Alternative Lengthening of Telomeres Mediated by Mitotic DNA Synthesis Engages Break-Induced Replication Processes

ABSTRACT Alternative lengthening of telomeres (ALT) is a telomerase-independent telomere maintenance mechanism that occurs in a subset of cancers. By analyzing telomerase-positive cells and their human TERC knockout-derived ALT human cell lines, we show that ALT cells harbor more fragile telomeres representing telomere replication problems. ALT-associated replication defects trigger mitotic DNA synthesis (MiDAS) at telomeres in a RAD52-dependent, but RAD51-independent, manner. Telomeric MiDAS is a conservative DNA synthesis process, potentially mediated by break-induced replication, similar to type II ALT survivors in Saccharomyces cerevisiae. Replication stresses induced by ectopic oncogenic expression of cyclin E, G-quadruplexes, or R-loop formation facilitate the ALT pathway and lead to telomere clustering, a hallmark of ALT cancers. The TIMELESS/TIPIN complex suppresses telomere clustering and telomeric MiDAS, whereas the SMC5/6 complex promotes them. In summary, ALT cells exhibit more telomere replication defects that result in persistent DNA damage responses at telomeres, leading to the engagement of telomeric MiDAS (spontaneous mitotic telomere synthesis) that is triggered by DNA replication stress, a potential driver of genomic duplications in cancer.

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

Science Advances ◽

10.1126/sciadv.aax6366 ◽

2019 ◽

Vol 5 (10) ◽

pp. eaax6366 ◽

Cited By ~ 8

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.

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Oncogenic herpesvirus KSHV triggers hallmarks of alternative lengthening of telomeres

Nature Communications ◽

10.1038/s41467-020-20819-4 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Timothy P. Lippert ◽

Paulina Marzec ◽

Aurora I. Idilli ◽

Grzegorz Sarek ◽

Aleksandra Vancevska ◽

...

Keyword(s):

Cancer Cells ◽

Cell Lines ◽

Telomere Maintenance ◽

Alternative Lengthening Of Telomeres ◽

Telomere Shortening ◽

Alternative Lengthening ◽

A Cell ◽

Infected Cells ◽

Break Induced Replication ◽

Alt Activity

AbstractTo achieve replicative immortality, cancer cells must activate telomere maintenance mechanisms to prevent telomere shortening. ~85% of cancers circumvent telomeric attrition by re-expressing telomerase, while the remaining ~15% of cancers induce alternative lengthening of telomeres (ALT), which relies on break-induced replication (BIR) and telomere recombination. Although ALT tumours were first reported over 20 years ago, the mechanism of ALT induction remains unclear and no study to date has described a cell-based model that permits the induction of ALT. Here, we demonstrate that infection with Kaposi’s sarcoma herpesvirus (KSHV) induces sustained acquisition of ALT-like features in previously non-ALT cell lines. KSHV-infected cells acquire hallmarks of ALT activity that are also observed in KSHV-associated tumour biopsies. Down-regulating BIR impairs KSHV latency, suggesting that KSHV co-opts ALT for viral functionality. This study uncovers KSHV infection as a means to study telomere maintenance by ALT and reveals features of ALT in KSHV-associated tumours.

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Alternative lengthening of telomeres in childhood neuroblastoma from genome to proteome

Nature Communications ◽

10.1038/s41467-021-21247-8 ◽

2021 ◽

Vol 12 (1) ◽

Cited By ~ 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.

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Abstract 4767: Generating and characterizing novel prostate cancer cell lines that employ the alternative lengthening of telomeres (ALT) telomere maintenance mechanism

10.1158/1538-7445.am2016-4767 ◽

2016 ◽

Author(s):

Mindy K. Graham ◽

Jacqueline Brosnan-Cashman ◽

Anthony Rizzo ◽

Michael Haffner ◽

Alan Meeker ◽

...

Keyword(s):

Prostate Cancer ◽

Cell Lines ◽

Cancer Cell ◽

Prostate Cancer Cell ◽

Cancer Cell Lines ◽

Telomere Maintenance ◽

Alternative Lengthening Of Telomeres ◽

Prostate Cancer Cell Lines ◽

Maintenance Mechanism ◽

Alternative Lengthening

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Alternative lengthening of telomeres is the major telomere maintenance mechanism in astrocytoma with isocitrate dehydrogenase 1 mutation

Journal of Neuro-Oncology ◽

10.1007/s11060-020-03394-y ◽

2020 ◽

Vol 147 (1) ◽

pp. 1-14 ◽

Cited By ~ 2

Author(s):

Monica Sofia Ventura Ferreira ◽

Mia Dahl Sørensen ◽

Stefan Pusch ◽

Dagmar Beier ◽

Anne-Sophie Bouillon ◽

...

Keyword(s):

Isocitrate Dehydrogenase ◽

Telomere Maintenance ◽

Alternative Lengthening Of Telomeres ◽

Isocitrate Dehydrogenase 1 ◽

Maintenance Mechanism ◽

Alternative Lengthening

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Alternative Lengthening of Telomeres through Two Distinct Break-Induced Replication Pathways

Cell Reports ◽

10.1016/j.celrep.2018.12.102 ◽

2019 ◽

Vol 26 (4) ◽

pp. 955-968.e3 ◽

Cited By ~ 67

Author(s):

Jia-Min Zhang ◽

Tribhuwan Yadav ◽

Jian Ouyang ◽

Li Lan ◽

Lee Zou

Keyword(s):

Alternative Lengthening Of Telomeres ◽

Alternative Lengthening ◽

Break Induced Replication

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Evidence for Alternative Lengthening of Telomeres in Chronic Lymphocytic Leukemia Patients.

Blood ◽

10.1182/blood.v106.11.1179.1179 ◽

2005 ◽

Vol 106 (11) ◽

pp. 1179-1179

Author(s):

Rajendra N. Damle ◽

Taraneh Banapour ◽

Cristina Sison ◽

Steven L. Allen ◽

Kanti R. Rai ◽

...

Keyword(s):

Chronic Lymphocytic Leukemia ◽

Telomerase Activity ◽

Lymphocytic Leukemia ◽

Nuclear Bodies ◽

Telomere Maintenance ◽

Osteosarcoma Cell ◽

Clonal Deletion ◽

Alternative Lengthening Of Telomeres ◽

Alternative Lengthening ◽

V Genes

Abstract Telomere shortening is a consequence of repetitive clonal replication and leads to clonal deletion unless DNA extension and repair occur. All tumors must circumvent this problem by up-regulating mechanisms that lead to chromosomal lengthening. Two mechanisms have been identified that maintain chromosome ends- telomerase that does so by reverse transcription and alternative lengthening of telomeres (ALT) that occurs by homologous recombination. The latter function is characterized by the presence of promyelocytic leukemia protein-associated nuclear bodies (PML-NBs) and the presence of PML-NB is used to mark cells that use this process. B cell Chronic lymphocytic leukemia (B-CLL) cells with unmutated Ig V genes have shorter mean telomere lengths compared with those exhibiting mutated Ig V genes. In addition, cells with unmutated Ig V genes demonstrate more telomerase activity than their mutated counterparts. The mutated cases show long and heterogeneously elongated telomeres in spite of the absence, in most cases, of detectable telomerase activity. Therefore we determined whether the ALT pathway plays a role in telomere maintenance in B-CLL, using a monoclonal anti-PML antibody and a flow-cytometric assay for assessment of PML protein. Telomerase-expressing Jurkat T cells and murine fibroblasts-L cells served as negative controls for PML staining, whereas the ALT positive Osteosarcoma cell line U2-OS served as a positive control. In a cohort of 20 B-CLL cases, PML protein was detected in all cases regardless of Ig V mutation status. In addition, a similar percentage of cells within the clones contained PML (10 - 90% of the members of unmutated clones and 11–96% of mutated clones), whereas peripheral blood B cells from 6/6 elderly normal donors did not show any PML staining. PML expression was compared with telomere length and telomerase activity in the same cases. The percentage of cells showing PML expression inversely correlated with telomerase activity (r= −0.58; p=0.029). Although in most published reports telomere maintenance by ALT occurs in the absence of telomerase activity, we found ALT (as suggested by PML positive cells) in cells with telomerase activity (detected by the standard TRAP assay). Thus, B-CLL cases can express PML bodies and some B-CLL cells can contain both PML-NB and express telomerase activity. These findings suggest that B-CLL cells can use two distinct mechanisms to assure telomere maintenance and perpetuate clonal survival and expansion.

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Cancer cells with alternative lengthening of telomeres do not display a general hypersensitivity to ATR inhibition

10.1101/053280 ◽

2016 ◽

Author(s):

Katharina I. Deeg ◽

Inn Chung ◽

Caroline Bauer ◽

Karsten Rippe

Keyword(s):

Cell Viability ◽

Cancer Cells ◽

Cell Lines ◽

Ataxia Telangiectasia ◽

Telomere Maintenance ◽

Alternative Lengthening Of Telomeres ◽

Isogenic Cell Line ◽

Alternative Lengthening ◽

Cellular Immortality ◽

Proliferation Potential

AbstractTelomere maintenance is a hallmark of cancer as it provides cancer cells with cellular immortality. A significant fraction of tumors uses the alternative lengthening of telomeres (ALT) pathway to elongate their telomeres and to gain an unlimited proliferation potential. Since the ALT pathway is unique to cancer cells, it represents a potentially valuable, currently unexploited target for anticancer therapies. Recently, it was proposed that ALT renders cells hypersensitive to ataxia telangiectasia-and RAD3-related (ATR) protein inhibitors (Flynn et al., Science 347, 273). Here, we measured the response of various ALT or telomerase positive cell lines to the ATR inhibitor VE-821. In addition, we compared the effect of the inhibitor on cell viability in an isogenic cell line, in which ALT was active or suppressed. In these experiments a general ATR inhibitor sensitivity of cells with ALT could not be confirmed. We rather propose that the observed variations in sensitivity reflect differences between cell lines that are unrelated to ALT.

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TCGA Pan-Cancer genomic analysis of Alternative Lengthening of Telomeres (ALT) related genes

10.1101/2020.04.27.063610 ◽

2020 ◽

Author(s):

Isaac Armendáriz-Castillo ◽

Andrés López-Cortés ◽

Jennyfer García-Cárdenas ◽

Patricia Guevara-Ramírez ◽

Paola E. Leone ◽

...

Keyword(s):

Homologous Recombination ◽

Molecular Mechanisms ◽

Ex Vivo ◽

Genomic Analysis ◽

Interaction Network ◽

Telomere Maintenance ◽

The Cancer Genome Atlas ◽

Alternative Lengthening Of Telomeres ◽

Alternative Lengthening ◽

Pan Cancer

AbstractTelomere maintenance mechanisms (TMM) are used by cancer cells to avoid apoptosis, 85-90% reactivate telomerase, while 10-15% use the alternative lengthening of telomeres (ALT). Due to anti-telomerase-based treatments, some tumors have the ability to switch from a telomerase-dependent mechanism to ALT, in fact, the co-existence between telomerase and the ALT pathway have been observed in a variety of cancer types. Despite different elements in the ALT pathway have been uncovered, the molecular mechanism and other factors are still poorly understood, which difficult the detection and treatment of ALT-positive cells, which are known to present poor prognosis. Therefore, with the aim to identify potential molecular markers to be used in the study of ALT, we combined simplistic in silico approaches in 411 telomere maintenance (TM) genes which have been previously validated or predicted to be involved in the ALT pathway. In consequence, we conducted a genomic analysis of these genes in 31 Pan-Cancer Atlas studies (n=9,282) from The Cancer Genome Atlas in the cBioPortal and found 325,936 genomic alterations, being mRNA high and low the top alterations with 65,.8% and 10.7% respectively. Moreover, we analyzed the highest frequency means of genomics alterations, identified and proposed 20 genes, which are highly mutated and up and down regulated in the cancer studies and could be used for future analysis in the study of ALT. Finally, we made a protein-protein interaction network and enrichment analysis to obtain an insight into the main pathways these genes are involved. We could observe their role in main processes related to the ALT mechanism like homologous recombination, homology directed repair (HDR), HDR through homologous recombination and telomere maintenance and organization.. Overall, due to the lack of understanding of the molecular mechanisms and detection of ALT-positive cancers, we identified and proposed more molecular targets that can be used for expression analysis and additional ex vivo assays to validate them as new potential therapeutic markers in the study of the ALT mechanism.

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