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1.
《Cell cycle (Georgetown, Tex.)》2013,12(16):3177-3185
Telomeres consist of an elaborate, higher-order DNA architecture, and a suite of proteins that provide protection for the chromosome terminus by blocking inappropriate recombination and nucleolytic attack, and facilitate telomeric DNA replication by physical interactions with telomerase and the lagging strand replication machinery. The prevailing view has been that two distinct telomere capping complexes evolved, shelterin in vertebrates and a trimeric complex comprised of Cdc13, Stn1 and Ten1 (CST) in yeast. The recent discovery of a CST-like complex in plants and humans raises new questions about the composition of telomeres and their regulatory mechanisms in multicellular eukaryotes. In this review we discuss the evolving functions and interactions of CST components and their contributions to chromosome end protection and DNA replication. 相似文献
2.
In the budding yeast Saccharomyces cerevisiae, the structure and function of telomeres are maintained by binding proteins, such as Cdc13-Stn1-Ten1 (CST), Yku, and the telomerase complex. Like CST and Yku, telomerase also plays a role in telomere protection or capping. Unlike CST and Yku, however, the underlying molecular mechanism of telomerase-mediated telomere protection remains unclear. In this study, we employed both the CDC13-EST1 fusion gene and the separation-of-function allele est1-D514A to elucidate that Est1 provided a telomere protection pathway that was independent of both the CST and Yku pathways. Est1's ability to convert single-stranded telomeric DNA into a G quadruplex was required for telomerase-mediated telomere protection function. Additionally, Est1 maintained the integrity of telomeres by suppressing the recombination of subtelomeric Y' elements. Our results demonstrate that one major functional role that Est1 brings to the telomerase complex is the capping or protection of telomeres. 相似文献
3.
The maintenance of an appropriate number of telomere repeats by telomerase is essential for proper chromosome protection. The action of telomerase at the telomere terminus is regulated by opposing activities that either recruit/activate the enzyme at shorter telomeres or inhibit it at longer ones, thus achieving a stable average telomere length. To elucidate the mechanistic details of telomerase regulation we engineered specific chromosome ends in yeast so that a single telomere could be suddenly shortened and, as a consequence of its reduced length, elongated by telomerase. We show that shortened telomeres replicate early in S phase, unlike normal-length telomeres, due to the early firing of origins of DNA replication in subtelomeric regions. Early telomere replication correlates with increased telomere length and telomerase activity. These data reveal an epigenetic effect of telomere length on the activity of nearby replication origins and an unanticipated link between telomere replication timing and telomerase action. 相似文献
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The telomere complex must allow nucleases and helicases to process chromosome ends to make them substrates for telomerase, while preventing these same activities from disrupting chromosome end-protection. Replication protein A (RPA) binds to single-stranded DNA and is required for DNA replication, recombination, repair, and telomere maintenance. In fission yeast, the telomere binding protein Taz1 protects telomeres and negatively regulates telomerase. Here, we show that taz1-d rad11-D223Y double mutants lose their telomeric DNA, indicating that RPA (Rad11) and Taz1 are synergistically required to prevent telomere loss. Telomere loss in the taz1-d rad11-D223Y double mutants was suppressed by additional mutation of the helicase domain in a RecQ helicase (Rqh1), or by overexpression of Pot1, a single-strand telomere binding protein that is essential for protection of chromosome ends. From our results, we propose that in the absence of Taz1 and functional RPA, Pot1 cannot function properly and the helicase activity of Rqh1 promotes telomere loss. Our results suggest that controlling the activity of Rqh1 at telomeres is critical for the prevention of genomic instability. 相似文献
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Dennis Kappei Falk Butter Christian Benda Marion Scheibe Irena Draškovič Michelle Stevense Clara Lopes Novo Claire Basquin Masatake Araki Kimi Araki Dragomir Blazhev Krastev Ralf Kittler Rolf Jessberger J Arturo Londoño‐Vallejo Matthias Mann Frank Buchholz 《The EMBO journal》2013,32(12):1681-1701
Telomeres are repetitive DNA structures that, together with the shelterin and the CST complex, protect the ends of chromosomes. Telomere shortening is mitigated in stem and cancer cells through the de novo addition of telomeric repeats by telomerase. Telomere elongation requires the delivery of the telomerase complex to telomeres through a not yet fully understood mechanism. Factors promoting telomerase–telomere interaction are expected to directly bind telomeres and physically interact with the telomerase complex. In search for such a factor we carried out a SILAC‐based DNA–protein interaction screen and identified HMBOX1, hereafter referred to as homeobox telomere‐binding protein 1 (HOT1). HOT1 directly and specifically binds double‐stranded telomere repeats, with the in vivo association correlating with binding to actively processed telomeres. Depletion and overexpression experiments classify HOT1 as a positive regulator of telomere length. Furthermore, immunoprecipitation and cell fractionation analyses show that HOT1 associates with the active telomerase complex and promotes chromatin association of telomerase. Collectively, these findings suggest that HOT1 supports telomerase‐dependent telomere elongation. 相似文献
8.
Etheridge KT Compton SA Barrientos KS Ozgur S Griffith JD Counter CM 《The Journal of biological chemistry》2008,283(11):6935-6941
Mammalian telomeres are composed of G-rich repetitive double-stranded (ds) DNA with a 3' single-stranded (ss) overhang and associated proteins that together maintain chromosome end stability. Complete replication of telomeric DNA requires de novo elongation of the ssDNA by the enzyme telomerase, with telomeric proteins playing a key role in regulating telomerase-mediated telomere replication. In regards to the protein component of mammalian telomeres, TRF1 and TRF2 bind to the dsDNA of telomeres, whereas POT1 binds to the ssDNA portion. These three proteins are linked through either direct interactions or by the proteins TIN2 and TPP1. To determine the biological consequence of connecting telomeric dsDNA to ssDNA through a multiprotein assembly, we compared the effect of expressing TRF1 and POT1 in trans versus in cis in the form of a fusion of these two proteins, on telomere length in telomerase-positive cells. When expressed in trans these two proteins induced extensive telomere elongation. Fusing TRF1 to POT1 abrogated this effect, inducing mild telomere shortening, and generated looped DNA structures, as assessed by electron microscopy, consistent with the protein forming a complex with dsDNA and ssDNA. We speculate that such a protein bridge between dsDNA and ssDNA may inhibit telomerase access, promoting telomere shortening. 相似文献
9.
Andrea Puglisi Alessandro Bianchi Laure Lemmens Pascal Damay David Shore 《The EMBO journal》2008,27(17):2328-2339
The budding yeast Cdc13, Stn1 and Ten1 (CST) proteins are proposed to function as an RPA-like complex at telomeres that protects (‘caps'') chromosome ends and regulates their elongation by telomerase. We show that Stn1 has a critical function in both processes through the deployment of two separable domains. The N terminus of Stn1 interacts with Ten1 and carries out its essential capping function. The C terminus of Stn1 binds both Cdc13 and Pol12, and we present genetic data indicating that the Stn1–Cdc13 interaction is required to limit continuous telomerase action. Stn1 telomere association, similar to that of Cdc13, peaks during S phase. Significantly, the magnitude of Stn1 telomere binding is independent of telomere TG tract length, suggesting that the negative effect of Stn1 on telomerase action might be regulated by a modification of CST activity or structure in cis at individual telomeres. Genetic analysis suggests that the Tel1 kinase exerts an effect in parallel with the Stn1 C terminus to counteract its inhibition of telomerase. These data provide new insights into the coordination of telomere capping and telomerase regulation. 相似文献
10.
Telomeric chromatin: replicating and wrapping up chromosome ends 总被引:14,自引:0,他引:14
Shore D 《Current opinion in genetics & development》2001,11(2):189-198
Recent advances in our understanding of the specialized chromatin structure at telomeres, the ends of eukaryotic chromosomes, have focused on three separate areas: replication of telomeres through the coordinated action of conventional DNA polymerases and the telomerase enzyme, protection of the chromosome end from DNA damage checkpoint sensors and DNA-repair processes, and the discovery of a novel deacetylase enzyme (Sir2p) required for the establishment and maintenance of telomeric heterochromatin. Although the number of proteins and the complexity of their interactions at telomeres continues to grow, a picture of at least some of the major players and mechanisms underlying telomere replication, end 'capping' and chromatin assembly is beginning to emerge. 相似文献
11.
Genome stability necessitates a mechanism to protect the termini of linear chromosomes from inappropriate degradation or recombination. In many species this protection depends on 'capping' proteins that bind telomeric DNA. The budding yeast Cdc13p binds single-stranded telomeric sequences, prevents lethal degradation of chromosome ends and regulates telomere extension by telomerase. Two Cdc13-interacting proteins, Stn1p and Ten1p, are also required for viability and telomere length regulation. It has been proposed that Cdc13p DNA binding directs a Cdc13p-Stn1p-Ten1p complex to telomeres to mediate end protection. However, the functional significance of these protein interactions, and their respective roles in maintaining telomere integrity, remain undefined. Here, we show that co-overexpressing TEN1 with a truncated form of STN1 efficiently bypasses the essential role of CDC13. We further show that this truncated Stn1p binds directly to Pol12p, a polymerase alpha-primase regulatory subunit, and that Pol12 activity is required for CDC13 bypass. Thus, Stn1p and Ten1p control a Cdc13p-independent telomere capping mechanism that is coupled to the conventional DNA replication machinery. 相似文献
12.
Chang-Ching Liu Veena Gopalakrishnan Lai-Fong Poon TingDong Yan Shang Li 《Molecular and cellular biology》2014,34(1):57-70
In budding yeast (Saccharomyces cerevisiae), the cell cycle-dependent telomere elongation by telomerase is controlled by the cyclin-dependent kinase 1 (Cdk1). The telomere length homeostasis is balanced between telomerase-unextendable and telomerase-extendable states that both require Cdc13. The recruitment of telomerase complex by Cdc13 promotes telomere elongation, while the formation of Cdc13-Stn1-Ten1 (CST) complex at the telomere blocks telomere elongation by telomerase. However, the cellular signaling that regulates the timing of the telomerase-extendable and telomerase-unextendable states is largely unknown. Phosphorylation of Cdc13 by Cdk1 promotes the interaction between Cdc13 and Est1 and hence telomere elongation. Here, we show that Cdk1 also phosphorylates Stn1 at threonine 223 and serine 250 both in vitro and in vivo, and these phosphorylation events are essential for the stability of the CST complexes at the telomeres. By controlling the timing of Cdc13 and Stn1 phosphorylations during cell cycle progression, Cdk1 regulates the temporal recruitment of telomerase complexes and CST complexes to the telomeres to facilitate telomere maintenance. 相似文献
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Peili Gu Shuting Jia Taylor Takasugi Eric Smith Jayakrishnan Nandakumar Eric Hendrickson Sandy Chang 《Aging cell》2018,17(4)
Coats plus (CP) is a rare autosomal recessive disorder caused by mutations in CTC1, a component of the CST (CTC1, STN1, and TEN1) complex important for telomere length maintenance. The molecular basis of how CP mutations impact upon telomere length remains unclear. The CP CTC1L1142H mutation has been previously shown to disrupt telomere maintenance. In this study, we used CRISPR/Cas9 to engineer this mutation into both alleles of HCT116 and RPE cells to demonstrate that CTC1:STN1 interaction is required to repress telomerase activity. CTC1L1142H interacts poorly with STN1, leading to telomerase‐mediated telomere elongation. Impaired interaction between CTC1L1142H:STN1 and DNA Pol‐α results in increased telomerase recruitment to telomeres and further telomere elongation, revealing that C:S binding to DNA Pol‐α is required to fully repress telomerase activity. CP CTC1 mutants that fail to interact with DNA Pol‐α resulted in loss of C‐strand maintenance and catastrophic telomere shortening. Our findings place the CST complex as an important regulator of both G‐strand extensions by telomerase and C‐strand synthesis by DNA Pol‐α. 相似文献
15.
Telomeres in drag: Dressing as DNA damage to engage telomerase 总被引:1,自引:0,他引:1
The telomere field concentrates both on mechanisms of telomere synthesis and the mechanisms by which telomeres protect chromosome termini from fusion and degradation. Recent studies show that the DNA damage response (DDR) machinery, formerly thought to be the culprit in deleterious telomeric fusion and degradation reactions, plays an active role not only in telomere protection but also in regulating telomere synthesis. Conversely, semi-conservative DNA replication, responsible for the bulk of telomere synthesis, now appears to be a pivotal event on the road to telomere de-protection. These advances prompt the notion that the two guises of telomere function are intricately entangled. Indeed, telomeres appear to expose themselves to the DDR upon passage of the replication fork, in turn attracting telomerase. 相似文献
16.
In Saccharomyces cerevisiae, telomere replication occurs in late S phase and is accompanied by dynamic remodeling of its protein components. Here, we show that MRX (Mre11-Rad50-Xrs2), an evolutionarily conserved protein complex involved in DNA double-strand break (DSB) repair, is recruited to the telomeres in late S phase. MRX is required for the late S phase-specific recruitment of ATR-like kinase Mec1 to the telomeres. Mec1, in turn, contributes to the assembly of the telomerase regulators Cdc13 and Est1 at the telomere ends. Our results provide a model for the hierarchical assembly of telomere-replication proteins in late S phase; this involves triggering by the loading of MRX onto the chromosome termini. The recruitment of DNA repair-related proteins to the telomeres at particular times in the cell cycle suggests that the normal terminus of a chromosome is recognized as a DSB during the course of replication. 相似文献
17.
CTC1 deletion results in defective telomere replication, leading to catastrophic telomere loss and stem cell exhaustion 总被引:1,自引:0,他引:1
The proper maintenance of telomeres is essential for genome stability. Mammalian telomere maintenance is governed by a number of telomere binding proteins, including the newly identified CTC1-STN1-TEN1 (CST) complex. However, the in vivo functions of mammalian CST remain unclear. To address this question, we conditionally deleted CTC1 from mice. We report here that CTC1 null mice experience rapid onset of global cellular proliferative defects and die prematurely from complete bone marrow failure due to the activation of an ATR-dependent G2/M checkpoint. Acute deletion of CTC1 does not result in telomere deprotection, suggesting that mammalian CST is not involved in capping telomeres. Rather, CTC1 facilitates telomere replication by promoting efficient restart of stalled replication forks. CTC1 deletion results in increased loss of leading C-strand telomeres, catastrophic telomere loss and accumulation of excessive ss telomere DNA. Our data demonstrate an essential role for CTC1 in promoting efficient replication and length maintenance of telomeres. 相似文献
18.
Protection of Telomeres 1 (POT1) is a conserved nucleic acid binding protein implicated in both telomere replication and chromosome end protection. We previously showed that Arabidopsis thaliana POT1a associates with the TER1 telomerase RNP, and is required for telomere length maintenance in vivo. Here we further dissect the function of POT1a and explore its interplay with the CST (CTC1/STN1/TEN1) telomere complex. Analysis of pot1a null mutants revealed that POT1a is not required for telomerase recruitment to telomeres, but is required for telomerase to maintain telomere tracts. We show that POT1a stimulates the synthesis of long telomere repeat arrays by telomerase, likely by enhancing repeat addition processivity. We demonstrate that POT1a binds STN1 and CTC1 in vitro, and further STN1 and CTC1, like POT1a, associate with enzymatically active telomerase in vivo. Unexpectedly, the in vitro interaction of STN1 with TEN1 and POT1a was mutually exclusive, indicating that POT1a and TEN1 may compete for the same binding site on STN1 in vivo. Finally, unlike CTC1 and STN1, TEN1 was not associated with active telomerase in vivo, consistent with our previous data showing that TEN1 negatively regulates telomerase enzyme activity. Altogether, our data support a two-state model in which POT1a promotes an extendable telomere state via contacts with the telomerase RNP as well as STN1 and CTC1, while TEN1 opposes these functions. 相似文献
19.
Luciano P Coulon S Faure V Corda Y Bos J Brill SJ Gilson E Simon MN Géli V 《The EMBO journal》2012,31(8):2034-2046
In Saccharomyces cerevisiae, the telomerase complex binds to chromosome ends and is activated in late S-phase through a process coupled to the progression of the replication fork. Here, we show that the single-stranded DNA-binding protein RPA (replication protein A) binds to the two daughter telomeres during telomere replication but only its binding to the leading-strand telomere depends on the Mre11/Rad50/Xrs2 (MRX) complex. We further demonstrate that RPA specifically co-precipitates with yKu, Cdc13 and telomerase. The interaction of RPA with telomerase appears to be mediated by both yKu and the telomerase subunit Est1. Moreover, a mutation in Rfa1 that affects both the interaction with yKu and telomerase reduces the dramatic increase in telomere length of a rif1Δ, rif2Δ double mutant. Finally, we show that the RPA/telomerase association and function are conserved in Schizosaccharomyces pombe. Our results indicate that in both yeasts, RPA directly facilitates telomerase activity at chromosome ends. 相似文献
20.
Replication proteins influence the maintenance of telomere length and telomerase protein stability
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We investigated the effects of fission yeast replication genes on telomere length maintenance and identified 20 mutant alleles that confer lengthening or shortening of telomeres. The telomere elongation was telomerase dependent in the replication mutants analyzed. Furthermore, the telomerase catalytic subunit, Trt1, and the principal initiation and lagging-strand synthesis DNA polymerase, Polalpha, were reciprocally coimmunoprecipitated, indicating these proteins physically coexist as a complex in vivo. In a polalpha mutant that exhibited abnormal telomere lengthening and slightly reduced telomere position effect, the cellular level of the Trt1 protein was significantly lower and the coimmunoprecipitation of Trt1 and Polalpha was severely compromised compared to those in the wild-type polalpha cells. Interestingly, ectopic expression of wild-type polalpha in this polalpha mutant restored the cellular Trt1 protein to the wild-type level and shortened the telomeres to near-wild-type length. These results suggest that there is a close physical relationship between the replication and telomerase complexes. Thus, mutation of a component of the replication complex can affect the telomeric complex in maintaining both telomere length equilibrium and telomerase protein stability. 相似文献