Human telomerase RNA and box H/ACA scaRNAs share a common Cajal body–specific localization signal

Telomerase is a ribonucleoprotein reverse transcriptase that uses its RNA component as a template for synthesis of telomeric DNA repeats at the ends of linear eukaryotic chromosomes. Here, fluorescence in situ hybridization demonstrates that in HeLa cancer cells, human telomerase RNA (hTR) accumulates in the nucleoplasmic Cajal bodies (CBs). Localization of transiently expressed hTR to CBs is supported by a short sequence motif (411-UGAG-414) that is located in the 3′-terminal box H/ACA RNA-like domain of hTR and that is structurally and functionally indistinguishable from the CB-specific localization signal of box H/ACA small CB-specific RNAs. In synchronized HeLa cells, hTR shows the most efficient accumulation in CBs during S phase, when telomeres are most likely synthesized. CBs may function in post-transcriptional maturation (e.g., cap hypermethylation of hTR), but they may also play a role in the assembly and/or function of telomerase holoenzyme.

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Comprehensive Structure-Function Analysis of the Core Domain of Human Telomerase RNA

Molecular and Cellular Biology ◽

10.1128/mcb.23.19.6849-6856.2003 ◽

2003 ◽

Vol 23 (19) ◽

pp. 6849-6856 ◽

Cited By ~ 62

Author(s):

Hinh Ly ◽

Elizabeth H. Blackburn ◽

Tristram G. Parslow

Keyword(s):

Mutational Analysis ◽

Function Analysis ◽

Telomerase Activity ◽

Telomerase Rna ◽

Dna Repeats ◽

Telomeric Dna ◽

Core Domain ◽

The Core ◽

Optimal Function ◽

Human Telomerase

ABSTRACT Telomerase is a cellular reverse transcriptase that uses part of its integral RNA (called TER) as the template to synthesize telomeric DNA repeats. Vertebrate TERs are thought to share a conserved, highly structured core domain that includes the templating sequence and a pseudoknot, but not all features of the predicted core structure have been verified directly or shown to affect telomerase enzymatic activity. Here, we report a systematic mutational analysis of the core domain (residues 1 to 210) of human telomerase RNA (hTER). Our data confirm that optimal hTER activity requires the integrity of four short helices (P2a.1, P2a, P2b, and P3) which create the proposed pseudoknot and that features of both the primary sequence and secondary structure in P2b and P3 contribute to optimal function. At least part of the long-range P1 pairing is also required, despite the lack of a known P1 counterpart in rodent TERs. Among the predicted single-stranded regions, we found that J2b/3, portions of J2a/3, and residues in and around the template make sequence-specific contributions to telomerase function. Additionally, we provide evidence that naturally occurring hTER sequence polymorphisms found in some patients with aplastic anemia can inhibit telomerase activity by disrupting critical structures within the hTER core domain.

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Telomerase RNA Accumulates in Cajal Bodies in Human Cancer Cells

Molecular Biology of the Cell ◽

10.1091/mbc.e03-07-0525 ◽

2004 ◽

Vol 15 (1) ◽

pp. 81-90 ◽

Cited By ~ 128

Author(s):

Yusheng Zhu ◽

Rebecca L. Tomlinson ◽

Andrew A. Lukowiak ◽

Rebecca M. Terns ◽

Michael P. Terns

Keyword(s):

Subcellular Localization ◽

Cancer Cells ◽

Human Cancer ◽

Cajal Bodies ◽

Dna Repeats ◽

Primary Cells ◽

Telomeric Dna ◽

Human Cancer Cells ◽

Human Telomerase

Telomerase synthesizes telomeric DNA repeats at the ends of eukaryotic chromosomes. The RNA component of the enzyme (hTR) provides the template for telomere synthesis, which is catalyzed by telomerase reverse transcriptase (hTERT). Little is known regarding the subcellular localization of hTR and hTERT and the pathway by which telomerase is assembled. Here we report the first glimpse of the detailed subcellular localization of endogenous hTR in human cells, which we obtained by fluorescence in situ hybridization (FISH). Our studies have revealed a distinctive hTR localization pattern in cancer cells. We have found that hTR accumulates within intranuclear foci called Cajal bodies in all typical tumor-derived cell lines examined (in which telomerase is active), but not in primary or ALT cells (where little or no hTERT is present). Accumulation of hTR in the Cajal bodies of primary cells is induced when hTERT is ectopically expressed. Moreover, we report that hTERT is also found in Cajal bodies. Our data suggest that Cajal bodies are involved in the assembly and/or function of human telomerase.

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Cell Cycle-dependent Recruitment of Telomerase RNA and Cajal Bodies to Human Telomeres

Molecular Biology of the Cell ◽

10.1091/mbc.e05-09-0904 ◽

2006 ◽

Vol 17 (2) ◽

pp. 944-954 ◽

Cited By ~ 124

Author(s):

Beáta E. Jády ◽

Patricia Richard ◽

Edouard Bertrand ◽

Tamás Kiss

Keyword(s):

Time Lapse ◽

S Phase ◽

Telomere Maintenance ◽

Cajal Bodies ◽

Telomeric Sequence ◽

Marker Proteins ◽

Oligonucleotide Hybridization ◽

Cycle Dependent ◽

Human Telomerase

Telomerase is a ribonucleoprotein enzyme that counteracts replicative telomere erosion by adding telomeric sequence repeats onto chromosome ends. Despite its well-established role in telomere synthesis, telomerase has not yet been detected at telomeres. The RNA component of human telomerase (hTR) resides in the nucleoplasmic Cajal bodies (CBs) of interphase cancer cells. Here, in situ hybridization demonstrates that in human HeLa and Hep2 S phase cells, besides accumulating in CBs, hTR specifically concentrates at a few telomeres that also accumulate the TRF1 and TRF2 telomere marker proteins. Surprisingly, telomeres accumulating hTR exhibit a great accessibility for in situ oligonucleotide hybridization without chromatin denaturation, suggesting that they represent a structurally distinct, minor subset of HeLa telomeres. Moreover, we demonstrate that more than 25% of telomeres accumulating hTR colocalize with CBs. Time-lapse fluorescence microscopy demonstrates that CBs moving in the nucleoplasm of S phase cells transiently associate for 10-40 min with telomeres. Our data raise the intriguing possibility that CBs may deliver hTR to telomeres and/or may function in other aspects of telomere maintenance.

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Faculty Opinions recommendation of A common sequence motif determines the Cajal body-specific localization of box H/ACA scaRNAs.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1016274.200323 ◽

2003 ◽

Author(s):

Gabriele Varani

Keyword(s):

Cajal Body ◽

Sequence Motif ◽

Specific Localization ◽

Common Sequence

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Sticky-flares for in situ monitoring of human telomerase RNA in living cells

Nanoscale ◽

10.1039/c8nr01260a ◽

2018 ◽

Vol 10 (19) ◽

pp. 9386-9392 ◽

Cited By ~ 5

Author(s):

Qilong Wu ◽

Zhengjie Liu ◽

Lei Su ◽

Guangmei Han ◽

Renyong Liu ◽

...

Keyword(s):

Living Cells ◽

In Situ Monitoring ◽

Telomerase Rna ◽

Human Telomerase

A novel DNA-conjugated AuNP probe termed sticky-flares has been designed for the in situ detection of intracellular human telomerase RNA.

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Characterization of Physical and Functional Anchor Site Interactions in Human Telomerase

Molecular and Cellular Biology ◽

10.1128/mcb.02368-06 ◽

2007 ◽

Vol 27 (8) ◽

pp. 3226-3240 ◽

Cited By ~ 48

Author(s):

Haley D. M. Wyatt ◽

Deirdre A. Lobb ◽

Tara L. Beattie

Keyword(s):

Critical Role ◽

Telomerase Rna ◽

Dna Interactions ◽

Telomeric Dna ◽

Protein Subunit ◽

Protein Dna Interactions ◽

N Terminus ◽

Human Telomerase ◽

Linear Chromosomes

ABSTRACT Telomerase is a ribonucleoprotein reverse transcriptase (RT) that processively synthesizes telomeric repeats onto the ends of linear chromosomes to maintain genomic stability. It has been proposed that the N terminus of the telomerase protein subunit, telomerase RT (TERT), contains an anchor site that forms stable interactions with DNA to prevent enzyme-DNA dissociation during translocation and to promote realignment events that accompany each round of telomere synthesis. However, it is not known whether human TERT (hTERT) can directly interact with DNA in the absence of the telomerase RNA subunit. Here we use a novel primer binding assay to establish that hTERT forms stable and specific contacts with telomeric DNA in the absence of the human telomerase RNA component (hTR). We show that hTERT-mediated primer binding can be functionally uncoupled from telomerase-mediated primer extension. Our results demonstrate that the first 350 amino acids of hTERT have a critical role in regulating the strength and specificity of protein-DNA interactions, providing additional evidence that the TERT N terminus contains an anchor site. Furthermore, we establish that the RT domain of hTERT mediates important protein-DNA interactions. Collectively, these data suggest that hTERT contains distinct anchor regions that cooperate to help regulate telomerase-mediated DNA recognition and elongation.

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Structural and Functional Characterization of Human Telomerase RNA Processing and Cajal Body Localization Signals

Molecular Cell ◽

10.1016/j.molcel.2007.07.017 ◽

2007 ◽

Vol 27 (6) ◽

pp. 869-881 ◽

Cited By ~ 61

Author(s):

Carla A. Theimer ◽

Beáta E. Jády ◽

Nicholas Chim ◽

Patricia Richard ◽

Katherine E. Breece ◽

...

Keyword(s):

Rna Processing ◽

Functional Characterization ◽

Cajal Body ◽

Telomerase Rna ◽

Human Telomerase

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A single telomerase RNA is sufficient for the synthesis of variable telomeric DNA repeats in ciliates of the genus Paramecium.

Molecular and Cellular Biology ◽

10.1128/mcb.16.4.1871 ◽

1996 ◽

Vol 16 (4) ◽

pp. 1871-1879 ◽

Cited By ~ 43

Author(s):

M McCormick-Graham ◽

D P Romero

Keyword(s):

Secondary Structure ◽

Random Distribution ◽

Single Gene ◽

Telomerase Rna ◽

Paramecium Tetraurelia ◽

Dna Repeats ◽

Telomeric Dna ◽

Telomeric Repeats ◽

Precise Nature ◽

Rna Genes

Paramecium telomeric DNA consists largely of a random distribution of TTGGGG and TTTGGG repeats. Given the precise nature of other ciliate telomerases, it has been postulated that there are two distinct types of the Paramecium enzyme, each synthesizing perfect telomeric repeats: one with a template RNA that specifies the addition of TTTGGG and the second dictating the synthesis of TTGGGG repeats. We have cloned and sequenced telomerase RNA genes from Paramecium tetraurelia, P. primaurelia, P. multimicronucleatum, and P. caudatum. Surprisingly, a single gene encodes telomerase RNA in all four species, although an apparently nontranscribed pseudogene is also present in the genome of P. primaurelia. The overall lengths of the telomerase RNAs range between 202 and 209 nucleotides, and they can be folded into a conserved secondary structure similar to that derived for other ciliate RNAs. All Paramecium telomerase RNAs examined include a template specific for the synthesis of TTGGGG telomeric repeats, which has not been posttranscriptionally edited to account for the conventional synthesis of TTTGGG repeats. On the basis of these results, possible mechanisms for the synthesis of variable telomeric repeats by Paramecium telomerase are discussed.

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