skip to main content
DOE PAGES title logo U.S. Department of Energy
Office of Scientific and Technical Information

Title: Structure of Tetrahymena telomerase reveals previously unknown subunits, functions, and interactions

Abstract

Telomerase helps maintain telomeres by processive synthesis of telomere repeat DNA at their 3'-ends, using an integral telomerase RNA (TER) and telomerase reverse transcriptase (TERT). In this paper, we report the cryo–electron microscopy structure of Tetrahymena telomerase at ~9 angstrom resolution. In addition to seven known holoenzyme proteins, we identify two additional proteins that form a complex (TEB) with single-stranded telomere DNA-binding protein Teb1, paralogous to heterotrimeric replication protein A (RPA). The p75-p45-p19 subcomplex is identified as another RPA-related complex, CST (CTC1-STN1-TEN1). This study reveals the paths of TER in the TERT-TER-p65 catalytic core and single-stranded DNA exit; extensive subunit interactions of the TERT essential N-terminal domain, p50, and TEB; and other subunit identities and structures, including p19 and p45C crystal structures. Finally, our findings provide structural and mechanistic insights into telomerase holoenzyme function.

Authors:
; ; ; ; ; ; ; ; ; ; ;
Publication Date:
Research Org.:
Univ. of California, Los Angeles, CA (United States)
Sponsoring Org.:
USDOE; National Inst. of Health (NIH) (United States); National Science Foundation (NSF); American Heart Association (United States)
Contributing Org.:
Univ. of California, Berkeley, CA (United States)
OSTI Identifier:
1355826
Alternate Identifier(s):
OSTI ID: 1226366
Grant/Contract Number:  
FC0302ER63421; AC02-06CH11357; FC03-02ER63421; GM048123; MCB1022379; GM071940; GM103479; R01GM054198; 14POST18870059; GM101874; GM007185; DGE-1106400; S10RR23057; S10OD018111; DBI-133813; P41 RR015301; P41 GM103403
Resource Type:
Published Article
Journal Name:
Science
Additional Journal Information:
Journal Name: Science Journal Volume: 350 Journal Issue: 6260; Journal ID: ISSN 0036-8075
Publisher:
American Association for the Advancement of Science (AAAS)
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; 60 APPLIED LIFE SCIENCES

Citation Formats

Jiang, J., Chan, H., Cash, D. D., Miracco, E. J., Ogorzalek Loo, R. R., Upton, H. E., Cascio, D., O'Brien Johnson, R., Collins, K., Loo, J. A., Zhou, Z. H., and Feigon, J. Structure of Tetrahymena telomerase reveals previously unknown subunits, functions, and interactions. United States: N. p., 2015. Web. doi:10.1126/science.aab4070.
Jiang, J., Chan, H., Cash, D. D., Miracco, E. J., Ogorzalek Loo, R. R., Upton, H. E., Cascio, D., O'Brien Johnson, R., Collins, K., Loo, J. A., Zhou, Z. H., & Feigon, J. Structure of Tetrahymena telomerase reveals previously unknown subunits, functions, and interactions. United States. doi:10.1126/science.aab4070.
Jiang, J., Chan, H., Cash, D. D., Miracco, E. J., Ogorzalek Loo, R. R., Upton, H. E., Cascio, D., O'Brien Johnson, R., Collins, K., Loo, J. A., Zhou, Z. H., and Feigon, J. Thu . "Structure of Tetrahymena telomerase reveals previously unknown subunits, functions, and interactions". United States. doi:10.1126/science.aab4070.
@article{osti_1355826,
title = {Structure of Tetrahymena telomerase reveals previously unknown subunits, functions, and interactions},
author = {Jiang, J. and Chan, H. and Cash, D. D. and Miracco, E. J. and Ogorzalek Loo, R. R. and Upton, H. E. and Cascio, D. and O'Brien Johnson, R. and Collins, K. and Loo, J. A. and Zhou, Z. H. and Feigon, J.},
abstractNote = {Telomerase helps maintain telomeres by processive synthesis of telomere repeat DNA at their 3'-ends, using an integral telomerase RNA (TER) and telomerase reverse transcriptase (TERT). In this paper, we report the cryo–electron microscopy structure of Tetrahymena telomerase at ~9 angstrom resolution. In addition to seven known holoenzyme proteins, we identify two additional proteins that form a complex (TEB) with single-stranded telomere DNA-binding protein Teb1, paralogous to heterotrimeric replication protein A (RPA). The p75-p45-p19 subcomplex is identified as another RPA-related complex, CST (CTC1-STN1-TEN1). This study reveals the paths of TER in the TERT-TER-p65 catalytic core and single-stranded DNA exit; extensive subunit interactions of the TERT essential N-terminal domain, p50, and TEB; and other subunit identities and structures, including p19 and p45C crystal structures. Finally, our findings provide structural and mechanistic insights into telomerase holoenzyme function.},
doi = {10.1126/science.aab4070},
journal = {Science},
number = 6260,
volume = 350,
place = {United States},
year = {2015},
month = {10}
}

Journal Article:
Free Publicly Available Full Text
Publisher's Version of Record
DOI: 10.1126/science.aab4070

Citation Metrics:
Cited by: 35 works
Citation information provided by
Web of Science

Save / Share:

Works referenced in this record:

The Pot1a-associated proteins Tpt1 and Pat1 coordinate telomere protection and length regulation in Tetrahymena
journal, November 2011

  • Linger, Benjamin R.; Morin, Gregg B.; Price, Carolyn M.
  • Molecular Biology of the Cell, Vol. 22, Issue 21
  • DOI: 10.1091/mbc.E11-06-0551

TPP1 is a homologue of ciliate TEBP-β and interacts with POT1 to recruit telomerase
journal, January 2007


Tetrahymena POT1a Regulates Telomere Length and Prevents Activation of a Cell Cycle Checkpoint
journal, December 2006

  • Jacob, N. K.; Lescasse, R.; Linger, B. R.
  • Molecular and Cellular Biology, Vol. 27, Issue 5
  • DOI: 10.1128/MCB.01975-06

Telomeres and telomerase in cancer
journal, November 2009


RNA-protein binding interface in the telomerase ribonucleoprotein
journal, November 2011

  • Bley, C. J.; Qi, X.; Rand, D. P.
  • Proceedings of the National Academy of Sciences, Vol. 108, Issue 51
  • DOI: 10.1073/pnas.1100270108

The human CST complex is a terminator of telomerase activity
journal, July 2012

  • Chen, Liuh-Yow; Redon, Sophie; Lingner, Joachim
  • Nature, Vol. 488, Issue 7412
  • DOI: 10.1038/nature11269

Telomerase and Telomere-Associated Proteins: Structural Insights into Mechanism and Evolution
journal, January 2012


A popular engagement at the ends
journal, January 2013

  • Lue, Neal F.; Yu, Eun Young; Lei, Ming
  • Nature Structural & Molecular Biology, Vol. 20, Issue 1
  • DOI: 10.1038/nsmb.2483

Accurate determination of local defocus and specimen tilt in electron microscopy
journal, June 2003


DoG Picker and TiltPicker: Software tools to facilitate particle selection in single particle electron microscopy
journal, May 2009

  • Voss, N. R.; Yoshioka, C. K.; Radermacher, M.
  • Journal of Structural Biology, Vol. 166, Issue 2
  • DOI: 10.1016/j.jsb.2009.01.004

Electron counting and beam-induced motion correction enable near-atomic-resolution single-particle cryo-EM
journal, May 2013

  • Li, Xueming; Mooney, Paul; Zheng, Shawn
  • Nature Methods, Vol. 10, Issue 6
  • DOI: 10.1038/nmeth.2472

Multi-resolution contour-based fitting of macromolecular structures
journal, March 2002


A critical stem-loop structure in the CR4-CR5 domain of mammalian telomerase RNA
journal, January 2002


Automated molecular microscopy: The new Leginon system
journal, July 2005

  • Suloway, Christian; Pulokas, James; Fellmann, Denis
  • Journal of Structural Biology, Vol. 151, Issue 1
  • DOI: 10.1016/j.jsb.2005.03.010

Structure of the Human Telomerase RNA Pseudoknot Reveals Conserved Tertiary Interactions Essential for Function
journal, March 2005


LNA (Locked Nucleic Acid):  High-Affinity Targeting of Complementary RNA and DNA
journal, October 2004

  • Vester, Birte; Wengel, Jesper
  • Biochemistry, Vol. 43, Issue 42
  • DOI: 10.1021/bi0485732

Towards automated crystallographic structure refinement with phenix.refine
journal, March 2012

  • Afonine, Pavel V.; Grosse-Kunstleve, Ralf W.; Echols, Nathaniel
  • Acta Crystallographica Section D Biological Crystallography, Vol. 68, Issue 4
  • DOI: 10.1107/S0907444912001308

Telomeres and telomerase: the path from maize, Tetrahymena and yeast to human cancer and aging
journal, October 2006

  • Blackburn, Elizabeth H.; Greider, Carol W.; Szostak, Jack W.
  • Nature Medicine, Vol. 12, Issue 10
  • DOI: 10.1038/nm1006-1133

Functional importance of telomerase pseudoknot revealed by single-molecule analysis
journal, May 2011

  • Mihalusova, M.; Wu, J. Y.; Zhuang, X.
  • Proceedings of the National Academy of Sciences, Vol. 108, Issue 51
  • DOI: 10.1073/pnas.1017686108

Structure of the Human Telomeric Stn1-Ten1 Capping Complex
journal, June 2013


Pyrimidine motif triple helix in the Kluyveromyces lactis telomerase RNA pseudoknot is essential for function in vivo
journal, June 2013

  • Cash, D. D.; Cohen-Zontag, O.; Kim, N. -K.
  • Proceedings of the National Academy of Sciences, Vol. 110, Issue 27
  • DOI: 10.1073/pnas.1309590110

Specificity Requirements for Human Telomere Protein Interaction with Telomerase Holoenzyme
journal, August 2012

  • Sexton, Alec N.; Youmans, Daniel T.; Collins, Kathleen
  • Journal of Biological Chemistry, Vol. 287, Issue 41
  • DOI: 10.1074/jbc.M112.394767

Gapped BLAST and PSI-BLAST: a new generation of protein database search programs
journal, September 1997

  • Altschul, Stephen F.; Madden, Thomas L.; Schäffer, Alejandro A.
  • Nucleic Acids Research, Vol. 25, Issue 17, p. 3389-3402
  • DOI: 10.1093/nar/25.17.3389

Enhanced FASP (eFASP) to Increase Proteome Coverage and Sample Recovery for Quantitative Proteomic Experiments
journal, March 2014

  • Erde, Jonathan; Loo, Rachel R. Ogorzalek; Loo, Joseph A.
  • Journal of Proteome Research, Vol. 13, Issue 4
  • DOI: 10.1021/pr4010019

Solution Structure and Dynamics of the Wild-type Pseudoknot of Human Telomerase RNA
journal, December 2008


Biogenesis of telomerase ribonucleoproteins
journal, August 2012


Multiple Mechanisms for Elongation Processivity within the Reconstituted Tetrahymena Telomerase Holoenzyme
journal, April 2010


It all comes together at the ends: Telomerase structure, function, and biogenesis
journal, February 2012

  • Podlevsky, Joshua D.; Chen, Julian J. -L.
  • Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis, Vol. 730, Issue 1-2
  • DOI: 10.1016/j.mrfmmm.2011.11.002

CST for the grand finale of telomere replication
journal, July 2013


Human telomerase specialization for repeat synthesis by unique handling of primer-template duplex
journal, March 2014


The telomere syndromes
journal, September 2012

  • Armanios, Mary; Blackburn, Elizabeth H.
  • Nature Reviews Genetics, Vol. 13, Issue 10
  • DOI: 10.1038/nrg3246

Stimulation of Yeast Telomerase Activity by the Ever Shorter Telomere 3 (Est3) Subunit Is Dependent on Direct Interaction with the Catalytic Protein Est2
journal, June 2011

  • Talley, Jennell M.; DeZwaan, Diane C.; Maness, Leslie D.
  • Journal of Biological Chemistry, Vol. 286, Issue 30
  • DOI: 10.1074/jbc.M111.228635

A Novel RNA Binding Domain in Tetrahymena Telomerase p65 Initiates Hierarchical Assembly of Telomerase Holoenzyme
journal, February 2006


Integration, scaling, space-group assignment and post-refinement
journal, January 2010

  • Kabsch, Wolfgang
  • Acta Crystallographica Section D Biological Crystallography, Vol. 66, Issue 2, p. 133-144
  • DOI: 10.1107/S0907444909047374

UCSF Chimera?A visualization system for exploratory research and analysis
journal, January 2004

  • Pettersen, Eric F.; Goddard, Thomas D.; Huang, Conrad C.
  • Journal of Computational Chemistry, Vol. 25, Issue 13
  • DOI: 10.1002/jcc.20084

Bsoft: Image processing and molecular modeling for electron microscopy
journal, January 2007


Genetic and molecular identification of three human TPP1 functions in telomerase action: recruitment, activation, and homeostasis set point regulation
journal, August 2014

  • Sexton, Alec N.; Regalado, Samuel G.; Lai, Christine S.
  • Genes & Development, Vol. 28, Issue 17
  • DOI: 10.1101/gad.246819.114

Quantifying the local resolution of cryo-EM density maps
journal, November 2013

  • Kucukelbir, Alp; Sigworth, Fred J.; Tagare, Hemant D.
  • Nature Methods, Vol. 11, Issue 1
  • DOI: 10.1038/nmeth.2727

Holoenzyme proteins required for the physiological assembly and activity of telomerase
journal, May 2004


Structural basis for protein-RNA recognition in telomerase
journal, May 2014

  • Huang, Jing; Brown, Andrew F.; Wu, Jian
  • Nature Structural & Molecular Biology, Vol. 21, Issue 6
  • DOI: 10.1038/nsmb.2819

Structure and conformational change of a replication protein A heterotrimer bound to ssDNA
journal, October 2012


Stepwise protein-mediated RNA folding directs assembly of telomerase ribonucleoprotein
journal, February 2007

  • Stone, Michael D.; Mihalusova, Mariana; O’Connor, Catherine M.
  • Nature, Vol. 446, Issue 7134
  • DOI: 10.1038/nature05600

Structure of the RNA-Binding Domain of Telomerase: Implications for RNA Recognition and Binding
journal, November 2007


Crystal structure of the essential N-terminal domain of telomerase reverse transcriptase
journal, February 2006

  • Jacobs, Steven A.; Podell, Elaine R.; Cech, Thomas R.
  • Nature Structural & Molecular Biology, Vol. 13, Issue 3
  • DOI: 10.1038/nsmb1054

Everything You Ever Wanted to Know About Saccharomyces cerevisiae Telomeres: Beginning to End
journal, August 2012


A synergistic approach to protein crystallization: Combination of a fixed-arm carrier with surface entropy reduction
journal, January 2010

  • Moon, Andrea F.; Mueller, Geoffrey A.; Zhong, Xuejun
  • Protein Science
  • DOI: 10.1002/pro.368

The TEL patch of telomere protein TPP1 mediates telomerase recruitment and processivity
journal, October 2012

  • Nandakumar, Jayakrishnan; Bell, Caitlin F.; Weidenfeld, Ina
  • Nature, Vol. 492, Issue 7428
  • DOI: 10.1038/nature11648

The Jpred 3 secondary structure prediction server
journal, May 2008

  • Cole, C.; Barber, J. D.; Barton, G. J.
  • Nucleic Acids Research, Vol. 36, Issue Web Server
  • DOI: 10.1093/nar/gkn238

Direct Single-Stranded DNA Binding by Teb1 Mediates the Recruitment of Tetrahymena thermophila Telomerase to Telomeres
journal, September 2014

  • Upton, H. E.; Hong, K.; Collins, K.
  • Molecular and Cellular Biology, Vol. 34, Issue 22
  • DOI: 10.1128/MCB.01030-14

Structure of Est3 reveals a bimodal surface with differential roles in telomere replication
journal, December 2013

  • Rao, T.; Lubin, J. W.; Armstrong, G. S.
  • Proceedings of the National Academy of Sciences, Vol. 111, Issue 1
  • DOI: 10.1073/pnas.1316453111

Coot model-building tools for molecular graphics
journal, November 2004

  • Emsley, Paul; Cowtan, Kevin
  • Acta Crystallographica Section D Biological Crystallography, Vol. 60, Issue 12, p. 2126-2132
  • DOI: 10.1107/S0907444904019158

Structural basis for telomerase catalytic subunit TERT binding to RNA template and telomeric DNA
journal, March 2010

  • Mitchell, Meghan; Gillis, Andrew; Futahashi, Mizuko
  • Nature Structural & Molecular Biology, Vol. 17, Issue 4
  • DOI: 10.1038/nsmb.1777

Maintaining the end: Roles of telomere proteins in end-protection, telomere replication and length regulation
journal, February 2012

  • Stewart, Jason A.; Chaiken, Mary F.; Wang, Feng
  • Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis, Vol. 730, Issue 1-2
  • DOI: 10.1016/j.mrfmmm.2011.08.011

Tetrahymena Telomerase Holoenzyme Assembly, Activation, and Inhibition by Domains of the p50 Central Hub
journal, August 2013

  • Hong, K.; Upton, H.; Miracco, E. J.
  • Molecular and Cellular Biology, Vol. 33, Issue 19
  • DOI: 10.1128/MCB.00792-13

RELION: Implementation of a Bayesian approach to cryo-EM structure determination
journal, December 2012


Human CST Has Independent Functions during Telomere Duplex Replication and C-Strand Fill-In
journal, November 2012


The architecture of Tetrahymena telomerase holoenzyme
journal, April 2013

  • Jiang, Jiansen; Miracco, Edward J.; Hong, Kyungah
  • Nature, Vol. 496, Issue 7444
  • DOI: 10.1038/nature12062

Finding the end: recruitment of telomerase to telomeres
journal, January 2013

  • Nandakumar, Jayakrishnan; Cech, Thomas R.
  • Nature Reviews Molecular Cell Biology, Vol. 14, Issue 2
  • DOI: 10.1038/nrm3505

Structural Basis for Telomerase RNA Recognition and RNP Assembly by the Holoenzyme La Family Protein p65
journal, July 2012


The POT1–TPP1 telomere complex is a telomerase processivity factor
journal, January 2007

  • Wang, Feng; Podell, Elaine R.; Zaug, Arthur J.
  • Nature, Vol. 445, Issue 7127
  • DOI: 10.1038/nature05454

A novel motif in telomerase reverse transcriptase regulates telomere repeat addition rate and processivity
journal, December 2009

  • Xie, Mingyi; Podlevsky, Joshua D.; Qi, Xiaodong
  • Nucleic Acids Research, Vol. 38, Issue 6
  • DOI: 10.1093/nar/gkp1198

TPP1 OB-Fold Domain Controls Telomere Maintenance by Recruiting Telomerase to Chromosome Ends
journal, August 2012


RNA/DNA hybrid binding affinity determines telomerase template-translocation efficiency: Telomerase template translocation mechanism
journal, October 2011


Telomeric 3′ Overhangs Derive from Resection by Exo1 and Apollo and Fill-In by POT1b-Associated CST
journal, July 2012


Transcriptome Analysis of the Model Protozoan, Tetrahymena thermophila, Using Deep RNA Sequencing
journal, February 2012


Mutation in TERT separates processivity from anchor-site function
journal, July 2008

  • Zaug, Arthur J.; Podell, Elaine R.; Cech, Thomas R.
  • Nature Structural & Molecular Biology, Vol. 15, Issue 8
  • DOI: 10.1038/nsmb.1462

Structure of the Tetrahymena thermophila telomerase RNA helix II template boundary element
journal, February 2006


Telomerase at the intersection of cancer and aging
journal, September 2013


Telomerase: An RNP Enzyme Synthesizes DNA
journal, July 2010


Structural basis for Tetrahymena telomerase processivity factor Teb1 binding to single-stranded telomeric-repeat DNA
journal, December 2011

  • Zeng, Z.; Min, B.; Huang, J.
  • Proceedings of the National Academy of Sciences, Vol. 108, Issue 51
  • DOI: 10.1073/pnas.1113624108

Structural Analysis Reveals Conformational Plasticity in the Recognition of RNA 3′ Ends by the Human La Protein
journal, June 2008


Triple-helix structure in telomerase RNA contributes to catalysis
journal, May 2008

  • Qiao, Feng; Cech, Thomas R.
  • Nature Structural & Molecular Biology, Vol. 15, Issue 6
  • DOI: 10.1038/nsmb.1420

Structure of the Tribolium castaneum telomerase catalytic subunit TERT
journal, August 2008

  • Gillis, Andrew J.; Schuller, Anthony P.; Skordalakes, Emmanuel
  • Nature, Vol. 455, Issue 7213
  • DOI: 10.1038/nature07283

Structure and function of telomerase RNA
journal, June 2006


Evolution of CST function in telomere maintenance
journal, August 2010

  • Price, Carolyn; Boltz, Kara A.; Chaiken, Mary F.
  • Cell Cycle, Vol. 9, Issue 16
  • DOI: 10.4161/cc.9.16.12547

Identification of human TERT elements necessary for telomerase recruitment to telomeres
journal, October 2014


Stn1-Ten1 is an Rpa2-Rpa3-like complex at telomeres
journal, December 2009


Regulated assembly and disassembly of the yeast telomerase quaternary complex
journal, September 2014


Roles of Telomerase Reverse Transcriptase N-terminal Domain in Assembly and Activity of Tetrahymena Telomerase Holoenzyme
journal, February 2012

  • Eckert, Barbara; Collins, Kathleen
  • Journal of Biological Chemistry, Vol. 287, Issue 16
  • DOI: 10.1074/jbc.M112.339853

Phaser crystallographic software
journal, July 2007

  • McCoy, Airlie J.; Grosse-Kunstleve, Ralf W.; Adams, Paul D.
  • Journal of Applied Crystallography, Vol. 40, Issue 4
  • DOI: 10.1107/S0021889807021206

EMAN: Semiautomated Software for High-Resolution Single-Particle Reconstructions
journal, December 1999

  • Ludtke, Steven J.; Baldwin, Philip R.; Chiu, Wah
  • Journal of Structural Biology, Vol. 128, Issue 1
  • DOI: 10.1006/jsbi.1999.4174

    Works referencing / citing this record:

    De novo computational RNA modeling into cryo-EM maps of large ribonucleoprotein complexes
    journal, October 2018


    Functional Proteomics of Nuclear Proteins in Tetrahymena thermophila: A Review
    journal, May 2019