Intracellular trafficking of yeast telomerase components

doi:10.1093/EMBO-REPORTS/KVF133

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Intracellular trafficking of yeast telomerase components

Journal Article•DOI•

Intracellular trafficking of yeast telomerase components

M.Teresa Teixeira¹, Klaus Förstemann¹, Susan M. Gasser², Joachim Lingner¹•Institutions (2)

ISREC¹, University of Geneva²

01 Jul 2002-EMBO Reports (John Wiley & Sons, Ltd)-Vol. 3, Iss: 7, pp 652-659

TL;DR: It is found that Est1p, Est2p and TLC1 can migrate independently of each other to the nucleus and a role of the nucleolus in telomerase biogenesis is suggested.

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Abstract: Telomerase uses an internal RNA moiety as template for the synthesis of telomere repeats. In Saccharomyces cerevisiae, the telomerase holoenzyme contains the telomerase reverse transcriptase subunit Est2p, the telomerase RNA moiety TLC1, the telomerase associated proteins Est1p and Est3p, and Sm proteins. Here we assess telomerase assembly by determining the localization of telomerase components. We found that Est1p, Est2p and TLC1 can migrate independently of each other to the nucleus. With limiting amounts of TLC1, overexpressed Est1p and Est2p accumulated in the nucleolus, whereas enzymatically active Est2p–TLC1 complexes are distributed over the entire nucleus. The distribution to the nucleoplasm depended on the specific interaction between Est2p and TLC1 but was independent of Est1p and Est3p. Altogether, our results suggest a role of the nucleolus in telomerase biogenesis. We also describe experiments that support a transient cytoplasmic localization of TLC1 RNA.

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Journal Article•DOI•

The unusually large Plasmodium telomerase reverse-transcriptase localizes in a discrete compartment associated with the nucleolus

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Luisa M. Figueiredo¹, Eduardo P. C. Rocha, Liliana Mancio-Silva, Christine Prevost, Danièle Hernandez-Verdun, Artur Scherf - Show less +2 more•Institutions (1)

Pasteur Institute¹

01 Jan 2005-Nucleic Acids Research

TL;DR: The gene that encodes the catalytic reverse transcriptase (RT) component of this enzyme in the malaria parasite Plasmodium falciparum (PfTERT) is identified as well as the orthologous genes from two rodent and one simian malaria species.

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Abstract: Telomerase replicates chromosome ends, a function necessary for maintaining genome integrity. We have identified the gene that encodes the catalytic reverse transcriptase (RT) component of this enzyme in the malaria parasite Plasmodium falciparum (PfTERT) as well as the orthologous genes from two rodent and one simian malaria species. PfTERT is predicted to encode a basic protein that contains the major sequence motifs previously identified in known telomerase RTs (TERTs). At approximately 2500 amino acids, PfTERT is three times larger than other characterized TERTs. We observed remarkable sequence diversity between TERT proteins of different Plasmodial species, with conserved domains alternating with hypervariable regions. Immunofluorescence analysis revealed that PfTERT is expressed in asexual blood stage parasites that have begun DNA synthesis. Surprisingly, rather than at telomere clusters, PfTERT typically localizes into a discrete nuclear compartment. We further demonstrate that this compartment is associated with the nucleolus, hereby defined for the first time in P.falciparum.

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69 citations

Journal Article•DOI•

Telomere maintenance, function and evolution: the yeast paradigm

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Maria Teresa Teixeira¹, Eric Gilson¹•Institutions (1)

Centre national de la recherche scientifique¹

25 Aug 2005-Chromosome Research

TL;DR: Comparative genomic analyses show that telomeres have evolved rapidly among yeast species and functional plasticity emerges as one of the driving forces of this evolution.

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Abstract: Telomeres are multifunctional genetic elements that cap chromosome ends, playing essential roles in genome stability, chromosome higher-order organization and proliferation control. The telomere field has largely benefited from the study of unicellular eukaryotic organisms such as yeasts. Easy cultivation in laboratory conditions and powerful genetics have placed mainly Saccharomyces cerevisiae, Kluveromyces lactis and Schizosaccharomyces pombe as crucial model organisms for telomere biology research. Studies in these species have made it possible to elucidate the basic mechanisms of telomere maintenance, function and evolution. Moreover, comparative genomic analyses show that telomeres have evolved rapidly among yeast species and functional plasticity emerges as one of the driving forces of this evolution. This provides a precious opportunity to further our understanding of telomere biology.

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62 citations

Cites background from "Intracellular trafficking of yeast ..."

...2002, Figueiredo et al. 2005, Teixeira et al. 2002)....
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Journal Article•DOI•

Structure—function relationships in telomerase genes

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Eva Sýkorová¹, Jiří Fajkus², Jiří Fajkus¹•Institutions (2)

Academy of Sciences of the Czech Republic¹, Masaryk University²

01 Jul 2009-Biology of the Cell

TL;DR: The structure of TERT genes, their alternative splicing products and their functions are discussed, and whether particular findings are generally applicable to telomerases or species‐specific are explored.

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57 citations

Cites background from "Intracellular trafficking of yeast ..."

...…connected, and differential regulation raises questions of intracellular/intranuclear trafficking and their assembly to form an active telomerase (Teixeira et al., 2002; Tomlinson et al., 2006; for reviews see Collins, 2006; Hug and Lingner, 2006; Cairney and Keith, 2008; Gallardo et al., 2008)....
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Journal Article•DOI•

A template-proximal RNA paired element contributes to Saccharomyces cerevisiae telomerase activity.

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Anita G. Seto¹, Kfir Umansky, Yehuda Tzfati, Arthur J. Zaug, Elizabeth H. Blackburn, Thomas R. Cech - Show less +2 more•Institutions (1)

Howard Hughes Medical Institute¹

01 Nov 2003-RNA

TL;DR: The identification of a paired element located immediately 5' of the template region in the Saccharomyces cerevisiae telomerase RNA indicates that the RNA element has additional functions in enzyme processivity and in directing template usage by telomersase.

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Abstract: The ribonucleoprotein complex telomerase is critical for replenishing chromosome-end sequence during eukaryotic DNA replication. The template for the addition of telomeric repeats is provided by the RNA component of telomerase. However, in budding yeast, little is known about the structure and function of most of the remainder of the telomerase RNA. Here, we report the identification of a paired element located immediately 5 of the template region in the Saccharomyces cerevisiae telomerase RNA. Mutations disrupting or replacing the helical element showed that this structure, but not its exact nucleotide sequence, is important for telomerase function in vivo and in vitro. Biochemical characterization of a paired element mutant showed that the mutant generated longer products and incorporated noncognate nucleotides. Sequencing of in vivo synthesized telomeres from this mutant showed that DNA synthesis proceeded beyond the normal template. Thus, the S. cerevisiae element resembles a similar element found in Kluyveromyces budding yeasts with respect to a function in template boundary specification. In addition, the in vitro activity of the paired element mutant indicates that the RNA element has additional functions in enzyme processivity and in directing template usage by telomerase.

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55 citations

Cites background or methods from "Intracellular trafficking of yeast ..."

...…DMB fragments into the StuI and HpaI sites of pSD107. pRS314-PGAL1-TLC1 (TRP, CEN, GAL-TLC1) was a gift of J. Lingner, ISREC, Lausanne, Switzerland (Teixeira et al. 2002). pGAL-DMA, pGAL-DMB, and pGAL-DMC were constructed by subcloning the BsrGI to NotI fragment from pTLC1DMA, pTLC1-DMB, or…...
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...DMC was generated by triple ligation of the StuI– NcoI DMA and NcoI–HpaI DMB fragments into the StuI and HpaI sites of pSD107. pRS314-PGAL1-TLC1 (TRP, CEN, GAL-TLC1) was a gift of J. Lingner, ISREC, Lausanne, Switzerland (Teixeira et al. 2002). pGAL-DMA, pGAL-DMB, and pGAL-DMC were constructed by subcloning the BsrGI to NotI fragment from pTLC1DMA, pTLC1-DMB, or pTLC1-DMC into the corresponding sites in pRS314-PGAL1-TLC1....
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...It has been previously shown that overexpression of both the RNA and reverse transcriptase components of telomerase results in a significant increase in in vitro activity (Teixeira et al. 2002)....
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Journal Article•DOI•

Hypermethylation of yeast telomerase RNA by the snRNA and snoRNA methyltransferase Tgs1

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Jacqueline Franke¹, Jessica Gehlen¹, Ann E. Ehrenhofer-Murray¹•Institutions (1)

University of Duisburg-Essen¹

01 Nov 2008-Journal of Cell Science

TL;DR: It is reported that the yeast snRNA and snoRNA methyltransferase Tgs1 is responsible for TLC1 m3G cap formation and that the loss of the m 3G cap of TLC2 causes premature aging.

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Abstract: Telomerase in Saccharomyces cerevisiae consists of three protein subunits and the RNA moiety TLC1, which together ensure the complete replication of chromosome ends. TLC1 shares several features with snRNA, among them the presence of a trimethylguanosine (m3G) cap structure at the 5′ end of the RNA. Here, we report that the yeast snRNA and snoRNA methyltransferase Tgs1 is responsible for TLC1 m3G cap formation. The absence of Tgs1 caused changes in telomere length and structure, improved telomeric silencing and stabilized telomeric recombination. Genetic analyses implicated a role for the TLC1 m3G cap in the coordination between telomerase and DNA polymerase for end replication. Furthermore, tgs1Δ cells displayed a shortened replicative lifespan, suggesting that the loss of the m3G cap of TLC1 causes premature aging.

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50 citations

Cites background from "Intracellular trafficking of yeast ..."

...cytoplasm and nucleus (Teixeira et al., 2002), that the importin...
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...Given that TLC1 can shuttle between cytoplasm and nucleus (Teixeira et al., 2002), that the importin Mtr10 is required for full telomerase activity (Ferrezuelo et al., 2002), and that Tgs1 is located in the nucleolus (Mouaikel et al., 2002), our data suggest the following alternative scenarios: (1)…...
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Journal Article•DOI•

A telomeric sequence in the RNA of Tetrahymena telomerase required for telomere repeat synthesis.

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Carol W. Greider¹, Elizabeth H. Blackburn²•Institutions (2)

Cold Spring Harbor Laboratory¹, University of California, Berkeley²

26 Jan 1989-Nature

TL;DR: The essential RNA component of this ribonucleoprotein enzyme has now been cloned and found to contain the sequence CAACCCCAA, which seems to be the template for the synthesis of TTGGGG repeats.

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Abstract: The telomerase enzyme of Tetrahymena synthesizes repeats of the telomeric DNA sequence TTGGGG de novo in the absence of added template. The essential RNA component of this ribonucleoprotein enzyme has now been cloned and found to contain the sequence CAACCCCAA, which seems to be the template for the synthesis of TTGGGG repeats.

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1,623 citations

"Intracellular trafficking of yeast ..." refers background in this paper

...Telomerase is a ribonucleoprotein (RNP) polymerase that uses an internal RNA moiety as template for the synthesis of telomere repeats (Greider and Blackburn, 1989; Lingner et al., 1997b)....
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Journal Article•DOI•

Reverse Transcriptase Motifs in the Catalytic Subunit of Telomerase

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Joachim Lingner¹, Timothy P. Hughes², Andrej Shevchenko², Matthias Mann², Victoria Lundblad², Thomas R. Cech² - Show less +2 more•Institutions (2)

Howard Hughes Medical Institute¹, Baylor College of Medicine²

25 Apr 1997-Science

TL;DR: The reverse transcriptase protein fold, previously known to be involved in retroviral replication and retrotransposition, is essential for normal chromosome telomere replication in diverse eukaryotes.

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Abstract: Telomerase is a ribonucleoprotein enzyme essential for the replication of chromosome termini in most eukaryotes. Telomerase RNA components have been identified from many organisms, but no protein component has been demonstrated to catalyze telomeric DNA extension. Telomerase was purified from Euplotes aediculatus, a ciliated protozoan, and one of its proteins was partially sequenced by nanoelectrospray tandem mass spectrometry. Cloning and sequence analysis of the corresponding gene revealed that this 123-kilodalton protein (p123) contains reverse transcriptase motifs. A yeast (Saccharomyces cerevisiae) homolog was found and subsequently identified as EST2 (ever shorter telomeres), deletion of which had independently been shown to produce telomere defects. Introduction of single amino acid substitutions within the reverse transcriptase motifs of Est2 protein led to telomere shortening and senescence in yeast, indicating that these motifs are important for catalysis of telomere elongation in vivo. In vitro telomeric DNA extension occurred with extracts from wild-type yeast but not from est2 mutants or mutants deficient in telomerase RNA. Thus, the reverse transcriptase protein fold, previously known to be involved in retroviral replication and retrotransposition, is essential for normal chromosome telomere replication in diverse eukaryotes.

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1,293 citations

"Intracellular trafficking of yeast ..." refers background or result in this paper

...Telomerase is a ribonucleoprotein (RNP) polymerase that uses an internal RNA moiety as template for the synthesis of telomere repeats (Greider and Blackburn, 1989; Lingner et al., 1997b)....
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...Thus, Est1p and Est3p do not influence the steady-state localization of Est2p and TLC1 or their association, which is consistent with previous studies showing that Est1p and Est3p are not required for telomerase activity in vitro (Cohn and Blackburn, 1995; Lingner et al., 1997a)....
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...Active telomerase is localized in the nucleoplasm Telomerase activity requires Est2p and TLC1, which together make up the catalytic core (Lingner et al., 1997b)....
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Journal Article•DOI•

A telomerase component is defective in the human disease dyskeratosis congenita

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James R. Mitchell¹, Emily Wood¹, Kathleen Collins¹•Institutions (1)

University of California, Berkeley¹

02 Dec 1999-Nature

TL;DR: It is found that primary fibroblasts and lymphoblasts from DKC-affected males are not detectably deficient in conventional H/ACA small nucleolar RNA accumulation or function; however, DKC cells have a lower level of telomerase RNA, produce lower levels of telomersase activity and have shorter telomeres than matched normal cells.

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Abstract: The X-linked form of the human disease dyskeratosis congenita (DKC) is caused by mutations in the gene encoding dyskerin1. Sufferers have defects in highly regenerative tissues such as skin and bone marrow, chromosome instability and a predisposition to develop certain types of malignancy. Dyskerin is a putative pseudouridine synthase, and it has been suggested that DKC may be caused by a defect in ribosomal RNA processing. Here we show that dyskerin is associated not only with H/ACA small nucleolar RNAs2, but also with human telomerase RNA, which contains an H/ACA RNA motif3. Telomerase adds simple sequence repeats to chromosome ends using an internal region of its RNA as a template4, and is required for the indefinite proliferation of primary human cells5. We find that primary fibroblasts and lymphoblasts from DKC-affected males are not detectably deficient in conventional H/ACA small nucleolar RNA accumulation or function; however, DKC cells have a lower level of telomerase RNA, produce lower levels of telomerase activity and have shorter telomeres than matched normal cells. The pathology of DKC is consistent with compromised telomerase function leading to a defect in telomere maintenance, which may limit the proliferative capacity of human somatic cells in epithelia and blood.

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1,122 citations

"Intracellular trafficking of yeast ..." refers background in this paper

...In support of this, it was shown that vertebrate telomerase RNA contains an H/ACA motif that targets the RNA to nucleoli where it was hypothesized to associate with the catalytic subunit of telomerase (Mitchell et al., 1999; Lukowiak et al., 2001)....
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Journal Article•DOI•

A mutant with a defect in telomere elongation leads to senescence in yeast.

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Victoria Lundblad¹, Jack W. Szostak¹•Institutions (1)

Harvard University¹

19 May 1989-Cell

TL;DR: Using this assay, a mutant that displays a progressive decrease in telomere length as well as an increased frequency of chromosome loss is isolated, which defines a new gene, designated EST1 (for ever shorter telomeres).

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931 citations

Journal Article•DOI•

Ribosome synthesis in Saccharomyces cerevisiae.

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J. Venema¹, David Tollervey•Institutions (1)

VU University Amsterdam¹

01 Jan 1999-Annual Review of Genetics

TL;DR: The recent, and often surprising, advances in the understanding of ribosome synthesis in the yeast Saccharomyces cerevisiae will underscore the unexpected complexity of eukaryotic ribosomes synthesis.

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Abstract: The synthesis of ribosomes is one of the major metabolic pathways in all cells. In addition to around 75 individual ribosomal proteins and 4 ribosomal RNAs, synthesis of a functional eukaryotic ribosome requires a remarkable number of trans-acting factors. Here, we will discuss the recent, and often surprising, advances in our understanding of ribosome synthesis in the yeast Saccharomyces cerevisiae. These will underscore the unexpected complexity of eukaryotic ribosome synthesis.

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779 citations

"Intracellular trafficking of yeast ..." refers background in this paper

...Since the nucleolus is considered to be a site with a high concentration of trans-acting factors required for the assembly of ribosomes and other RNPs (Pederson, 1998; Sleeman and Lamond, 1999; Venema and Tollervey, 1999), it may provide factors that assist in the assembly of telomerase....
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