Advertisement

Histochemistry and Cell Biology

, Volume 141, Issue 2, pp 137–152 | Cite as

Catalytically active telomerase holoenzyme is assembled in the dense fibrillar component of the nucleolus during S phase

  • Ji Hoon Lee
  • Yang Sin Lee
  • Sun Ah Jeong
  • Prabhat Khadka
  • Jürgen RothEmail author
  • In Kwon ChungEmail author
Original Paper

Abstract

The maintenance of human telomeres requires the ribonucleoprotein enzyme telomerase, which is composed of telomerase reverse transcriptase (TERT), telomerase RNA component, and several additional proteins for assembly and activity. Telomere elongation by telomerase in human cancer cells involves multiple steps including telomerase RNA biogenesis, holoenzyme assembly, intranuclear trafficking, and telomerase recruitment to telomeres. Although telomerase has been shown to accumulate in Cajal bodies for association with telomeric chromatin, it is unclear where and how the assembly and trafficking of catalytically active telomerase is regulated in the context of nuclear architecture. Here, we show that the catalytically active holoenzyme is initially assembled in the dense fibrillar component of the nucleolus during S phase. The telomerase RNP is retained in nucleoli through the interaction of hTERT with nucleolin, a major nucleolar phosphoprotein. Upon association with TCAB1 in S phase, the telomerase RNP is transported from nucleoli to Cajal bodies, suggesting that TCAB1 acts as an S-phase-specific holoenzyme component. Furthermore, depletion of TCAB1 caused an increase in the amount of telomerase RNP associated with nucleolin. These results suggest that the TCAB1-dependent trafficking of telomerase to Cajal bodies occurs in a step separate from the holoenzyme assembly in nucleoli. Thus, we propose that the dense fibrillar component is the provider of active telomerase RNP for supporting the continued proliferation of cancer and stem cells.

Keywords

Telomere Telomerase holoenzyme assembly Dense fibrillar component of nucleolus hTERT TCAB1 Cajal body 

Notes

Acknowledgments

This work was supported by World Class University Fund from the Korean Ministry of Education, Science, and Technology R31-2009-000-10086-0 to IKC and JR.

Supplementary material

418_2013_1166_MOESM1_ESM.pdf (1.4 mb)
Supplementary material 1 (PDF 1399 kb)

References

  1. Armanios M (2009) Syndromes of telomere shortening. Annu Rev Genomics Hum Genet 10:45–61PubMedCentralPubMedCrossRefGoogle Scholar
  2. Autexier C, Lue NF (2006) The structure and function of telomerase reverse transcriptase. Annu Rev Biochem 75:493–517PubMedCrossRefGoogle Scholar
  3. Batista LF, Pech MF, Zhong FL, Nguyen HN, Xie KT, Zaug AJ, Crary SM, Choi J, Sebastiano V, Cherry A, Giri N, Wernig M, Alter BP, Cech TR, Savage SA, Reijo Pera RA, Artandi SE (2011) Telomere shortening and loss of self-renewal in dyskeratosis congenita induced pluripotent stem cells. Nature 474:399–402PubMedCentralPubMedCrossRefGoogle Scholar
  4. Bianchi A, Shore D (2008) How telomerase reaches its end: mechanism of telomerase regulation by the telomeric complex. Mol Cell 31:153–165PubMedCrossRefGoogle Scholar
  5. Blackburn EH (2001) Switching and signaling at the telomere. Cell 106:661–673PubMedCrossRefGoogle Scholar
  6. Blasco MA, Lee HW, Hande MP, Samper E, Lansdorp PM, DePinho RA, Greider CW (1997) Telomere shortening and tumor formation by mouse cells lacking telomerase RNA. Cell 91:25–34PubMedCrossRefGoogle Scholar
  7. Bodnar AG, Ouellette M, Frolkis M, Holt SE, Chiu CP, Morin GB, Harley CB, Shay JW, Lichtsteiner S, Wright WE (1998) Extension of life-span by introduction of telomerase into normal human cells. Science 279:349–352PubMedCrossRefGoogle Scholar
  8. Bryan TM, Englezou A, Dalla-Pozza L, Dunham MA, Reddel RR (1997) Evidence for an alternative mechanism for maintaining telomere length in human tumors and tumor-derived cell lines. Nat Med 3:1271–1274PubMedCrossRefGoogle Scholar
  9. Chen JL, Blasco MA, Greider CW (2000) Secondary structure of vertebrate telomerase RNA. Cell 100:503–514PubMedCrossRefGoogle Scholar
  10. Collins K (2008) Physiological assembly and activity of human telomerase complexes. Mech Ageing Dev 129:91–98PubMedCentralPubMedCrossRefGoogle Scholar
  11. Cristofari G, Adolf E, Reichenbach P, Sikora K, Terns RM, Terns MP, Lingner J (2007) Human telomerase RNA accumulation in Cajal bodies facilitates telomerase recruitment to telomeres and telomere elongation. Mol Cell 27:882–889PubMedCrossRefGoogle Scholar
  12. Darzacq X, Kittur N, Roy S, Shav-Tal Y, Singer RH, Meier UT (2006) Stepwise RNP assembly at the site of H/ACA RNA transcription in human cells. J Cell Biol 173:207–218PubMedCrossRefGoogle Scholar
  13. de Lange T (2005) Shelterin: the protein complex that shapes and safeguards human telomeres. Genes Dev 19:2100–2110PubMedCrossRefGoogle Scholar
  14. Dunham MA, Neumann AA, Fasching CL, Reddel RR (2000) Telomere maintenance by recombination in human cells. Nat Genet 26:447–450PubMedCrossRefGoogle Scholar
  15. Egan ED, Collins K (2010) Specificity and stoichiometry of subunit interactions in the human telomerase holoenzyme assembled in vivo. Mol Cell Biol 30:2775–2786PubMedCentralPubMedCrossRefGoogle Scholar
  16. Egan ED, Collins K (2012) Biogenesis of telomerase ribonucleoproteins. RNA 18:1747–1759PubMedCrossRefGoogle Scholar
  17. Fu D, Collins K (2003) Distinct biogenesis pathways for human telomerase RNA and H/ACA small nucleolar RNAs. Mol Cell 11:1361–1372PubMedCrossRefGoogle Scholar
  18. Hahn WC, Counter CM, Lundberg AS, Beijersbergen RL, Brooks MW, Weinberg RA (1999) Creation of human tumor cells with defined genetic elements. Nature 400:464–468PubMedCrossRefGoogle Scholar
  19. Jády BE, Bertrand E, Kiss T (2004) Human telomerase RNA and box H/ACA scaRNAs share a common Cajal body-specific localization signal. J Cell Biol 164:647–652PubMedCrossRefGoogle Scholar
  20. Jády BE, Richard P, Bertrand E, Kiss T (2006) Cell cycle-dependent recruitment of telomerase RNA and Cajal bodies to human telomeres. Mol Biol Cell 17:944–954PubMedCentralPubMedCrossRefGoogle Scholar
  21. Khurts S, Masutomi K, Delgermaa L, Arai K, Oishi N, Mizuno H, Hayashi N, Hahn WC, Murakami S (2004) Nucleolin interacts with telomerase. J Biol Chem 279:51508–51515PubMedCrossRefGoogle Scholar
  22. Kim NW, Wu F (1997) Advances in quantification and characterization of telomerase activity by the telomeric repeat amplification protocol (TRAP). Nucleic Acids Res 25:2595–2597PubMedCentralPubMedCrossRefGoogle Scholar
  23. Kim NW, Piatyszek MA, Prowse KR, Harley CB, West MD, Ho PL, Coviello GM, Wright WE, Weinrich SL, Shay JW (1994) Specific association of human telomerase activity with immortal cells and cancer. Science 266:2011–2015PubMedCrossRefGoogle Scholar
  24. Kim JH, Kim JH, Lee GE, Lee JE, Chung IK (2003) Potent inhibition of human telomerase by nitrostyrene derivatives. Mol Pharmacol 63:1117–1124PubMedCrossRefGoogle Scholar
  25. Kiss T, Fayet-Lebaron E, Jády BE (2010) Box H/ACA small ribonucleoproteins. Mol Cell 37:597–606PubMedCrossRefGoogle Scholar
  26. Lai CK, Mitchell JR, Collins K (2001) RNA binding domain of telomerase reverse transcriptase. Mol Cell Biol 21:990–1000PubMedCentralPubMedCrossRefGoogle Scholar
  27. Lee GE, Yu EY, Cho CH, Lee J, Muller MT, Chung IK (2004) DNA-protein kinase catalytic subunit-interacting protein KIP binds telomerase by interacting with human telomerase reverse transcriptase. J Biol Chem 279:34750–34755PubMedCrossRefGoogle Scholar
  28. Lee JH, Khadka P, Baek SH, Chung IK (2010) CHIP promotes human telomerase reverse transcriptase degradation and negatively regulates telomerase activity. J Biol Chem 285:42033–42045PubMedCrossRefGoogle Scholar
  29. Lingner J, Cooper JP, Cech TR (1995) Telomerase and DNA end replication: no longer a lagging strand problem? Science 269:1533–1534PubMedCrossRefGoogle Scholar
  30. Liu D, O’Connor MS, Qin J, Songyang Z (2004) Telosome, a mammalian telomere-associated complex formed by multiple telomeric proteins. J Biol Chem 279:51338–51342PubMedCrossRefGoogle Scholar
  31. Marcand S, Brevet V, Mann C, Gilson E (2000) Cell cycle restriction of telomere elongation. Curr Biol 10:487–490PubMedCrossRefGoogle Scholar
  32. Mitchell JR, Collins K (2000) Human telomerase activation requires two independent interactions between telomerase RNA and telomerase reverse transcriptase in vivo and in vitro. Mol Cell 6:361–371PubMedCrossRefGoogle Scholar
  33. Mitchell JR, Cheng J, Collins K (1999) A box H/ACA small nucleolar RNA-like domain at the human telomerase RNA 39 end. Mol Cell Biol 19:567–576PubMedCentralPubMedGoogle Scholar
  34. Palm W, de Lange T (2008) How shelterin protects mammalian telomeres. Annu Rev Genet 42:301–334PubMedCrossRefGoogle Scholar
  35. Richard P, Kiss AM, Darzacq X, Kiss T (2006) Cotranscriptional recognition of human intronic box H/ACA snoRNAs occurs in a splicing-independent manner. Mol Cell Biol 26:2540–2549PubMedCentralPubMedCrossRefGoogle Scholar
  36. Robart AR, Collins K (2011) Human telomerase domain interactions capture DNA for TEN domain-dependent processive elongation. Mol Cell 42:308–318PubMedCentralPubMedCrossRefGoogle Scholar
  37. Roth J, Bendayan M, Orci L (1978) Ultrastructural localization of intracellular antigens by the use of protein A-gold complex. J Histochem Cytochem 26:1074–1081PubMedCrossRefGoogle Scholar
  38. Savage SA, Bertuch AA (2010) The genetics and clinical manifestations of telomere biology disorders. Genet Med 12:753–764PubMedCrossRefGoogle Scholar
  39. Sexton AN, Youmans DT, Collins K (2012) Specificity requirements for human telomere protein interaction with telomerase holoenzyme. J Biol Chem 287:34455–34464PubMedCrossRefGoogle Scholar
  40. Sfeir A, de Lange T (2012) Removal of shelterin reveals the telomere end-protection problem. Science 336:593–597PubMedCentralPubMedCrossRefGoogle Scholar
  41. Smogorzewska A, de Lange T (2004) Regulation of telomerase by telomeric proteins. Annu Rev Biochem 73:177–208PubMedCrossRefGoogle Scholar
  42. Stern JL, Zyner KG, Pickett HA, Cohen SB, Bryan TM (2012) Telomerase recruitment requires both TCAB1 and Cajal bodies independently. Mol Cell Biol 32:2384–2395PubMedCentralPubMedCrossRefGoogle Scholar
  43. Tomlinson RL, Ziegler TD, Supakorndej T, Terns RM, Terns MP (2006) Cell cycle-regulated trafficking of human telomerase to telomeres. Mol Biol Cell 17:955–965PubMedCentralPubMedCrossRefGoogle Scholar
  44. Tomlinson RL, Abreu EB, Ziegler T, Ly H, Counter CM, Terns RM, Terns MP (2008) Telomerase reverse transcriptase is required for the localization of telomerase RNA to Cajal bodies and telomeres in human cancer cells. Mol Biol Cell 19:3793–3800PubMedCentralPubMedCrossRefGoogle Scholar
  45. Trahan C, Dragon F (2009) Dyskeratosis congenita mutations in the H/ACA domain of human telomerase RNA affect its assembly into a pre-RNP. RNA 15:235–243PubMedCrossRefGoogle Scholar
  46. Tycowski KT, Shu MD, Kukoyi A, Steitz JA (2009) A conserved WD40 protein binds the Cajal body localization signal of scaRNP particles. Mol Cell 34:47–57PubMedCentralPubMedCrossRefGoogle Scholar
  47. Venteicher AS, Artandi SE (2009) TCAB1: driving telomerase to Cajal bodies. Cell Cycle 8:1329–1331PubMedCrossRefGoogle Scholar
  48. Venteicher AS, Meng Z, Mason PJ, Veenstra TD, Artandi SE (2008) Identification of ATPases pontin and reptin as telomerase components essential for holoenzyme assembly. Cell 132:945–957PubMedCentralPubMedCrossRefGoogle Scholar
  49. Venteicher AS, Abreu EB, Meng Z, McCann KE, Terns RM, Veenstra TD, Terns MP, Artandi SE (2009) A human telomerase holoenzyme protein required for Cajal body localization and telomere synthesis. Science 323:644–648PubMedCentralPubMedCrossRefGoogle Scholar
  50. Wang C, Meier UT (2004) Architecture and assembly of mammalian H/ACA small nucleolar and telomerase ribonucleoproteins. EMBO J 23:1857–1867PubMedCrossRefGoogle Scholar
  51. Zhong F, Savage SA, Shkreli M, Giri N, Jessop L, Myers T, Chen R, Alter BP, Artandi SE (2011) Disruption of telomerase trafficking by TCAB1 mutation causes dyskeratosis congenita. Genes Dev 25:11–16PubMedCrossRefGoogle Scholar
  52. Zhong FL, Batista LF, Freund A, Pech MF, Venteicher AS, Artandi SE (2012) TPP1 OB-fold domain controls telomere maintenance by recruiting telomerase to chromosome ends. Cell 150:481–494PubMedCentralPubMedCrossRefGoogle Scholar
  53. Zhu Y, Tomlinson RL, Lukowiak AA, Terns RM, Terns MP (2004) Telomerase RNA accumulates in Cajal bodies in human cancer cells. Mol Biol Cell 15:81–90PubMedCentralPubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2013

Authors and Affiliations

  1. 1.Department of Integrated Omics for Biomedical ScienceGraduate School of Yonsei UniversitySeoulKorea
  2. 2.Department of Systems Biology, College of Life Science and BiotechnologyYonsei UniversitySeoulKorea

Personalised recommendations