scholarly journals Selective Elimination of Osteosarcoma Cell Lines with Short Telomeres by ATR Inhibitors

2020 ◽  
Author(s):  
Tomas Goncalves ◽  
Georgia Zoumpoulidou ◽  
Carlos Alvarez-Mendoza ◽  
Caterina Mancusi ◽  
Laura C. Collopy ◽  
...  
Keyword(s):  
Telomere Length ◽  
Cell Lines ◽  
Replicative Senescence ◽  
Telomere Maintenance ◽  
Osteosarcoma Cell ◽  
Cancer Evolution ◽  
Induced Apoptosis ◽  
Chromosome Bridges ◽  
Atr Kinase ◽  
Telomere Lengthening

AbstractTo avoid replicative senescence or telomere-induced apoptosis, cancers employ telomere maintenance mechanisms (TMMs) involving either the upregulation of telomerase or the acquisition of recombination-based alternative telomere lengthening (ALT). The choice of TMM may differentially influence cancer evolution and be exploitable in targeted therapies. Here, we examine TMMs in a panel of seventeen osteosarcoma-derived cell lines defining three separate groups according to TMM. Eight were ALT-positive, including the previously uncharacterised lines, KPD and LM7. ALT-negative cell lines were further classified into two groups according to their telomere length. HOS-MNNG, OHSN, SJSA-1, HAL, 143b and HOS displayed sub-normally short telomere length, while MG-63, MHM and HuO-3N1 displayed long telomeres. Importantly, sub-normally short telomeres were significantly associated with hypersensitivity to three different therapeutics targeting the ataxia telangiectasia and Rad3-related (ATR) kinase - AZD-6738/Ceralasertib, VE-822/Berzoserib and BAY-1895344 - compared to long telomeres, maintained via ALT or telomerase. Within 24 hours of ATR inhibition, cells with short but not long telomeres displayed chromosome bridges and underwent cell death, indicating a selective dependency on ATR for chromosome stability. Collectively, our work provides a resource to identify links between TMMs and drug sensitivity in osteosarcoma and indicates that telomere length predicts ATR-inhibitor sensitivity in cancer.

scholarly journals RAD50 and RAD51 Define Two Pathways That Collaborate to Maintain Telomeres in the Absence of Telomerase

Genetics ◽  
1999 ◽  
Vol 152 (1) ◽  
pp. 143-152 ◽  
Author(s):  
Siyuan Le ◽  
J Kent Moore ◽  
James E Haber ◽  
Carol W Greider
Keyword(s):  
Cell Death ◽  
Telomere Length ◽  
Gene Conversion ◽  
De Novo ◽  
Dna Recombination ◽  
Telomere Maintenance ◽  
Triple Mutant ◽  
Yeast Telomerase ◽  
Telomere Lengthening

Abstract Telomere length is maintained by the de novo addition of telomere repeats by telomerase, yet recombination can elongate telomeres in the absence of telomerase. When the yeast telomerase RNA component, TLC1, is deleted, telomeres shorten and most cells die. However, gene conversion mediated by the RAD52 pathway allows telomere lengthening in rare survivor cells. To further investigate the role of recombination in telomere maintenance, we assayed telomere length and the ability to generate survivors in several isogenic DNA recombination mutants, including rad50, rad51, rad52, rad54, rad57, xrs2, and mre11. The rad51, rad52, rad54, and rad57 mutations increased the rate of cell death in the absence of TLC1. In contrast, although the rad50, xrs2, and mre11 strains initially had short telomeres, double mutants with tlc1 did not affect the rate of cell death, and survivors were generated at later times than tlc1 alone. While none of the double mutants of recombination genes and tlc1 (except rad52 tlc1) blocked the ability to generate survivors, a rad50 rad51 tlc1 triple mutant did not allow the generation of survivors. Thus RAD50 and RAD51 define two separate pathways that collaborate to allow cells to survive in the absence of telomerase.

scholarly journals Rhotekin 2 silencing inhibits proliferation and induces apoptosis in human osteosarcoma cells

2018 ◽  
Vol 38 (6) ◽  
Author(s):  
Xiong Wang ◽  
Lei Zhang ◽  
Wenji Wang ◽  
Yuchen Wang ◽  
Ye Chen ◽  
...  
Keyword(s):  
Cell Lines ◽  
Phase Cell ◽  
Osteosarcoma Cell ◽  
Cyclin Dependent Kinase ◽  
Potential Therapeutic Target ◽  
Human Osteosarcoma ◽  
Cycle Arrest ◽  
Human Osteosarcoma Cells ◽  
Human Osteosarcoma Cell ◽  
Induced Apoptosis

Human osteosarcoma is the most frequent primary malignant of bone, and often occurs in adolescents. However, molecular mechanism of this disease remains unclear. In the present study, we found that the level of Rhotekin 2 (RTKN2) was up-regulated in osteosarcoma tissues and cell lines. In addition, silencing of RTKN2 of human osteosarcoma cell lines U2OS, inhibited proliferation, and induced G1 phase cell cycle arrest via reducing the level of the cyclin-dependent kinase 2 (CDK2). Furthermore, RTKN2 knockdown in the U2OS cells induced apoptosis by increasing the level of Bax and decreasing the level of Bcl2. These results suggested that RTKN2 is involved in the progression of human osteosarcoma, and may be a potential therapeutic target.

scholarly journals Differential Regulation of Telomeric Complex by BCR-ABL1 Kinase in Human Cellular Models of Chronic Myeloid Leukemia—From Single Cell Analysis to Next-Generation Sequencing

Genes ◽  
2020 ◽  
Vol 11 (10) ◽  
pp. 1145
Author(s):  
Anna Deregowska ◽  
Monika Pepek ◽  
Katarzyna Pruszczyk ◽  
Marcin M. Machnicki ◽  
Maciej Wnuk ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Chronic Myeloid Leukemia ◽  
Telomere Length ◽  
Cell Lines ◽  
Copy Number ◽  
Myeloid Leukemia ◽  
Telomere Maintenance ◽  
Next Generation ◽  
Cellular Models ◽  
Generation Sequencing

Telomeres are specialized nucleoprotein complexes, localized at the physical ends of chromosomes, that contribute to the maintenance of genome stability. One of the features of chronic myeloid leukemia (CML) cells is a reduction in telomere length which may result in increased genomic instability and progression of the disease. Aberrant telomere maintenance in CML is not fully understood and other mechanisms such as the alternative lengthening of telomeres (ALT) are involved. In this work, we employed five BCR-ABL1-positive cell lines, namely K562, KU-812, LAMA-84, MEG-A2, and MOLM-1, commonly used in the laboratories to study the link between mutation, copy number, and expression of telomere maintenance genes with the expression, copy number, and activity of BCR-ABL1. Our results demonstrated that the copy number and expression of BCR-ABL1 are crucial for telomere lengthening. We observed a correlation between BCR-ABL1 expression and telomere length as well as shelterins upregulation. Next-generation sequencing revealed pathogenic variants and copy number alterations in major tumor suppressors, such as TP53 and CDKN2A, but not in telomere-associated genes. Taken together, we showed that BCR-ABL1 kinase expression and activity play a crucial role in the maintenance of telomeres in CML cell lines. Our results may help to validate and properly interpret results obtained by many laboratories employing these in vitro models of CML.

scholarly journals The Rap1p-Telomere Complex Does Not Determine the Replicative Capacity of Telomerase-Deficient Yeast

2003 ◽  
Vol 23 (23) ◽  
pp. 8729-8739 ◽  
Author(s):  
Sarit Smolikov ◽  
Anat Krauskopf
Keyword(s):  
Telomere Length ◽  
Replicative Senescence ◽  
Budding Yeast ◽  
Kluyveromyces Lactis ◽  
Dna Binding Protein ◽  
Replicative Capacity ◽  
Telomeric Dna ◽  
Telomeric Repeats ◽  
Telomere Lengthening

ABSTRACT Telomeres are nucleoprotein structures that cap the ends of chromosomes and thereby protect their stability and integrity. In the presence of telomerase, the enzyme that synthesizes telomeric repeats, telomere length is controlled primarily by Rap1p, the budding yeast telomeric DNA binding protein which, through its C-terminal domain, nucleates a protein complex that limits telomere lengthening. In the absence of telomerase, telomeres shorten with every cell division, and eventually, cells enter replicative senescence. We have set out to identify the telomeric property that determines the replicative capacity of telomerase-deficient budding yeast. We show that in cells deficient for both telomerase and homologous recombination, replicative capacity is dependent on telomere length but not on the binding of Rap1p to the telomeric repeats. Strikingly, inhibition of Rap1p binding or truncation of the C-terminal tail of Rap1p in Kluyveromyces lactis and deletion of the Rap1p-recruited complex in Saccharomyces cerevisiae lead to a dramatic increase in replicative capacity. The study of the role of telomere binding proteins and telomere length on replicative capacity in yeast may have significant implications for our understanding of cellular senescence in higher organisms.

scholarly journals The Function of DNA Polymerase α at Telomeric G Tails Is Important for Telomere Homeostasis

2000 ◽  
Vol 20 (3) ◽  
pp. 786-796 ◽  
Author(s):  
Aegina Adams Martin ◽  
Isabelle Dionne ◽  
Raymund J. Wellinger ◽  
Connie Holm
Keyword(s):  
Telomere Length ◽  
Dna Polymerase ◽  
Telomere Maintenance ◽  
Replication Machinery ◽  
Telomeric Dna ◽  
Dna Polymerase Α ◽  
Telomeric Silencing ◽  
Pass Through ◽  
Telomere Lengthening ◽  
Lagging Strand

ABSTRACT Telomere length control is influenced by several factors, including telomerase, the components of telomeric chromatin structure, and the conventional replication machinery. Although known components of the replication machinery can influence telomere length equilibrium, little is known about why mutations in certain replication proteins cause dramatic telomere lengthening. To investigate the cause of telomere elongation in cdc17/pol1 (DNA polymerase α) mutants, we examined telomeric chromatin, as measured by its ability to repress transcription on telomere-proximal genes, and telomeric DNA end structures in pol1-17 mutants. pol1-17 mutants with elongated telomeres show a dramatic loss of the repression of telomere-proximal genes, or telomeric silencing. In addition,cdc17/pol1 mutants grown under telomere-elongating conditions exhibit significant increases in single-stranded character in telomeric DNA but not at internal sequences. The single strandedness is manifested as a terminal extension of the G-rich strand (G tails) that can occur independently of telomerase, suggesting thatcdc17/pol1 mutants exhibit defects in telomeric lagging-strand synthesis. Interestingly, the loss of telomeric silencing and the increase in the sizes of the G tails at the telomeres temporally coincide and occur before any detectable telomere lengthening is observed. Moreover, the G tails observed incdc17/pol1 mutants incubated at the semipermissive temperature appear only when the cells pass through S phase and are processed by the time cells reach G1. These results suggest that lagging-strand synthesis is coordinated with telomerase-mediated telomere maintenance to ensure proper telomere length control.

scholarly journals Evaluations of short telomere risk-associated single nucleotide polymorphisms in telomerase reverse transcriptase gene on telomere length maintenance

2019 ◽  
Author(s):  
Jialin Xu ◽  
Matthew A. Trudeau ◽  
Andrew J. Sandford ◽  
Judy M.Y. Wong
Keyword(s):  
Single Nucleotide Polymorphisms ◽  
Reverse Transcriptase ◽  
Telomere Length ◽  
Replicative Senescence ◽  
Telomere Maintenance ◽  
Telomerase Reverse Transcriptase ◽  
Nucleotide Polymorphisms ◽  
Primary Cells ◽  
Single Nucleotide ◽  
Telomere Biology

ABSTRACTTelomere biology disorders (TBDs) refer to a spectrum of tissue degenerative disorders caused by genetic mutations in telomere biology genes. Most patients with TBDs suffer from telomere maintenance defects secondary to telomerase deficiency. While the highly penetrant mutations in the telomerase reverse transcriptase (TERT) gene that drive disease onset and progression of TBDs are relatively rare, there exist several single nucleotide polymorphisms (SNPs) in TERT that have been linked to various diseases in the TBD spectrum. In this study, we investigated the biochemical properties of five TERT variants. In an ex vivo cell model, we found that primary human fibroblasts expressing nonsynonymous TERT SNPs had comparable cell growth kinetics to primary cells expressing WT-TERT, while a parallel vector control expressing-cell line entered replicative senescence. At the molecular level, primary cells expressing the minor alleles of two of the five TERT variants (A279T, ΔE441) had replication-dependent loss of telomere length. In an in vitro primer extension assay, these two variants showed reduced telomerase nucleotide addition processivity. Together, our data suggested that selective, common TERT variants could be revealed to harbour telomere maintenance defects, leading to a plausible explanation for their observed associations to telomere biology disorders.

The flow cytometric analysis of telomere length in antigen-specific CD8+ T cells during acute Epstein-Barr virus infection

Blood ◽  
2001 ◽  
Vol 97 (3) ◽  
pp. 700-707 ◽  
Author(s):  
Fiona J. Plunkett ◽  
Maria Vieira D. Soares ◽  
Nicola Annels ◽  
Andrew Hislop ◽  
Kamal Ivory ◽  
...  
Keyword(s):  
T Cells ◽  
Telomere Length ◽  
Cd8 T Cells ◽  
Epstein Barr Virus ◽  
Replicative Senescence ◽  
Telomere Maintenance ◽  
Epstein Barr Virus Infection ◽  
Telomere Shortening ◽  
Barr Virus ◽  
Epstein Barr

Abstract Acute infectious mononucleosis (AIM) induced by Epstein-Barr virus (EBV) infection is characterized by extensive expansion of antigen-specific CD8+ T cells. One potential consequence of this considerable proliferative activity is telomere shortening, which predisposes the EBV-specific cells to replicative senescence. To investigate this, a method was developed that enables the simultaneous identification of EBV specificity of the CD8+ T cells, using major histocompatibility complex (MHC) class I/peptide complexes, together with telomere length, which is determined by fluorescence in situ hybridization. Despite the considerable expansion, CD8+ EBV-specific T cells in patients with AIM maintain their telomere length relative to CD8+ T cells in normal individuals and relative to CD4+ T cells within the patients themselves and this is associated with the induction of the enzyme telomerase. In 4 patients who were studied up to 12 months after resolution of AIM, telomere lengths of EBV-specific CD8+ T cells were unchanged in 3 but shortened in one individual, who was studied only 5 months after initial onset of infection. Substantial telomere shortening in EBV-specific CD8+ T cells was observed in 3 patients who were studied between 15 months and 14 years after recovery from AIM. Thus, although telomerase activation may preserve the replicative potential of EBV-specific cells in AIM and after initial stages of disease resolution, the capacity of these cells to up-regulate this enzyme after restimulation by the persisting virus may dictate the extent of telomere maintenance in the memory CD8+ T-cell pool over time.

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