<i>Euplotes crassus</i>has genes encoding telomere-binding proteins and telomere-binding protein homologs

Wang, Wenlan; Skopp, Rose; Scofield, Margaret A.; Price, Carolyn M.

doi:10.1093/nar/20.24.6621

Cited by 70 publications

(64 citation statements)

References 50 publications

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“…Such an S-phase-specific function would explain the low levels of POT1b in asynchronous populations of cells and our difficulty in detecting POT1b binding to telomeres. Indeed, a POT1 paralog with such an S-phasespecific function was identified for Euplotes some years ago (44). The Euplotes replication telomere protein (rTP) is expressed only in replicating cells, and the protein localizes very specifically to replication bands, the sites of DNA replication in the Euplotes macronucleus (42).…”

Section: Discussionmentioning

confidence: 99%

“…This lack of a severe telomereuncapping phenotype, together with the unexpected finding that POT1 is a component of the core telomere protein complex, suggests that the function of POT1 is more com-plex than first envisioned. Interestingly, while humans have only one POT1 protein, Euplotes, plants, and mice have two or more TEBP/POT1 paralogs which show some separation of function (16,39,44). In particular, the mouse POT1b seems to regulate resection of the telomeric C strand, while POT1a plays a greater role in preventing a telomeric DNA damage response (16).…”

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confidence: 99%

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Tetrahymena POT1a Regulates Telomere Length and Prevents Activation of a Cell CycleCheckpoint

Jacob

Lescasse

Linger

et al. 2007

Molecular and Cellular Biology

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The POT1/TEBP telomere proteins are a group of single-stranded DNA (ssDNA)-binding proteins that have long been assumed to protect the G overhang on the telomeric 3 strand. We have found that the Tetrahymena thermophila genome contains two POT1 gene homologs, POT1a and POT1b. The POT1a gene is essential, but POT1b is not. We have generated a conditional POT1a cell line and shown that POT1a depletion results in a monster cell phenotype and growth arrest. However, G-overhang structure is essentially unchanged, indicating that POT1a is not required for overhang protection. In contrast, POT1a is required for telomere length regulation. After POT1a depletion, most telomeres elongate by 400 to 500 bp, but some increase by up to 10 kb. This elongation occurs in the absence of further cell division. The growth arrest caused by POT1a depletion can be reversed by reexpression of POT1a or addition of caffeine. Thus, POT1a is required to prevent a cell cycle checkpoint that is most likely mediated by ATM or ATR (ATM and ATR are protein kinases of the PI-3 protein kinase-like family). Our findings indicate that the essential function of POT1a is to prevent a catastrophic DNA damage response. This response may be activated when nontelomeric ssDNA-binding proteins bind and protect the G overhang.In order to maintain genome integrity, the telomeric DNA from cells with linear chromosomes is packaged into a protective nucleoprotein complex (10). In the absence of this complex, the telomeres are recognized as DNA damage and subject to repair by nonhomologous end joining (33). The resulting chromosome fusions lead to genome instability (36). The protective telomeric complex is composed of a series of unique telomere proteins that bind the double-stranded region of the telomeric DNA and/or the single-strand overhang on the 3Ј G-rich strand (37). Although the exact composition of the complex varies between species, vertebrates, yeasts, plants, and ciliates all use a series of structurally related proteins to protect their telomeres. The G-overhang binding proteins all bind Gstrand DNA through a conserved OB fold motif, while the double-stranded DNA-binding proteins bind via a conserved myb motif (23,37,46).In vertebrate cells, telomeres are packaged by a core complex of six proteins which function both in telomere protection and in telomere length regulation (10). This core complex (which is sometimes called shelterin) contains two doublestranded DNA-binding proteins, TRF1 and TRF2, which anchor the complex along the length of the telomere, and the TRF1/2 interaction partners Rap1, TIN2, and TPP1. The Gstrand binding protein POT1 is also a component of the complex. Although POT1 is secured into the complex through interactions with TPP1 (17, 31, 50), POT1 molecules are also thought to bind the G-strand overhang via their OB foldcontaining DNA-binding domain (45,46). Additional telomere-associated proteins include a number of DNA damage response factors (33). However, these factors appear to bind only transiently during replication of...

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

Tetrahymena POT1a Regulates Telomere Length and Prevents Activation of a Cell CycleCheckpoint

Jacob

Lescasse

Linger

et al. 2007

Molecular and Cellular Biology

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“…The ciliates Oxytrichia and Euplotes have a telomere end-binding protein, the ab heterodimer, that caps the ends of their telomeres, preventing degradation (Gray et al, 1991;Hicke et al, 1990;Wang et al, 1992). A homologue of the a subunit, Pot1, was recently identi®ed in S pombe and humans and was shown to bind G-strand telomeric DNA (Baumann and Cech, 2001).…”

Section: Introductionmentioning

confidence: 99%

Balancing instability: dual roles for telomerase and telomere dysfunction in tumorigenesis

2002

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“…Euplotes crassus genes encoding a basic 446-aa telomere protein and an acidic 460-aa telomere protein homologue were identified on the basis of sequence similarity with a gene for the Oxytricha nova alpha telomere protein (40). The 460-aa telomere protein homologue (EcRTP in Figure 1) is transiently expressed coincident with DNA synthesis and is thought to be involved with telomere replication (35).…”

Section: Introductionmentioning

confidence: 99%

DNA Binding Affinity and Sequence Permutation Preference of the Telomere Protein from Euplotes crassus

et al. 2006

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Telomere end binding proteins from diverse organisms use various forms of an ancient protein structure to recognize and bind with single strand DNA found at the ends of telomeres. To further understand the biochemistry and evolution of these proteins we have characterized the DNA-binding properties of the telomere end binding protein from Euplotes crassus (EcTEBP). EcTEBP and its predicted amino-terminal DNA-binding domain, EcTEBP-N, were expressed in E. coli and purified. Each protein formed stoichiometric (1:1) complexes with single strand DNA oligos derived from the precisely defined d(TTTTGGGGTTTTGG) sequence found at DNA termini in Euplotes. Dissociation constants for DNA•EcTEBP and DNA•EcTEBP-N were comparable, with K D-DNA = 38 ± 2 nM for the full-length protein and K D-DNA = 60 ± 4 nM for the N-terminal domain, indicating that the N-terminal domain retains high affinity for DNA even in the absence of potentially stabilizing moieties located in the C-terminal domain. Rate constants for DNA association and DNA dissociation corroborated a slightly improved DNA binding performance for the full-length protein (k a = 45 ± 4 μM -1 s -1 , k d = 0.10 ± 0.02 s -1 ) relative to the N-terminal domain (k a = 18 ± 1 μM -1 s -1 , k d = 0.15 ± 0.01 s -1 ). Equilibrium dissociation constants measured for sequence permutations of the telomere repeat spanned a 55 -1400 nM range, with EcTEBP and EcTEBP-N binding most tightly to d (TTGGGGTTTTGG) -the sequence corresponding with that of mature DNA termini. Additionally, competition experiments showed that EcTEBP recognizes and binds the telomere-derived 14-nucleotide DNA in preference to shorter 5′ -truncation variants. Compared with multi-subunit complexes assembled with telomere single strand DNA from Oxytricha nova, our results highlight the relative simplicity of the Euplotes crassus system where a telomere end binding protein has biochemical properties indicating one protein subunit caps the single strand DNA.

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Euplotes crassushas genes encoding telomere-binding proteins and telomere-binding protein homologs

Cited by 70 publications

References 50 publications

Tetrahymena POT1a Regulates Telomere Length and Prevents Activation of a Cell CycleCheckpoint

Tetrahymena POT1a Regulates Telomere Length and Prevents Activation of a Cell CycleCheckpoint

Balancing instability: dual roles for telomerase and telomere dysfunction in tumorigenesis

DNA Binding Affinity and Sequence Permutation Preference of the Telomere Protein from Euplotes crassus

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