scholarly journals RNA-DNA hybrids support recombination-based telomere maintenance in fission yeast

2018 ◽  
Author(s):  
Yan Hu ◽  
Henrietta W. Bennett ◽  
Na Liu ◽  
Martin Moravec ◽  
Jessica F. Williams ◽  
...  
Keyword(s):  
Schizosaccharomyces Pombe ◽  
Fission Yeast ◽  
Telomeric Repeat ◽  
Telomere Maintenance ◽  
Telomeric Dna ◽  
Homology Directed Repair ◽  
Alternative Lengthening ◽  
Linear Chromosomes ◽  
Severe Growth ◽  
Telomere Erosion

ABSTRACTA subset of cancers rely on telomerase-independent mechanisms to maintain their chromosome ends. The predominant “alternative lengthening of telomeres” pathway appears dependent on homology-directed repair (HDR) to maintain telomeric DNA. However, the molecular changes needed for cells to productively engage in telomeric HDR are poorly understood. To gain new insights into this transition, we monitored the state of telomeres during serial culture of fission yeast (Schizosaccharomyces pombe) lacking the telomerase recruitment factor Ccq1. Rad52 is loaded onto critically short telomeres shortly after germination despite continued telomere erosion, suggesting that recruitment of recombination factors is not sufficient to maintain telomeres in the absence of telomerase function. Instead, survivor formation coincides with the derepression of telomeric repeat-containing RNA (TERRA). In this context, degradation of TERRA associated with the telomere in the form of R-loops drives a severe growth crisis, ultimately leading to a novel type of survivor with linear chromosomes and altered cytological telomere characteristics, including the loss of the shelterin component Rap1 (but not the TRF1/TRF2 orthologue, Taz1) from the telomere. We demonstrate that deletion of Rap1 is protective in this context, preventing the growth crisis that is otherwise triggered by degradation of telomeric R-loops in survivors with linear chromosomes. These findings suggest that up-regulation of telomere-engaged TERRA or altered recruitment of shelterin components can support telomerase-independent telomere maintenance.

scholarly journals Alternative lengthening of telomeres in childhood neuroblastoma from genome to proteome

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Sabine A. Hartlieb ◽  
Lina Sieverling ◽  
Michal Nadler-Holly ◽  
Matthias Ziehm ◽  
Umut H. Toprak ◽  
...  
Keyword(s):  
High Frequency ◽  
Clinical Course ◽  
Population Based ◽  
Telomeric Repeat ◽  
Telomere Maintenance ◽  
Study Cohort ◽  
Alternative Lengthening Of Telomeres ◽  
Course Of Disease ◽  
Low Protein ◽  
Alternative Lengthening

AbstractTelomere maintenance by telomerase activation or alternative lengthening of telomeres (ALT) is a major determinant of poor outcome in neuroblastoma. Here, we screen for ALT in primary and relapsed neuroblastomas (n = 760) and characterize its features using multi-omics profiling. ALT-positive tumors are molecularly distinct from other neuroblastoma subtypes and enriched in a population-based clinical sequencing study cohort for relapsed cases. They display reduced ATRX/DAXX complex abundance, due to either ATRX mutations (55%) or low protein expression. The heterochromatic histone mark H3K9me3 recognized by ATRX is enriched at the telomeres of ALT-positive tumors. Notably, we find a high frequency of telomeric repeat loci with a neuroblastoma ALT-specific hotspot on chr1q42.2 and loss of the adjacent chromosomal segment forming a neo-telomere. ALT-positive neuroblastomas proliferate slowly, which is reflected by a protracted clinical course of disease. Nevertheless, children with an ALT-positive neuroblastoma have dismal outcome.

scholarly journals Nuclear receptors regulate alternative lengthening of telomeres through a novel noncanonical FANCD2 pathway

2019 ◽  
Vol 5 (10) ◽  
pp. eaax6366 ◽  
Author(s):  
Mafei Xu ◽  
Jun Qin ◽  
Leiming Wang ◽  
Hui-Ju Lee ◽  
Chung-Yang Kao ◽  
...  
Keyword(s):  
Nuclear Receptors ◽  
Direct Interaction ◽  
Telomere Maintenance ◽  
Telomeric Dna ◽  
Core Complex ◽  
Alternative Lengthening Of Telomeres ◽  
Independent Manner ◽  
Replication Complex ◽  
Alternative Lengthening ◽  
Key Steps

Alternative lengthening of telomeres (ALT) is known to use homologous recombination (HR) to replicate telomeric DNA in a telomerase-independent manner. However, the detailed process remains largely undefined. It was reported that nuclear receptors COUP-TFII and TR4 are recruited to the enriched GGGTCA variant repeats embedded within ALT telomeres, implicating nuclear receptors in regulating ALT activity. Here, we identified a function of nuclear receptors in ALT telomere maintenance that involves a direct interaction between COUP-TFII/TR4 and FANCD2, the key protein in the Fanconi anemia (FA) DNA repair pathway. The COUP-TFII/TR4-FANCD2 complex actively induces the DNA damage response by recruiting endonuclease MUS81 and promoting the loading of the PCNA-POLD3 replication complex in ALT telomeres. Furthermore, the COUP-TFII/TR4-mediated ALT telomere pathway does not require the FA core complex or the monoubiquitylation of FANCD2, key steps in the canonical FA pathway. Thus, our findings reveal that COUP-TFII/TR4 regulates ALT telomere maintenance through a novel noncanonical FANCD2 pathway.

scholarly journals G-quadruplex Stabilization Fuels the ALT Pathway in ALT-positive Osteosarcoma Cells

Genes ◽  
2020 ◽  
Vol 11 (3) ◽  
pp. 304 ◽  
Author(s):  
Roberta Amato ◽  
Martina Valenzuela ◽  
Francesco Berardinelli ◽  
Erica Salvati ◽  
Carmen Maresca ◽  
...  
Keyword(s):  
Dna Damage ◽  
Sister Chromatid Exchanges ◽  
Telomere Maintenance ◽  
Osteosarcoma Cell ◽  
Telomeric Dna ◽  
Replicative Stress ◽  
Alternative Lengthening ◽  
G Quadruplex ◽  
Reduce Cell Proliferation ◽  
The Impact

Most human tumors maintain telomere lengths by telomerase, whereas a portion of them (10–15%) uses a mechanism named alternative lengthening of telomeres (ALT). The telomeric G-quadruplex (G4) ligand RHPS4 is known for its potent antiproliferative effect, as shown in telomerase-positive cancer models. Moreover, RHPS4 is also able to reduce cell proliferation in ALT cells, although the influence of G4 stabilization on the ALT mechanism has so far been poorly investigated. Here we show that sensitivity to RHPS4 is comparable in ALT-positive (U2OS; SAOS-2) and telomerase-positive (HOS) osteosarcoma cell lines, unlinking the telomere maintenance mechanism and RHPS4 responsiveness. To investigate the impact of G4 stabilization on ALT, the cardinal ALT hallmarks were analyzed. A significant induction of telomeric doublets, telomeric clusterized DNA damage, ALT-associated Promyelocytic Leukaemia-bodies (APBs), telomere sister chromatid exchanges (T-SCE) and c-circles was found exclusively in RHPS4-treated ALT cells. We surmise that RHPS4 affects ALT mechanisms through the induction of replicative stress that in turn is converted in DNA damage at telomeres, fueling recombination. In conclusion, our work indicates that RHPS4-induced telomeric DNA damage promotes overactivation of telomeric recombination in ALT cells, opening new questions on the therapeutic employment of G4 ligands in the treatment of ALT positive tumors.

scholarly journals G-Quadruplexes at Telomeres: Friend or Foe?

Molecules ◽  
2020 ◽  
Vol 25 (16) ◽  
pp. 3686 ◽  
Author(s):  
Tracy M. Bryan
Keyword(s):  
Tandem Repeats ◽  
Genome Instability ◽  
Protein Complexes ◽  
Telomere Maintenance ◽  
Telomeric Dna ◽  
Positive Role ◽  
Telomere Biology ◽  
Linear Chromosomes

Telomeres are DNA-protein complexes that cap and protect the ends of linear chromosomes. In almost all species, telomeric DNA has a G/C strand bias, and the short tandem repeats of the G-rich strand have the capacity to form into secondary structures in vitro, such as four-stranded G-quadruplexes. This has long prompted speculation that G-quadruplexes play a positive role in telomere biology, resulting in selection for G-rich tandem telomere repeats during evolution. There is some evidence that G-quadruplexes at telomeres may play a protective capping role, at least in yeast, and that they may positively affect telomere maintenance by either the enzyme telomerase or by recombination-based mechanisms. On the other hand, G-quadruplex formation in telomeric DNA, as elsewhere in the genome, can form an impediment to DNA replication and a source of genome instability. This review summarizes recent evidence for the in vivo existence of G-quadruplexes at telomeres, with a focus on human telomeres, and highlights some of the many unanswered questions regarding the location, form, and functions of these structures.

scholarly journals Fission Yeast Taz1 and RPA Are Synergistically Required to Prevent Rapid Telomere Loss

2007 ◽  
Vol 18 (6) ◽  
pp. 2378-2387 ◽  
Author(s):  
Tatsuya Kibe ◽  
Yuuki Ono ◽  
Koichiro Sato ◽  
Masaru Ueno
Keyword(s):  
Fission Yeast ◽  
Binding Protein ◽  
Protein A ◽  
Telomere Maintenance ◽  
Helicase Domain ◽  
Recq Helicase ◽  
Telomeric Dna ◽  
Telomere Binding Protein ◽  
Recombination Repair ◽  
Telomere Loss

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.

scholarly journals Genetic Inactivation ofATRXLeads to a Decrease in the Amount of Telomeric Cohesin and Level of Telomere Transcription in Human Glioma Cells

2015 ◽  
Vol 35 (16) ◽  
pp. 2818-2830 ◽  
Author(s):  
Rita Eid ◽  
Marie-Véronique Demattei ◽  
Harikleia Episkopou ◽  
Corinne Augé-Gouillou ◽  
Anabelle Decottignies ◽  
...  
Keyword(s):  
Rna Polymerase Ii ◽  
Chromatin Immunoprecipitation ◽  
Glioma Cells ◽  
Telomere Maintenance ◽  
Telomeric Dna ◽  
Hairpin Rna ◽  
Genetic Inactivation ◽  
Alternative Lengthening ◽  
Rnap Ii ◽  
Subtelomeric Regions

Mutations in ATRX (alphathalassemia/mentalretardation syndromeX-linked), a chromatin-remodeling protein, are associated with the telomerase-independent ALT (alternative lengthening of telomeres) pathway of telomere maintenance in several types of cancer, including human gliomas. In telomerase-positive glioma cells, we found by immunofluorescence that ATRX localized not far from the chromosome ends but not exactly at the telomere termini. Chromatin immunoprecipitation (ChIP) experiments confirmed a subtelomeric localization for ATRX, yet short hairpin RNA (shRNA)-mediated genetic inactivation ofATRXfailed to trigger the ALT pathway. Cohesin has been recently shown to be part of telomeric chromatin. Here, using ChIP, we showed that genetic inactivation ofATRXprovoked diminution in the amount of cohesin in subtelomeric regions of telomerase-positive glioma cells. Inactivation ofATRXalso led to diminution in the amount of TERRAs, noncoding RNAs resulting from transcription of telomeric DNA, as well as to a decrease in RNA polymerase II (RNAP II) levels at the telomeres. Our data suggest that ATRX might establish functional interactions with cohesin on telomeric chromatin in order to control TERRA levels and that one or the other or both of these events might be relevant to the triggering of the ALT pathway in cancer cells that exhibit genetic inactivation ofATRX.

scholarly journals Recombination-Based Telomere Maintenance Is Dependent on Tel1-MRN and Rap1 and Inhibited by Telomerase, Taz1, and Ku in Fission Yeast

2007 ◽  
Vol 28 (5) ◽  
pp. 1443-1455 ◽  
Author(s):  
Lakxmi Subramanian ◽  
Bettina A. Moser ◽  
Toru M. Nakamura
Keyword(s):  
Dna Repair ◽  
Fission Yeast ◽  
Catalytic Subunit ◽  
Telomere Maintenance ◽  
Yeast Cells ◽  
Eukaryotic Cells ◽  
Checkpoint Kinase ◽  
Telomere Binding Protein ◽  
Evolutionarily Conserved ◽  
Linear Chromosomes

ABSTRACT Fission yeast cells survive loss of the telomerase catalytic subunit Trt1 (TERT) through recombination-based telomere maintenance or through chromosome circularization. Although trt1Δ survivors with linear chromosomes can be obtained, they often spontaneously circularize their chromosomes. Therefore, it was difficult to establish genetic requirements for telomerase-independent telomere maintenance. In contrast, when the telomere-binding protein Taz1 is also deleted, taz1Δ trt1Δ cells are able to stably maintain telomeres. Thus, taz1Δ trt1Δ cells can serve as a valuable tool in understanding the regulation of telomerase-independent telomere maintenance. In this study, we show that the checkpoint kinase Tel1 (ATM) and the DNA repair complex Rad32-Rad50-Nbs1 (MRN) are required for telomere maintenance in taz1Δ trt1Δ cells. Surprisingly, Rap1 is also essential for telomere maintenance in taz1Δ trt1Δ cells, even though recruitment of Rap1 to telomeres depends on Taz1. Expression of catalytically inactive Trt1 can efficiently inhibit recombination-based telomere maintenance, but the inhibition requires both Est1 and Ku70. While Est1 is essential for recruitment of Trt1 to telomeres, Ku70 is dispensable. Thus, we conclude that Taz1, TERT-Est1, and Ku70-Ku80 prevent telomere recombination, whereas MRN-Tel1 and Rap1 promote recombination-based telomere maintenance. Evolutionarily conserved proteins in higher eukaryotic cells might similarly contribute to telomere recombination.

scholarly journals Telomere damage induces internal loops that generate telomeric circles

2020 ◽  
Author(s):  
Giulia Mazzucco ◽  
Armela Huda ◽  
Martina Galli ◽  
Daniele Piccini ◽  
Michele Giannattasio ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Tumor Cells ◽  
Telomere Maintenance ◽  
Telomeric Repeats ◽  
Single Stranded Dna ◽  
Alternative Lengthening ◽  
Electron Microscopy Analysis ◽  
Microscopy Analysis ◽  
Internal Loops ◽  
Telomere Erosion

AbstractExtrachromosomal telomeric circles are commonly invoked as important players in telomere maintenance, but their origin has remained elusive. Using electron microscopy analysis on purified telomeres we show that, apart from known structures, telomeric repeats accumulate internal loops (i-loops), that occur in proximity of nicks and single-stranded DNA gaps. I-loops are induced by single-stranded damage at normal telomeres and represent the majority of telomeric structures detected in ALT (Alternative Lengthening of Telomeres) tumor cells. Our data indicate that i-loops form as a consequence of the exposure of single-stranded DNA at telomeric repeats. Finally, we show that these damage-induced i-loops can be excised to generate extrachromosomal telomeric circles resulting in loss of telomeric repeats. Our results identify damage-induced i-loops as a new intermediate in telomere metabolism and reveal a simple mechanism that links telomere damage with the accumulation of extrachromosomal telomeric circles and telomere erosion.

scholarly journals Genetic Control of Telomere Integrity in Schizosaccharomyces pombe: rad3+ and tel1+ Are Parts of Two Regulatory Networks Independent of the Downstream Protein Kinases chk1+ and cds1+

Genetics ◽  
1999 ◽  
Vol 152 (4) ◽  
pp. 1501-1512 ◽  
Author(s):  
Akira Matsuura ◽  
Taku Naito ◽  
Fuyuki Ishikawa
Keyword(s):  
Schizosaccharomyces Pombe ◽  
Protein Kinases ◽  
Regulatory Networks ◽  
Telomere Maintenance ◽  
Telomeric Dna ◽  
Telomere Shortening ◽  
Telomere Repeat ◽  
Homologous Protein ◽  
Drastic Effect

Abstract The Schizosaccharomyces pombe checkpoint gene named rad3+ encodes an ATM-homologous protein kinase that shares a highly conserved motif with proteins involved in DNA metabolism. Previous studies have shown that Rad3 fulfills its function via the regulation of the Chk1 and Cds1 protein kinases. Here we describe a novel role for Rad3 in the control of telomere integrity. Mutations in the rad3+ gene alleviated telomeric silencing and produced shortened lengths in the telomere repeat tracts. Genetic analysis revealed that the other checkpoint rad mutations rad1, rad17, and rad26 belong to the same phenotypic class with rad3 with regard to control of the telomere length. Of these mutations, rad3 and rad26 have a drastic effect on telomere shortening. tel1+, another ATM homologue in S. pombe, carries out its telomere maintenance function in parallel with the checkpoint rad genes. Furthermore, either a single or double disruption of cds1+ and chk1+ caused no obvious changes in the telomeric DNA structure. Our results demonstrate a novel role of the S. pombe ATM homologues that is independent of chk1+ and cds1+.

scholarly journals TERRA G-quadruplex RNA interaction with TRF2 GAR domain is required for telomere integrity

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Yang Mei ◽  
Zhong Deng ◽  
Olga Vladimirova ◽  
Nitish Gulve ◽  
F. Brad Johnson ◽  
...  
Keyword(s):  
Telomeric Repeat ◽  
Telomere Maintenance ◽  
Structural Basis ◽  
Telomeric Dna ◽  
Recognition Element ◽  
Age Related ◽  
Binding Factor ◽  
G Quadruplex ◽  
Genetic Deletion ◽  
Rna Interaction

AbstractTelomere dysfunction causes chromosomal instability which is associated with many cancers and age-related diseases. The non-coding telomeric repeat-containing RNA (TERRA) forms a structural and regulatory component of the telomere that is implicated in telomere maintenance and chromosomal end protection. The basic N-terminal Gly/Arg-rich (GAR) domain of telomeric repeat-binding factor 2 (TRF2) can bind TERRA but the structural basis and significance of this interaction remains poorly understood. Here, we show that TRF2 GAR recognizes G-quadruplex features of TERRA. We show that small molecules that disrupt the TERRA-TRF2 GAR complex, such as N-methyl mesoporphyrin IX (NMM) or genetic deletion of TRF2 GAR domain, result in the loss of TERRA, and the induction of γH2AX-associated telomeric DNA damage associated with decreased telomere length, and increased telomere aberrations, including telomere fragility. Taken together, our data indicates that the G-quadruplex structure of TERRA is an important recognition element for TRF2 GAR domain and this interaction between TRF2 GAR and TERRA is essential to maintain telomere stability.

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