med. 352, 1413–1424 (2005)

  8. 8.

    , & A telomerase component is defective in the human disease dyskeratosis congenita. Nature 402, 551–555 (1999)

  9. 9.

    et al. Protein composition of catalytically active human telomerase from immortal cells. Science 315, 1850–1853 (2007)

  10. 10.

    et al. Reverse transcriptase motifs in the catalytic subunit of telomerase. Science 276, 561–567 (1997)

  11. 11.

    , & Structure of the Tribolium castaneum telomerase catalytic subunit TERT. Nature 455, 633–637 (2008)

  12. 12.

    , , & TER1, the RNA subunit of fission yeast telomerase. Nature Struct. Mol. Biol. 15, 26–33 (2008)

  13. 13.

    & Identification and characterization of the Schizosaccharomyces pombe TER1 telomerase RNA. Nature Struct. Mol. Biol. 15, 34–42 (2008)

  14. 14.

    & Pot1, the putative telomere end-binding protein in fission yeast and humans. Science 292, 1171–1175 (2001)

  15. 15.

    & Telomere binding of the Rap1 protein is required for meiosis in fission yeast. Curr. Biol. 11, 1618–1623 (2001)

  16. 16.

    , , & Regulation of telomere length and function by a Myb-domain protein in fission yeast. Nature 385, 744–747 (1997)

  17. 17.

    & spRap1 and spRif1, recruited to telomeres by Taz1, are essential for telomere function in fission yeast. Curr. Biol. 11, 1624–1630 (2001)

  18. 18.

    , , & Fission yeast Pot1–Tpp1 protects telomeres and regulates telomere length. Science 320, 1341–1344 (2008)

  19. 19.

    , & Polyadenylation of telomerase RNA in budding yeast. RNA 3, 1337–1351 (1997)

  20. 20.

    & Distinct biogenesis pathways for human telomerase RNA and H/ACA small nucleolar RNAs. Mol. Cell 11, 1361–1372 (2003)

  21. 21.

    & Fission yeast gene structure and recognition. Nucleic Acids Res. 22, 1750–1759 (1994)

  22. 22.

    , , , & Saccharomyces cerevisiae telomerase is an Sm small nuclear ribonucleoprotein particle. Nature 401, 177–180 (1999)

  23. 23.

    , & U1 snRNA:pre-mRNA base pairing interaction is required early in yeast spliceosome assembly but does not uniquely define the 5′ cleavage site. EMBO J. 7, 2533–2538 (1988)

  24. 24.

    & 5′ splice site selection in yeast: genetic alterations in base-pairing with U1 reveal additional requirements. Genes Dev. 2, 1258–1267 (1988)

  25. 25.

    & A compensatory base change in U1 snRNA suppresses a 5′ splice site mutation. Cell 46, 827–835 (1986)

  26. 26.

    , , , & Mutational analysis of U1 function in Schizosaccharomyces pombe: pre-mRNAs differ in the extent and nature of their requirements for this snRNA in vivo. RNA 2, 404–418 (1996)

  27. 27.

    & Mutational analysis of the interactions between U1 small nuclear RNA and pre-mRNA of yeast. Gene 82, 145–151 (1989)

  28. 28.

    et al. Lariat structures are in vivo intermediates in yeast pre-mRNA splicing. Cell 39, 611–621 (1984)

  29. 29.

    & Both the polypyrimidine tract and the 3′ splice site function prior to the first step of splicing in fission yeast. Nucleic Acids Res. 25, 4658–4665 (1997)

  30. 30.

    , , & Distinct requirements for Pot1 in limiting telomere length and maintaining chromosome stability. Mol. Cell. Biol. 25, 5567–5578 (2005)

  31. 31.

    , , , & A flexible template boundary element in the RNA subunit of fission yeast telomerase. J. Biol. Chem. 283, 24224–24233 (2008)

  32. 32.

    , & in The RNA World (eds Gesteland, R. F., Cech, T. R. & Atkins, J. F.) 525–560 (Cold Spring Harbor Laboratory Press, 1999)

  33. 33.

    Comparison of Schizosaccharomyces pombe expression systems. Nucleic Acids Res. 21, 2955–2956 (1993)

  34. 34.

    & Genomic sequencing. Proc. Natl Acad. Sci. USA 81, 1991–1995 (1984)

  35. 35.

    , , , & Identification and functional analysis of 20 Box H/ACA small nucleolar RNAs (snoRNAs) from Schizosaccharomyces pombe. J. Biol. Chem. 280, 16446–16455 (2005)

Download references

Acknowledgements

The authors thank H. Yang and the other members of the Baumann laboratory for help and discussions, Y. Tzfati and L. Tomaska for sharing results before publication, and A. Berglund, M. Blanchette, R. Conaway and T. Cech for discussions and comments on the manuscript. We also thank the Molecular Biology Core Facility for site-directed mutagenesis and sequencing, M. Gogol and R. Voelker for computational analysis, and D. Baumann and R. Helston for proofreading of the manuscript. This work was funded by the Stowers Institute for Medical Research and a Pew Scholars in the Biomedical Sciences Award to P.B.

Author Contributions P.B. made the initial observations, oversaw the project and designed the experiments. J.T.B. and P.B. developed protocols for RNA isolation, northern blotting and primer extension analysis. J.A.B. contributed plasmids and strains and performed telomere length analysis and RT–PCR assays. J.T.B. conducted northern blotting and primer extension analysis. W.T. characterized the TER1-Sm1 mutant. All authors contributed to data analysis, and P.B. wrote the manuscript.

Author information

Author notes

    • Jessica A. Box
    •  & Jeremy T. Bunch

    These authors contributed equally to this work.

Affiliations

  1. Stowers Institute for Medical Research, Kansas City, Missouri 64110, USA

    • Jessica A. Box
    • , Jeremy T. Bunch
    • , Wen Tang
    •  & Peter Baumann
  2. Department of Molecular and Integrative Physiology, University of Kansas Medical Center, Kansas City, Kansas 66160, USA

    • Wen Tang
    •  & Peter Baumann

Authors

  1. Search for Jessica A. Box in:

  2. Search for Jeremy T. Bunch in:

  3. Search for Wen Tang in:

  4. Search for Peter Baumann in:

Corresponding author

Correspondence to Peter Baumann.

Supplementary information

PDF files

  1. 1.

    Supplementary Information

    This file contais Supplementary Figures 1-6 illustrating the 5′ splice site consensus in S. pombe, the effects of mutations in TER1 on splicing, the mapping of the branch point by primer extension and TER1 processing at various levels of expression. A detailed schematic of the heterologous intron and mutations therein is also provided.

About this article

Publication history

Received

Accepted

Published

DOI

https://doi.org/10.1038/nature07584

Further reading

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.