scholarly journals Closed chromatin loops at the ends of chromosomes

2004 ◽  
Vol 166 (2) ◽  
pp. 161-165 ◽  
Author(s):  
Tatiana Nikitina ◽  
Christopher L. Woodcock
Keyword(s):  
Electron Microscopy ◽  
Telomere Length ◽  
Telomerase Activity ◽  
Chromatin Conformation ◽  
Chicken Erythrocyte ◽  
Telomere Shortening ◽  
Mouse Lymphocyte ◽  
Associated Proteins ◽  
Protective Structures ◽  
Telomere Chromatin

The termini of eukaryotic chromosomes contain specialized protective structures, the telomeres, composed of TTAGGG repeats and associated proteins which, together with telomerase, control telomere length. Telomere shortening is associated with senescence and inappropriate telomerase activity may lead to cancer. Little is known about the chromatin context of telomeres, because, in most cells, telomere chromatin is tightly anchored within the nucleus. We now report the successful release of telomere chromatin from chicken erythrocyte and mouse lymphocyte nuclei, both of which have a reduced karyoskeleton. Electron microscopy reveals telomere chromatin fibers in the form of closed terminal loops, which correspond to the “t-loop” structures adopted by telomere DNA. The ability to recognize isolated telomeres in their native chromatin conformation opens the way for detailed structural and compositional studies.

Extensive Chromosome Instability in Acute Myeloid Leukemia Is Associated with Critical Telomere Shortening.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 3376-3376
Author(s):  
Susan J.J. Swiggers ◽  
Marianne A. Kuijpers ◽  
Maartje J. de Cort ◽  
Berna Beverloo ◽  
J. Mark J.M. Zijlmans
Keyword(s):  
Telomere Length ◽  
Reciprocal Translocation ◽  
Genetic Instability ◽  
Myeloid Leukemia ◽  
Chromosomal Abnormalities ◽  
Chromosome Instability ◽  
Telomerase Activity ◽  
In Vitro Models ◽  
Telomere Shortening

Abstract Telomeres, the ends of linear chromosomes, have a critical role in protection against chromosome end-to-end fusion. Telomeres shorten in every cell division due to the end replication problem. Telomerase is a reverse transcriptase that adds telomeric DNA repeats to the ultimate chromosome end. In vitro models of long-term fibroblast cultures have identified two sequential mortality stages, senescence (M1) and crisis (M2). Senescence can be bypassed by loss of p53 or Rb function, whereas escape from crisis can only be achieved by activating a telomere maintenance mechanism, mostly telomerase reactivation. Cells that bypass senescence (M1) did not reactivate telomerase, resulting in further telomere shortening to a critical telomere length upon reaching crisis (M2). In these models, critical telomere shortening induces extensive chromosome instability, most likely via chromosome end-to-end fusions. Dicentric chromosomes lead to anaphase breakage-fusion-bridges resulting in multiple chromosomal aberrations. To investigate whether similar mechanisms may be involved in the development of genetic instability in human cancer, we studied telomere length and expression of critical telomeric proteins (TRF2 and POT1) in acute myeloid leukemia (AML) patients. AML is a good model for these studies since distinct subgroups of AML are characterized by either exchanges along chromosome arms (translocation or inversion), or by a complex karyotype with multiple chromosome aberrations. Groups were age-matched. Telomere length was studied in metaphase arrested leukemic cells using quantitative fluorescence in situ hybridization (Q-FISH) using a telomere-specific probe. Subsequently, metaphase spreads were hybridized with a leukemia-specific probe to confirm leukemic origin of each metaphase. Telomeres were significantly shorter in AML samples with multiple chromosomal abnormalities in comparison to AML samples with a reciprocal translocation/inversion or no abnormalities (mean±SEM=16±1.7 AFU, n=12 versus 29±4.3 AFU, n=18; p=0.015). Interestingly, telomerase activity level is significantly higher in AML samples with multiple chromosomal abnormalities, compared to AML samples with a reciprocal translocation or inversion (mean±SEM=330±95, n=11 versus 70±21, n=13; p=0.02). Expression levels of telomeric proteins TRF2 and POT1 were similar in these AML groups. Our observations suggest that, consistent with previous in vitro models in fibroblasts, critical telomere shortening may have a role in the development of genetic instability in human AML. Critically short telomeres in association with high levels of telomerase activity suggest that AML cells with multiple chromosomal abnormalities have bypassed crisis (M2). The longer telomeres and low levels of telomerase activity in AML cells with a reciprocal translocation or inversion suggest that they originate from an earlier stage, preceding crisis. Consequently, telomere length modulation may have a role in cancer prevention.

Telomerase Activity and Telomere Length of CD4+CD25+ Regulatory T-Cells under Conditions of In Vitro and In Vivo Expansion.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 3857-3857
Author(s):  
Dominik G.F. Wolf ◽  
Anna M. Wolf ◽  
Christian Koppelstaetter ◽  
Holger F. Rumpold ◽  
Gert Mayer ◽  
...  
Keyword(s):  
T Cells ◽  
Regulatory T Cells ◽  
Telomere Length ◽  
Cancer Patients ◽  
Telomerase Activity ◽  
Telomere Maintenance ◽  
Healthy Human ◽  
Telomere Shortening

Abstract The expandability of CD4+CD25+ regulatory T-cells (Treg) has been shown in vitro and in vivo. Activation of telomerase activity is a prerequisite for clonal expansion and telomere maintenance in T-cells. There is currently no data available on the expression and function of telomerase in proliferating Treg. Analyses of telomere length by flow-FISH, real-time PCR and Southern blotting revealed that Treg isolated from healthy human volunteers have significantly shortened telomeres when compared to CD4+CD25− T-cells. However, telomere length is not further shortened in Treg isolated from the peripheral blood of cancer patients, despite the observation that the regulatory T-cell pool of these patients was significantly enlarged. To gain further insight into maintenance of telomere length of Treg, we induced in vitro proliferation of Treg by stimulation with anti-CD3 and IL-2. This led to a rapid increase of telomerase activity, as determined by PCR-ELISA. However, when we focused on the proliferating fraction of Treg using a sorting strategy based on the dilution of CFSE, we could show a significant telomere shortening in Treg with high proliferative and immmuno-suppressive capacity. Of note, proliferating CFSElow Treg are characterized by high telomerase activity, which however seems to be insufficient to avoid further telomere shortening under conditions of strong in vitro stimulation. In contrast, under conditions of in vivo expansion of Treg in cancer patients, the induction of telomerase activity is likely to compensate for further telomere erosion. These data might be of importance when considering the application of in vitro expanded Treg for the treatment of GvHD or autoimmune diseases, as telomere shortening might be associated with genomic instability.

Maintenance of Telomere Length in Peripheral Blood CD4+CD25+ Regulatory T-Cells of Cancer Patients Despite Active Proliferation.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 3309-3309
Author(s):  
Dominik Wolf ◽  
Holger Rumpold ◽  
Christian Koppelstaetter ◽  
Guenther Gastl ◽  
Eberhard Gunsilius ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Regulatory T Cells ◽  
Telomere Length ◽  
Cancer Patients ◽  
Peripheral Blood ◽  
Telomerase Activity ◽  
Telomere Shortening ◽  
Active Proliferation

Abstract CD4+CD25+ regulatory T-cells (Treg) are increased in the peripheral blood of cancer patients. It remains unclear whether this is due to redistribution or active proliferation. The latter would require the up-regulation of telomerase activity, whose regulation also remains unknown for Treg. We therefore isolated Treg and the respective CD4+CD25− control T-cell population from peripheral blood of cancer patients (n=23) and healthy age-matched controls (n=17). Analysis of their content of T-cell receptor excision circles (TREC) revealed that the observed increase of Treg frequencies in peripheral blood is due to active cycling rather than to redistribution from other compartments (i.e. secondary lymphoid organs or bone-marrow), as Treg from cancer patients are characterized by a significant decrease of TREC content when compared to TREC content of Treg isolated from healthy age-matched controls. Surprisingly, despite their proven in vivo proliferation, telomere length is not further shortened in Treg from peripheral blood of cancer patients as shown by Flow-Fish, Real-Time PCR and Southern Blotting. Accodingly, telomerase activity of Treg was readily inducible in vitro by OKT3 together with IL-2. Notably, sorting of in vitro proliferating Treg using dilution of CFSE revealed a significant telomere shortening in Treg with high proliferative capacity (i.e. CFSElow fraction) under conditions of strong in vitro stimulatory growth conditions despite a high telomerase activity. Thus, under conditions of strong in vitro stimulation induction of telomerase seems to be insufficient to avoid progressive telomere shortening. In contrast, in actively proliferating peripheral blood Treg from patients with epithelial malignancies induction of telomerase activity is likely to compensate for further telomere erosion.

Functional p53 Is Required for Effective Telomerase Inhibition in BCRABL- Positive CML Cells in Vitro

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 572-572
Author(s):  
Ute Brassat ◽  
Stefan Balabanov ◽  
Melanie Braig ◽  
Daniel Bali ◽  
Kerstin Borgmann ◽  
...  
Keyword(s):  
Telomere Length ◽  
Growth Kinetics ◽  
Blast Crisis ◽  
Telomerase Activity ◽  
Gfp Expression ◽  
Telomere Shortening ◽  
Telomerase Inhibition ◽  
Group A ◽  
Group B

Abstract Telomeres consist of repeat structures such as (TTAGGG)n in vertebrates and are localized at the end of chromosomes. Replication-dependent telomere shortening due to the end-replication problem can be counteracted by upregulation of an endogenous reverse transcriptase called telomerase. Increasing evidence suggests that critical telomere shortening results in genetic instability which may promote tumour evolution and telomerase activation during which critically short telomeres are stabilised and ongoing tumour growth is facilitated. In Chronic myeloid leukemia (CML) the high turnover of the malignant clone is driven by the oncogene BCR-ABL and leads to accelerated telomere shortening in chronic phase (CP) compared to telomere length in healthy individuals. Telomere shortening has been demonstrated to be correlated with disease stage, duration, prognosis and response to molecular targeted treatment. Despite of the accelerated telomere shortening observed, telomerase activity is increased in CP CML and further upregulated with progression of the disease to accelerated phase or blast crisis (AP/BC). To investigate the effect of telomerase inhibition on BCR-ABL-positive cells, we expressed a dominant-negative mutant of hTERT (vector pOS DNhTERT-IRES-GFP) in K562 cells. The cells were single sorted and clones in addition to bulk cultures were long term expanded in vitro. The expression of the transgene DNhTERT was monitored by the expression of GFP and function of DNhTERT was analyzed by measurement of telomere length (by flow-FISH) and telomerase activity (TRAP assay). Evaluation of these parameters showed the following patterns of growth kinetics and telomere biology in individual clones: Two clones lost telomere repeats and were transiently delayed in growth kinetics but eventually escaped from crisis without loss of GFP expression (indicated by a re-increase in telomere length and growth rate, group A) Three other clones lost GFP expression after initial and significant telomere reduction indicating loss of the transgene (group B). Finally, telomere length and growth kinetics of two remaining clones and of the bulk culture cells remained unaffected by expression of DN-hTERT (group C). Of note, none of the clones analyzed either died or entered cell cycle arrest. Further analyses of one clone of group A revealed impaired DNA damage response indicated by two fold increase in number of γH2AX foci in comparison to control cells. Moreover, the expression pattern of genes involved in DNA repair was significantly altered (Dual chip®). Network analysis of the altered genes using MetaCore® software confirmed p53 as a key regulator in signaling of DNA damage in these cells. CML blast crisis cell lines such as K562 are typically negative for functional p53 and p16INK4. Therefore, we went on and investigate if the presence of functional p53 is required for the induction of telomere-mediated apoptosis or senescence in BCR-ABL-positive cells. For this purpose, we restored p53 in telomerase-negative clones by using an inducible system (vector pBABE p53ERtam) in two clones from group A and group B. Induction of p53 in cells with critically short telomeres (telomere length 4–5 kb) lead to immediate induction of apoptosis while vector control cells continued to escape from crisis. These results suggest that the success of strategies aimed at telomerase inhibition in CML is dependent on the presence of functional p53 in BCR-ABL-positive cells which argues in favour of applying these strategies preferentially in CP as opposed to BC.

Feline chronic kidney disease is associated with shortened telomeres and increased cellular senescence

2013 ◽  
Vol 305 (3) ◽  
pp. F295-F303 ◽  
Author(s):  
Jessica M. Quimby ◽  
David G. Maranon ◽  
Christine L. R. Battaglia ◽  
Shannon M. McLeland ◽  
William T. Brock ◽  
...  
Keyword(s):  
Chronic Kidney Disease ◽  
Kidney Disease ◽  
Statistical Significance ◽  
Interventional Therapy ◽  
Telomerase Activity ◽  
Telomere Shortening ◽  
Amplification Protocol ◽  
Protective Structures ◽  
Kidney Liver

Telomeres are protective structures at the ends of chromosomes that have important implications for aging. To address the question of whether telomeres contribute to feline chronic kidney disease (CKD), we evaluated kidney, liver, and skin samples from 12 cats with naturally occurring CKD, 12 young normal cats, and 6 old normal cats. Telomere length was assessed using standard telomere fluorescent in situ hybridization (TEL-FISH) combined with immunohistochemistry (TELI-FISH) to identify proximal (PTEC) and distal tubular epithelial cells (DTEC), whereas senescence-associated β-galactosidase (SABG) staining was used to evaluate senescence. Results revealed statistically significant decreases in the average telomere fluorescence intensity (TFI) of PTEC in CKD cats compared with young and geriatric normal cats, and in the DTEC of CKD cats compared with young normal cats. When histograms of individual TFI were compared, statistically significant decreases in the PTEC and DTEC of CKD cats were observed compared with young and geriatric normal cats. Concomitantly, a statistically significant increase in SABG staining was seen in CKD kidney samples compared with young normal cats. CKD cats tended to have increased SABG staining in the kidney compared with normal geriatric cats, but this did not reach statistical significance. No significant telomere shortening in liver or skin from any group was observed. Real-time quantitative telomeric repeat amplification protocol assessment of renal telomerase activity revealed comparable low levels of telomerase activity in all groups. Our results suggest that shortened telomeres and increased senescence in the kidneys of CKD cats may represent novel targets for interventional therapy.

scholarly journals Functional Diversification of Replication Protein A Paralogs and Telomere Length Maintenance in Arabidopsis

Genetics ◽  
2020 ◽  
Vol 215 (4) ◽  
pp. 989-1002
Author(s):  
Behailu B. Aklilu ◽  
François Peurois ◽  
Carole Saintomé ◽  
Kevin M. Culligan ◽  
Daniela Kobbe ◽  
...  
Keyword(s):  
Homologous Recombination ◽  
Telomere Length ◽  
Protein A ◽  
Replication Protein A ◽  
Telomerase Activity ◽  
Replication Protein ◽  
Telomeric Dna ◽  
Telomere Shortening ◽  
Replication Group

Replication protein A (RPA) is essential for many facets of DNA metabolism. The RPA gene family expanded in Arabidopsis thaliana with five phylogenetically distinct RPA1 subunits (RPA1A-E), two RPA2 (RPA2A and B), and two RPA3 (RPA3A and B). RPA1 paralogs exhibit partial redundancy and functional specialization in DNA replication (RPA1B and RPA1D), repair (RPA1C and RPA1E), and meiotic recombination (RPA1A and RPA1C). Here, we show that RPA subunits also differentially impact telomere length set point. Loss of RPA1 resets bulk telomeres at a shorter length, with a functional hierarchy for replication group over repair and meiosis group RPA1 subunits. Plants lacking RPA2A, but not RPA2B, harbor short telomeres similar to the replication group. Telomere shortening does not correlate with decreased telomerase activity or deprotection of chromosome ends in rpa mutants. However, in vitro assays show that RPA1B2A3B unfolds telomeric G-quadruplexes known to inhibit replications fork progression. We also found that ATR deficiency can partially rescue short telomeres in rpa2a mutants, although plants exhibit defects in growth and development. Unexpectedly, the telomere shortening phenotype of rpa2a mutants is completely abolished in plants lacking the RTEL1 helicase. RTEL1 has been implicated in a variety of nucleic acid transactions, including suppression of homologous recombination. Thus, the lack of telomere shortening in rpa2a mutants upon RTEL1 deletion suggests that telomere replication defects incurred by loss of RPA may be bypassed by homologous recombination. Taken together, these findings provide new insight into how RPA cooperates with replication and recombination machinery to sustain telomeric DNA.

scholarly journals Chronic stress elevates telomerase activity in rats

2012 ◽  
Vol 8 (6) ◽  
pp. 1063-1066 ◽  
Author(s):  
Annaliese K. Beery ◽  
Jue Lin ◽  
Joshua S. Biddle ◽  
Darlene D. Francis ◽  
Elizabeth H. Blackburn ◽  
...  
Keyword(s):  
Chronic Stress ◽  
Dna Sequences ◽  
Replicative Senescence ◽  
Immune Cell ◽  
Telomerase Activity ◽  
Male Rats ◽  
Repetitive Dna Sequences ◽  
Potential Mechanism ◽  
Telomere Shortening ◽  
Protective Structures

The enzyme telomerase lengthens telomeres—protective structures containing repetitive DNA sequences at chromosome ends. Telomere shortening is associated with diseases of ageing in mammals. Chronic stress has been related to shorter immune-cell telomeres, but telomerase activity under stress may be low, permitting telomere loss, or high, partially attenuating it. We developed an experimental model to examine the impacts of extended unpredictable stress on telomerase activity in male rats. Telomerase activity was 54 per cent higher in stressed rats than in controls, and associated with stress-related physiological and behavioural outcomes. This significant increase suggests a potential mechanism for resilience to stress-related replicative senescence.

scholarly journals Influence of music therapy on the telomere length: a brief review and a pilot study

Biologija ◽  
2019 ◽  
Vol 64 (4) ◽  
Author(s):  
Irina M. Spivak ◽  
Tatiana Yu. Smirnova ◽  
Arina S. Urazova ◽  
Andrey L. Runov ◽  
Anastasia A. Vasilishina ◽  
...  
Keyword(s):  
Music Therapy ◽  
Telomere Length ◽  
Music Perception ◽  
Classical Music ◽  
Telomerase Activity ◽  
Adaptive Performance ◽  
Telomere Shortening ◽  
Age Related ◽  
Psychological Training ◽  
Before And After

Relation between music perception and telomere length is discussed. Telomeres are now considered as markers of general health and as possible predictor of life expectancy, while their length correlates with either the risk of age-related pathologies or with higher adaptive performance. The ability to withstand agerelated telomere shortening by means of practicing psychological training is discussed. Problems and prospects of present-day music therapy are reviewed, as well as absence of studies of alteration of telomere length, related to music perception. Sixty-three practically healthy young (aged 24 ± 5 years) randomly selected Russian-speaking students were divided into three subgroups, each of which listened to music of a specific type for 14 days, 90 minutes each day. Before and after the two-week course, the telomere length and telomerase activity in blood samples were measured. A limited but statistically reliable increase in the telomere length and in telomerase activity is demonstrated to have occurred as a result of completing the music course. Classical music tends to bring about less stress, positively affect one’s mood, and physiology, and, finally, such essential aging markers as telomerase activity and the telomere length. Non-classical music conditioned manifestation of some stress, which tended to affect the telomere length in a negative way. In order to cope with this difficulty, telomerase was activated.

Inhibition of Homologous Recombination Pathway Promotes Telomere Shortening and Cell Survival without Affecting Telomerase Activity In Myeloma

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 786-786
Author(s):  
Jagannath Pal ◽  
Jason Wong ◽  
Puru Nanjappa ◽  
Saem Lee ◽  
Masood Shammas ◽  
...  
Keyword(s):  
Homologous Recombination ◽  
Telomere Length ◽  
Board Of Directors ◽  
Telomerase Activity ◽  
Human Cells ◽  
Telomere Maintenance ◽  
Advisory Committees ◽  
Telomere Shortening ◽  
Telomerase Inhibitor ◽  
Normal Human

Abstract Abstract 786 Recombinase (RAD51) expression and homologous recombination (HR) activity are low in normal human cells including plasma cells. It is significantly induced following exposure of normal human cells to carcinogen, and is constitutively elevated in cancer cells including multiple myeloma (MM) cells. Besides its effect on genomic stability, elevated or dysregulated HR has also been implicated in telomere maintenance in tumor and immortalized cells. These cells usually lack telomerase activity and maintain telomere length by ALT mechanism (alternate lengthening of telomeres). Inhibitors of homologous recombination, therefore, have potential not only to prevent/reduce genomic instability, but also inhibit telomere maintenance, and cancer survival. We have here investigated the effect of inhibitor of HR on telomere maintenance mechanism in MM. We have evaluated effect of Nilotinib, a tyrosine kinase inhibitor and RAD51 shRNA on HR in MM. First we observed that nilotinib inhibits and RAD51 phosphorylation in MM. Nilotinib at both 5 and 10 mM concentration also led to dose-dependent inhibition of recombinase expression in MM cells. Importantly, Nilotinib also inhibited HR activity in MM cells as well as other cancer cell lines, as measured by a plasmid based assay in which leuciferase activity is generated following homologous recombination. We next evaluated effect of nilotinib on telomere maintenance alone as well as in combination with agents inhibiting telomere maintenance. The MM cells were treated for 48 hrs, either with nilotinib, telomerase inhibitor, or both nilotinib and telomerase inhibitor and evaluated for telomerase activity as well as effect on telomere length. As expected, the treatment of myeloma cells with telomerase inhibitor at 1 mM led to 88% inhibition of telomerase activity relative to control cells. Nilotinib, either alone or in the presence of telomerase inhibitor, did not have any major effect on telomerase activity in these cells. The cells were cultured in the presence of these agents for 2 weeks and evaluated for telomere length, using telomere specific real time PCR. Cells in presence of Telomerase inhibitor at 1 mM in fact had slightly increased telomere length (9%), probably due to presence or activation of ALT mechanism, following loss of telomerase activity. Importantly, nilotinib alone at 10 mM led to 20% reduction in telomere length and when combined with telomerease inhibitor at 1 mM concentrations led to reduction in the telomere length in MM cells by 52%. Moreover we have observed that transduction of MM cells with shRNA targeting RAD51 combined with telomerase inhibitor induced greater and quicker MM cell kill compared to either of these treatments alone. These data indicate that elevated HR pathway contributes to telomere maintenance in MM and combining inhibitors of HR with telomerase would expedite telomere shortening and cell death providing more effective therapeutic strategy. Disclosures: Munshi: Millennium Pharmaceuticals: Honoraria, Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; Onyx: Membership on an entity's Board of Directors or advisory committees.

scholarly journals Revisiting Telomere Shortening in Cancer

Cells ◽  
2019 ◽  
Vol 8 (2) ◽  
pp. 107 ◽  
Author(s):  
Keiji Okamoto ◽  
Hiroyuki Seimiya
Keyword(s):  
Cancer Cells ◽  
Telomere Length ◽  
Prognostic Biomarker ◽  
Telomere Maintenance ◽  
Epigenetic Alterations ◽  
Telomere Shortening ◽  
Normal Tissues ◽  
Genome Wide ◽  
Unlimited Growth ◽  
Protective Structures

Telomeres, the protective structures of chromosome ends are gradually shortened by each cell division, eventually leading to senescence or apoptosis. Cancer cells maintain the telomere length for unlimited growth by telomerase reactivation or a recombination-based mechanism. Recent genome-wide analyses have unveiled genetic and epigenetic alterations of the telomere maintenance machinery in cancer. While telomerase inhibition reveals that longer telomeres are more advantageous for cell survival, cancer cells often have paradoxically shorter telomeres compared with those found in the normal tissues. In this review, we summarize the latest knowledge about telomere length alterations in cancer and revisit its rationality. Finally, we discuss the potential utility of telomere length as a prognostic biomarker.

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