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Genome-wide analysis showed that putative G-quadruplex DNA structures are prevalent in the human genome. The presence of G-quadruplex structure in the telomere and promoter region of certain oncogenes inspired people to use G-quadruplex ligand as anti-cancer agents. G-quadruplex structures, stabilized by ligand at telomere are resolved by telomerase making the cancer cells resistant to G-quadruplex ligand. So, identification of a new G-quadruplex ligand having anti-telomerase activity would be a promising strategy for cancer therapy as about 85% of human cancers are telomerase positive. A set of the drug-like compounds were screened from the ZINC database randomly and 2284 ligands were chosen following Lipinski’s rule of five that were docked with five different G-quadruplex DNA sequences in idock. We screened 43 potential G-quadruplex binders using Z-score as a normalization scoring function. The compound (ZINC ID-05220992) gave the best score (average idock = −10.17 kcal/mol, average normalized idock = −3.42). We performed G4 FID assay, CD analysis to understand its binding with three different G-quadruplex DNA sequences, and checked its anti-telomerase activity in A549 cells using TRAP assay. We observed that this compound had an intrinsic fluorescence, capability to stain live cells with a blue fluorescence, and a specific affinity to only 22AG out of three different G-quadruplex DNA sequences under study. It showed cytotoxicity, good permeability to live cells, and a significant reduction of telomerase activity in human A549 cells at a very low dose. So, this compound has strong potential to be an anti-cancer drug.Graphical AbstractHighlightsA set of compounds were screened randomly by a High throughput method and Lipinski’s rule of five from ZINC database to identify potential G4-binders.The compound (ZINC ID-05220992) was screened after docking with five G-quadruplex DNAIt binds G-quadruplex DNA 22AG as detected by TO displacement and CD spectroscopy.It inhibits telomerase activity in A549 cells and also cytotoxic to this cell.It penetrates live A549 cell in culture and stains it with blue fluorescence

Noncanonical DNA structures, termed G-quadruplexes, are present in human genomic DNA and are important elements in many DNA metabolic processes. Multiple sites in the human genome have G-rich DNA stretches able to support formation of several consecutive G-quadruplexes. One of those sites is the telomeric overhang region that has multiple repeats of TTAGGG and is tightly associated with both cancer and aging. We investigated the folding of consecutive G-quadruplexes in both potassium- and sodium-containing solutions using single-molecule FRET spectroscopy, circular dichroism, thermal melting and molecular dynamics simulations. Our observations show coexistence of partially and fully folded DNA, the latter consisting of consecutive G-quadruplexes. Following the folding process over hours in sodium-containing buffers revealed fast G-quadruplex folding but slow establishment of thermodynamic equilibrium. We find that full consecutive G-quadruplex formation is inhibited by the many DNA structures randomly nucleating on the DNA, some of which are off-path conformations that need to unfold to allow full folding. Our study allows describing consecutive G-quadruplex formation in both nonequilibrium and equilibrium conditions by a unified picture, where, due to the many possible DNA conformations, full folding with consecutive G-quadruplexes as beads on a string is not necessarily achieved.

Telomerase is a cellular enzyme that prevents telomere shortening and promoting viability. The literature has reported shortened telomere length in patients with major depressive disorder (MDD).35 patients with diagnosis of major depressive disorder (MDD) fulfilling DMS-5 criteria in the age range of 18-60 years, treatment-naïve after assessing the severity on HAM-D and HAM-A and 35 age and sex matched healthy controls were included in the study. Baseline peripheral blood monocytes (PBMC) telomerase enzyme was assessed in cases and controls and repeated in cases of MDD at 8th week after intervention with escitalopram for 8 weeks.Pretreatment telomerase activity (TA) was elevated in cases as compared to controls and it was also significantly correlated to the severity of depression (p = 0.00). There was a significant positive difference in telomerase activity between non-responders (higher TA) and responders at baseline (p = 0.001) and 8th week (p = 0.012). The TA did not vary significantly amidst pretreatment and post-treatment, although it was slightly lower in the post-treatment group.The study has few limitations in the form of small sample size, shorter duration of follow-up, and leucocyte telomeres length (LTL) was not assessed.The index study concludes that TA is higher in drug naïve patients with MDD than age and sex matched healthy control. The non-responders had significantly higher TA as compared to responders at baseline and post-treatment which indicates TA as a potential biomarker in the underlying biological mechanism of MDD and in response to antidepressant pharmacotherapy.

Background The pathogenesis of psoriasis vulgaris involves changes in DNA molecules, genomic instability, telomere attrition, and epigenetic alterations among them. These changes are also considered important mechanisms of aging in cells and tissues. Objective This study dealt with oxidation damage, telomere length and methylation status in DNA originating from peripheral blood of 41 psoriatic patients and 30 healthy controls. Methods Oxidative damage of serum DNA/RNA was determined immunochemically. Real-time PCR was used for the analysis of the telomere length. ELISA technique determined levels of 5-methylcytosine in blood cells' DNA. Results Oxidative damage of serum DNA/RNA was higher in patients than in controls (median, 3758 vs. 2286 pg/mL, p < 0.001). A higher length of telomeres per chromosome was found in patients whole-cell DNA than in controls (3.57 vs. 3.04 kilobases, p = 0.011). A negative correlation of the length of telomeres with an age of the control subjects was revealed (Spearman’s rho = -0.420, p = 0.028). Insignificantly different levels of 5-methylcytosine in patients and controls were observed (33.20 vs. 23.35%, p = 0.234). No influences of sex, smoking, BMI, PASI score, and metabolic syndrome on the methylation status were found. Study limitations i) A relatively small number of the participants, particularly for reliable subgroup analyses, ii) the Caucasian origin of the participants possibly influencing the results of the parameters determined, and iii) Telomerase activity was not directly measured in serum or blood cells. Conclusion The study demonstrated increased levels of oxidized DNA/RNA molecules in the serum of patients with exacerbated psoriasis vulgaris. The results were minimally influenced by sex, the presence of metabolic syndrome, or cigarette smoking. In the psoriatic blood cells' DNA, the authors observed longer telomeres compared to healthy controls, particularly in females. Insignificantly higher global DNA methylation in psoriasis cases compared to the controls indicated marginal clinical importance of this epigenetic test performed in the blood cells' DNA.

Numerous genetic discoveries and the advent of clinical telomere length testing have led to the recognition of a spectrum of telomere biology disorders (TBDs) beyond the classic dyskeratosis congenita (DC) triad of nail dysplasia, abnormal skin pigmentation, and oral leukoplakia occurring with pediatric bone marrow failure. Patients with DC/TBDs have very short telomeres for their age and are at high risk of bone marrow failure, cancer, pulmonary fibrosis (PF), pulmonary arteriovenous malformations, liver disease, stenosis of the urethra, esophagus, and/or lacrimal ducts, avascular necrosis of the hips and/or shoulders, and other medical problems. However, many patients with TBDs do not develop classic DC features; they may present in middle age and/or with just 1 feature, such as PF or aplastic anemia. TBD-associated clinical manifestations are progressive and attributed to aberrant telomere biology caused by the X-linked recessive, autosomal dominant, autosomal recessive, or de novo occurrence of pathogenic germline variants in at least 18 different genes. This review describes the genetics and clinical manifestations of TBDs and highlights areas in need of additional clinical and basic science research.

Previous studies have suggested a potential association between sleep and telomere length (TL), but its genetic basis remains unclear. In this study, we aimed to explore the genetic correlation and potential causal association between TL and insomnia.The genome-wide association study (GWAS) datasets of TL and insomnia-related traits were used, including insomnia, snoring, daytime dozing and napping. Based on the polygenic risk scores (PRS) of TL, linear regression and linkage disequilibrium score (LDSC) regression were used to preliminarily explore the association between TL and insomnia parameters in the UK Biobank cohort. Then, we investigated the causal association between TL and insomnia by mendelian randomization (MR) analysis and colocalization analysis.In the UK Biobank cohort, the association between TL and insomnia was observed in the female samples (t = 2.968, P = 3.00 × 10Our study found no evidence for causal association between insomnia and TL in individuals of European ancestry. We detected a candidate gene associated with both insomnia and TL, providing novel clues for understanding the roles of this association.

In brief Oxidative stress is recognized as an underlying driving factor of both telomere dysfunction and human subfertility/infertility. This review briefly reassesses telomere integrity as a fertility biomarker before proposing a novel, mechanistic rationale for the role of oxidative stress in the seemingly paradoxical lengthening of sperm telomeres with aging. Abstract The maintenance of redox balance in the male reproductive tract is critical to sperm health and function. Physiological levels of reactive oxygen species (ROS) promote sperm capacitation, while excess ROS exposure, or depleted antioxidant defenses, yields a state of oxidative stress which disrupts their fertilizing capacity and DNA structural integrity. The guanine moiety is the most readily oxidized of the four DNA bases and gets converted to the mutagenic lesion 8-hydroxy-deoxyguanosine (8-OHdG). Numerous studies have also confirmed oxidative stress as a driving factor behind accelerated telomere shortening and dysfunction. Although a clear consensus has not been reached, clinical studies also appear to associate telomere integrity with fertility outcomes in the assisted reproductive technology setting. Intriguingly, while sperm cellular and molecular characteristics make them more susceptible to oxidative insult than any other cell type, they are also the only cell type in which telomere lengthening accompanies aging. This article focuses on the oxidative stress response pathways to propose a mechanism for the explanation of this apparent paradox.

Telomeres have been advocated to be important markers of biological age in evolutionary and ecological studies. Telomeres usually shorten with age, and shortening is frequently associated with environmental stressors and increased subsequent mortality. Telomere lengthening – an apparent increase in telomere length between repeated samples from the same individual – also occurs. However, the exact circumstances, and consequences, of telomere lengthening are poorly understood. Using longitudinal data from the Seychelles warbler (Acrocephalus sechellensis), we tested whether telomere lengthening – which occurs in adults of this species – is associated with specific stressors (reproductive effort, food availability, malarial infection and cooperative breeding) and predicts subsequent survival. In females, telomere shortening was observed under greater stress (i.e. low food availability, malaria infection), while telomere lengthening was observed in females experiencing lower stress (i.e. high food availability, assisted by helpers, without malaria). The telomere dynamics of males were not associated with the key stressors tested. These results indicate that, at least for females, telomere lengthening occurs in circumstances more conducive to self-maintenance. Importantly, both females and males with lengthened telomeres had improved subsequent survival relative to individuals that displayed unchanged, or shortened, telomeres – indicating that telomere lengthening is associated with individual fitness. These results demonstrate that telomere dynamics are bidirectionally responsive to the level of stress that an individual faces, but may poorly reflect the accumulation of stress over the lifetime. This study challenges how we think of telomeres as a marker of biological age.

Omega-3 (n-3) and omega-6 (n-6) fatty acids may contribute to oxidative stress and inflammation, which are related to telomere shortening. Evidence supporting an association between intake of n-3 or n-6 fatty acids and leukocyte telomere length (LTL) in males has been limited.We conducted a cross-sectional study to examine the associations of total or individual n-3 or total n-6 fatty acid intake with LTL in US males.We included 2,494 US males with LTL measurement from 4 nested case-control studies within the Health Professionals Follow-Up Study. Individuals with previous histories of cancers, diabetes, and cardiovascular diseases at or before blood collection were excluded. Blood collection was performed between 1993 and 1995, and relevant information including n-3 and n-6 intake was collected in 1994 by questionnaire. The LTL was log-transformed and Z scores of the LTL were calculated for statistical analyses by standardizing the LTL in comparison with the mean within each selected nested case-control study.We found that consumption of DHA (22:6n-3) was positively associated with LTL. In the multivariable-adjusted model, compared with individuals who had the lowest intake of DHA (i.e., first quartile group), the percentage differences (95% CIs) of LTL were -3.7 (-13.7, 7.5), 7.0 (-4.3, 19.7), and 8.2 (-3.5, 21.3) for individuals in the second, third, and fourth quartiles of consumption, respectively (P-trend = 0.0498). We did not find significant associations between total n-3 or total n-6 fatty acid intakes and LTL. In addition, we found that males who consumed canned tuna had longer LTL than those who did not; in the multivariable-adjusted model, the percentage difference of LTL was 10.5 (95% CI: 1.3, 20.4) (P = 0.02).Our results suggest that higher intakes of DHA and canned tuna consumption are associated with longer LTL.

Telomeres are protective, nucleoprotein structures at the end of chromosomes that have been associated with lifespan across taxa. However, the extent to which these associations can be attributed to absolute length vs. the rate of telomere shortening prior to sampling remains unresolved. In a longitudinal study, we examined the relationship between lifespan, telomere length and the rate of telomere shortening in wild, purple-crowned fairy-wrens (Malurus coronatus coronatus). To this end, we measured telomere length using quantitative polymerase chain reaction in the blood of 59 individuals sampled as nestlings and 4-14 months thereafter, and in 141 known-age individuals sampled on average three times across adulthood. We applied within-subject centring analyses to simultaneously test for associations between lifespan and average telomere length and telomere shortening. We reveal that the rate of telomere shortening and to a lesser extent telomere length in the first year of life independently predicted lifespan, with individuals with faster shortening rates and/or shorter telomeres living less long. In contrast, in adulthood neither telomere shortening nor telomere length predicted lifespan, despite a considerably larger data set. Our results suggest that telomere length measured very early in life (during development) and longitudinal assessments of telomere shortening during the first year of life constitute more useful biomarkers of total life expectancy than either telomere length measured after development, or telomere shortening later in adulthood.

Individual variation in telomere length is predictive of health and mortality risk across a range of species. However, the relative influence of environmental and genetic variation on individual telomere length in wild populations remains poorly understood. Heritability of telomere length has primarily been calculated using parent-offspring regression which can be confounded by shared environments. To control for confounding variables, quantitative genetic "animal models" can be used, but few studies have applied animal models in wild populations. Furthermore, parental age at conception may also influence offspring telomere length, but most studies have been cross-sectional. We investigated within- and between-parental age at conception effects and heritability of telomere length in the Seychelles warbler using measures from birds caught over 20 years and a multigenerational pedigree. We found a weak negative within-paternal age at conception effect (as fathers aged, their offspring had shorter telomeres) and a weak positive between-maternal age at conception effect (females that survived to older ages had offspring with longer telomeres). Animal models provided evidence that heritability and evolvability of telomere length were low in this population, and that variation in telomere length was not driven by early-life effects of hatch period or parental identities. Quantitative polymerase chain reaction plate had a large influence on telomere length variation and not accounting for it in the models would have underestimated heritability. Our study illustrates the need to include and account for technical variation in order to accurately estimate heritability, as well as other environmental effects, on telomere length in natural populations.

Evidence that telomere length (TL) and dynamics can be interpreted as proxy for ‘life stress’ experienced by individuals stems largely from correlational studies. We tested for effects of an experimental increase of workload on telomere dynamics by equipping male great tits (Parus major) with a 0.9 gram backpack for a full year. In addition, we analysed associations between natural life-history variation, TL and TL dynamics. Carrying 5% extra weight for a year did not significantly accelerate telomere attrition. This agrees with our earlier finding that this experiment did not affect survival or future reproduction. Apparently, great tit males were able to compensate behaviourally or physiologically for the increase in locomotion costs we imposed. We found no cross-sectional association between reproductive success and TL, but individuals with higher reproductive success (number of recruits) lost fewer telomere base pairs in the subsequent year. We used the TRF method to measure TL, which method yields a TL distribution for each sample, and the association between reproductive success and telomere loss was more pronounced in the higher percentiles of the telomere distribution, in agreement with the higher impact of ageing on longer telomeres within individuals. Individuals with longer telomeres and less telomere shortening were more likely to survive to the next breeding season, but these patterns did not reach statistical significance. Whether successful individuals are characterized by losing fewer or more base pairs from their telomeres varies between species, and we discuss aspects of ecology and social organisation that may explain this variation.

The genome consists of non-B-DNA structures such as G-quadruplexes (G4) that are involved in the regulation of genome stability and transcription. Telomeric-repeat containing RNA (TERRA) is capable of folding into G-quadruplex and interacting with chromatin remodeler ATRX. Here we show that TERRA modulates ATRX occupancy on repetitive sequences and over genes, and maintains DNA G-quadruplex structures at TERRA target and non-target sites in mouse embryonic stem cells. TERRA prevents ATRX from binding to subtelomeric regions and represses H3K9me3 formation. G4 ChIP-seq reveals that G4 abundance decreases at accessible chromatin regions, particularly at transcription start sites (TSS) after TERRA depletion; such G4 reduction at TSS is associated with elevated ATRX occupancy and differentially expressed genes. Loss of ATRX alleviates the effect of gene repression caused by TERRA depletion. Immunostaining analyses demonstrate that knockdown of TERRA diminishes DNA G4 signals, whereas silencing ATRX elevates G4 formation. Our results uncover an epigenetic regulation by TERRA that sequesters ATRX and preserves DNA G4 structures.

SummaryTelomeres in Drosophila are composed of sequential non-LTR retrotransposons: HeT-A, TART, and TAHRE. Although they are repressed by the piRNA pathway in the germline, how these retrotransposons are regulated in somatic cells is poorly understood. Here, we show that specific splice variants of Mod(mdg4) repress HeT-A by blocking subtelomeric enhancers in ovarian somatic cells. We found that the Mod(mdg4)-N variant represses HeT-A expression most efficiently among the variants. Mod(mdg4)-N mutant flies show elevated HeT-A expression and female sterility. Mod(mdg4)-N-binding subtelomeric sequences exhibit enhancer-blocking activity, and recruitment of RNA polymerase II (Pol II) on subtelomeres by Mod(mdg4)-N is essential for this enhancer-blocking. Moreover, Mod(mdg4)-N functions to form chromatin boundaries of higher-order chromatin conformation but this mechanism is independent of its Pol II recruitment activity at telomeres/subtelomeres. This study provides a link between enhancer-blocking and telomere regulation, and two different molecular mechanisms exhibited by an insulator protein to orchestrate precise gene expression.

Telomeric repeat-containing RNA (TERRA) is a type of long non-coding RNA transcribed from telomeres, and it forms R-loops by invasion into telomeric DNA. Since either an excessive or inadequate number of R-loops leads to telomere instability, the TERRA levels need to be delicately modulated. In this study, we found that m6A modification presents on the subtelomeric regions of TERRA and stabilizes it, and the loss of METTL3 impacts telomere stability. Mechanically, the m6A modification on TERRA is catalyzed by METTL3, recognized and stabilized by the m6A reader YTHDC1. Knockdown of either METTL3 or YTHDC1 enhances TERRA degradation. The m6A-modified TERRA forms R-loops and promotes homologous recombination which is essential for the alternative lengthening of telomeres (ALT) pathway in cancer cells. METTL3 depletion leads to R-loop reduction, telomere shortening and instability. Altogether, these findings reveal that METTL3 protects telomeres by catalyzing m6A modification on TERRA, indicating that inhibition or deletion of METTL3 is potentially a new avenue for ALT cancer therapy.

Pancreatic neuroendocrine tumors (PanNETs) are a heterogeneous group of neoplasms with increasing incidence and an ill-defined pathobiology. Although many PanNETs are indolent and remain stable for years, a subset may behave aggressively and metastasize widely. Thus, the increasing and frequent detection of PanNETs presents a treatment dilemma. Current prognostic systems are susceptible to interpretation errors, sampling issues, and do not accurately reflect the clinical behavior of these neoplasms. Hence, additional biomarkers are needed to improve the prognostic stratification of patients diagnosed with a PanNET. Recent studies have identified alterations in death domain-associated protein 6 (DAXX) and alpha-thalassemia/mental retardation X-linked (ATRX), as well as alternative lengthening of telomeres (ALT), as promising prognostic biomarkers. This review summarizes the identification, clinical utility, and specific nuances in testing for DAXX/ATRX by immunohistochemistry and ALT by telomere-specific fluorescence in situ hybridization in PanNETs. Furthermore, a discussion on diagnostic indications for DAXX, ATRX, and ALT status is provided to include the distinction between PanNETs and pancreatic neuroendocrine carcinomas (PanNECs), and determining pancreatic origin for metastatic neuroendocrine tumors in the setting of an unknown primary.

In brief COVID-19 does not affect the telomeres or fertility outcomes in mild cases. However, in women with severe symptoms, telomeres of granulosa cells are shorter, and the oocyte maturation rate is decreased. Abstract The coronavirus SARS-CoV-2 causes COVID-19 disease and affects primarily the lungs and also other organs, causing accelerated cell aging. One of the main pathways involved in aging is telomere attrition, which ultimately leads to defective tissue regeneration and organ dysfunction. Indeed, short telomeres in aged people aggravate the COVID-19 symptoms, and COVID-19 survivors showed shorter telomeres in blood cells. The SARS-CoV-2 has been detected in testis, but the ovaries, which express the viral entry factors, have not been fully explored. Our objective was to analyze telomeres and reproductive outcomes in women who had COVID-19 and controls. In this prospective cohort study, granulosa cells (GCs) and blood were collected from 65 women. Telomere length (TL) was measured by high-throughput in situ hybridization. Mean TL of GCs and peripheral blood mononuclear cells (PBMCs) was alike in control and mild cases. However, mean TL of GCs was lower in severe cases compared to controls (P = 0.017). Control and COVID groups had similar ovarian reserve and number of total oocytes after puncture. However, the oocyte maturation rate was lower in severe cases (P = 0.018). Interestingly, a positive correlation between the oocyte maturation rate and TL of GCs was found in the control group (P = 0.024). Our findings point to a potential impact of the coronavirus infection on telomeres and reproductive outcomes in severe cases. This might be considered upon possible new SARS-CoV threats, to favor treatments that enhance oocyte maturation in women severely affected by coronavirus undergoing ART.

The incidence of rheumatoid arthritis (RA) and its complications are expected to increase with age. Remarkably, RA patients were identified features of accelerated aging, particularly in immunosenescence. As is known, T cells in RA patients readily differentiate into pro-inflammatory phenotypes that maintain chronic and persistent inflammatory changes in joints and many other organ systems. Recent evidence suggests that T cells are most sensitive to aging, and aged CD4+ T cells contribute to inflammaging, which plays a crucial role in accelerating the disease process. In recent years, the molecular mechanisms of T cell immunosenescence were beginning to be understood. Immune aging in RA T cells is associated with thymus insufficiency, metabolic abnormalities, shortened telomere length, and chronic energy stress. Therefore, we summarized the role and mechanism of T cell immunosenescence in RA.A computer-based online search was performed using the PubMed database for published articles concerning T cells aging and rheumatoid arthritis.In this review, we assess the roles of CD4+ T cells in the center of inflammaging especially in RA and emphasize arthritogenic effector functions of senescent T cell; also we discuss the possible molecular mechanisms of senescent T cells and therapeutic targets to intervene T cells immunosenescence for improvement of RA.

ConspectusG-quadruplexes (G4s) are distinctive four-stranded DNA or RNA structures found within cells that are thought to play functional roles in gene regulation and transcription, translation, recombination, and DNA damage/repair. While G4 structures can be uni-, bi-, or tetramolecular with respect to strands, folded unimolecular conformations are most significant

Introduction: Aging is characterized as a syndrome of deleterious, progressive, universal, and irreversible function changes affecting every structural and functional aspect of the organism and accompanied by a generalized increase in mortality. Although a substantial number of candidates for biomarkers of aging have been proposed, none has been validated or universally accepted. Human telomeres constitute hexameric repetitive DNA sequence nucleoprotein complexes that cap chromosome ends, regulating gene expression and modulating stress-related pathways. Telomere length (TL) shortening is observed both in cellular senescence and advanced age, leading to the investigation of TL as a biomarker for aging and a risk factor indicator for the development and progression of the most common age-related diseases. Objective: The present review underlines the connection between TL and the pathophysiology of the diseases associated with telomere attrition. Methods: We performed a structured search of the PubMed database for peer-reviewed research of the literature regarding leukocyte TL and cardiovascular diseases (CVD), more specifically stroke and heart disease, and focused on the relevant articles published during the last 5 years. We also applied Hill’s criteria of causation to strengthen this association. Result: We analyzed the recent literature regarding TL length, stroke, and CVD. Although approximately one-third of the available studies support the connection, the results of different studies seem to be rather conflicting as a result of different study designs, divergent methods of TL determination, small study samples, and patient population heterogeneity. After applying Hill’s criteria, we can observe that the literature conforms to them weakly, with chronology being the only Hill criterion of causality that probably cannot be contested. Conclusion: The present review attempted to examine the purported relation between leukocyte TL and age-related diseases such as CVD and more specific stroke and heart disease in view of the best established, comprehensive, medical and epidemiological criteria that have characterized the focused recent relevant research. Although several recommendations have been made that may contribute significantly to the field, a call for novel technical approaches and studies is mandatory to further elucidate the possible association.

Telomeres cap the ends of eukaryotic chromosomes, and the telomere length (TL) is related to cellular age. The mitochondrial DNA copy number (mtDNAcn) reflects the abundance of mitochondria in a cell. In addition to generating energy, mitochondria are also the main producers of reactive oxygen species, which in turn can accelerate TL attrition and impair mitochondrial function. Nutrition in early life could influence mtDNAcn and TL in later life. In the present study, we investigated the effects of feeding different levels of milk replacer (MR) on TL shortening and energetic status by examining mtDNAcn of heifers during their first year of life. In this study, whole blood samples were obtained from German Holstein heifer calves 36 to 48 h after birth (wk 1) and at wk 12 and wk 16 of life (n = 37), as well as from 31 calves when reaching 1 yr of age. Calves were fed either a high level of MR (14% solids) at 10 L/d (1.4 kg of MR/d; n = 18) or a restrictive low level at 5.7 L/d (0.8 kg of MR/d; n = 19) until linear weaning in wk 13 to 14 of life. Additional whole blood samples were taken from their respective dams 36 to 48 h after calving. Relative TL (qT) and mtDNAcn in cells from whole blood were measured by multiplex quantitative PCR. The greatest qT values were observed in neonates (36-48 h after birth), with decreasing qT values thereafter. Delta qT values were calculated as ΔqT = qT (first year of life) - initial qT (36-48 h after birth). We found no effect of the feeding regimen on qT values, but qT decreased with age. The mtDNAcn was lowest in neonates, increased until wk 12 of life, and then remained at a constant level until after weaning (wk 16). After the first year of life, mtDNAcn was decreased and returned to levels comparable to those of the neonatal stage. No differences in mtDNAcn were detectable between feeding groups within each time point. When comparing the values of qT and mtDNAcn between the calves and their dams after calving (36-48 h after birth and after calving), greater values were observed in calves than in dams. Delta qT values were negative in all but 2 calves (on the restricted diet), indicating that the change in TL with age was not uniform among individual animals, whereas no difference in mean ΔqT values occurred between the feeding groups. Additional analyses of the correlation between qT, mtDNAcn, and various indicators of oxidative status from birth until wk 16 of life did not indicate major interactions between oxidative status, qT and mtDNAcn. The results of this study support an age-dependent decrease of TL in calves independent of the MR feeding level and show the dynamic changes of mtDNAcn in early life.

Telomere biology disorders (TBDs) are a group of rare diseases caused by mutations that impair telomere maintenance. Mutations that cause reduced levels of TERC/hTR, the telomerase RNA component, are found in most TBD patients and include loss-of-function mutations in hTR itself, in hTR-binding proteins [NOP10, NHP2, NAF1, ZCCHC8, and dyskerin (DKC1)], and in proteins required for hTR processing (PARN). These patients show diverse clinical presentations that most commonly include bone marrow failure (BMF)/aplastic anemia (AA), pulmonary fibrosis, and liver cirrhosis. There are no curative therapies for TBD patients. An understanding of hTR biogenesis, maturation, and degradation has identified pathways and pharmacological agents targeting the poly(A) polymerase PAPD5, which adds 3'-oligoadenosine tails to hTR to promote hTR degradation, and TGS1, which modifies the 5'-cap structure of hTR to enhance degradation, as possible therapeutic approaches. Critical next steps will be clinical trials to establish the effectiveness and potential side effects of these compounds in TBD patients.

Parenchymal lung disease is the fourth leading cause of death in the United States; among the top causes, it continues on the rise. Telomeres and telomerase have historically been linked to cellular processes related to aging and cancer, but surprisingly, in the recent decade genetic discoveries have linked the most apparent manifestations of telomere and telomerase dysfunction in humans to the etiology of lung disease: both idiopathic pulmonary fibrosis (IPF) and emphysema. The short telomere defect is pervasive in a subset of IPF patients, and human IPF is the phenotype most intimately tied to germline defects in telomere maintenance. One-third of families with pulmonary fibrosis carry germline mutations in telomerase or other telomere maintenance genes, and one-half of patients with apparently sporadic IPF have short telomere length. Beyond explaining genetic susceptibility, short telomere length uncovers clinically relevant syndromic extrapulmonary disease, including a T-cell immunodeficiency and a propensity to myeloid malignancies. Recognition of this subset of patients who share a unifying molecular defect has provided a precision medicine paradigm wherein the telomere-mediated lung disease diagnosis provides more prognostic value than histopathology or multidisciplinary evaluation. Here, we critically evaluate this progress, emphasizing how the genetic findings put forth a new pathogenesis paradigm of age-related lung disease that links telomere abnormalities to alveolar stem senescence, remodeling, and defective gas exchange.

Both myeloid cells, which contribute to innate immunity, and lymphoid cells, which dominate adaptive immunity, partake in defending against SARS-CoV-2. In response to the virus, the otherwise slow haematopoietic production supply chain quickly unleashes its preconfigured myeloid element, which largely resists a bullwhip-like effect. By contrast, the lymphoid element risks a bullwhip-like effect when it produces T cells and B cells that are specifically designed to clear the virus. As T-cell production is telomere-length dependent and telomeres shorten with age, older adults are at higher risk of a T-cell shortfall when contracting SARS-CoV-2 than are younger adults. A poorly calibrated adaptive immune response, stemming from a bullwhip-like effect, compounded by a T-cell deficit, might thus contribute to the propensity of people with inherently short T-cell telomeres to develop severe COVID-19. The immune systems of these individuals might also generate an inadequate T-cell response to anti-SARS-CoV-2 vaccination.

Guanine-rich DNA sequences may fold back into non-canonical four-stranded secondary structures termed as G-quadruplexes. The role of G-quadruplexes has already been well established in different diseases like cancer, neurological and viral disorders etc. Also, several small molecules have been reported, which can influence the involvement of G-quadruplexes either through stabilization or destabilization in the cellular environment. Growing statistics have associated G-quadruplex assemblies to a discrete biological process in vivo, including DNA replication, transcription, genomic stability, and epigenetic regulation. DNA G-quadruplex existence in human telomere is well recognized attractive target for anticancer drugs. G-quadruplex-interactive ligands have been known to prevent telomerase access as well as telomere capping. To the best of our understanding, the role of G-quadruplexes in virology, neuropharmacology, cancer progression and its treatment has not been discussed on a single platform till date. This review aims to enhance our knowledge regarding these magical sticky quadruplex structures, which have been quite significantly proved to be the part of many cellular processes along with their established in vivo existence. Understanding regarding stabilizing or destabilizing ligands of these multistranded guanine quadruplex structures might be proved as the facilitator of drug discovery process for many incurable diseases in future.

Telomeres are DNA sequences of tandem TTAGGG repeats that protect chromosome ends from degradation and instability. Constitutional loss-of-function telomerase mutations result in rapid telomere shortening, premature senescence and cell death. Liver cirrhosis is rare and has only been reported in adults. We present five family members of Bedouin-Muslim origin, all of which carry the same mutation, and yet demonstrate an extremely variable phenotypical presentation, including liver cirrhosis during early childhood.A multidisciplinary long-term follow-up of two healthy and three affected patients was analysed. The mutation (r.95GC) was identified in all patients using Sanger sequencing. Telomere length samples were obtained and analysed.Clinical phenotypes were extremely variable, including age at first symptoms, organ involvement, disease severity and patient prognosis. The most prominent clinical phenotype is liver involvement, including end-stage liver disease early in life, which affects three members of the family. Affected patients had markedly shorter telomeres.We describe an unusual presentation of early liver failure in telomere disease patients. Little, if any, is known about the association between the genotype and phenotype among children with telomere disease and whether the mutation we have described (r.95GC) is predisposed to early severe hepatic involvement.

Zbtb34 is a novel zinc finger protein, which is revealed by biological software analysis to have 3 zinc fingers, but its functions remain unknown. In this study, mouseiZbtb34/icDNA was amplified by PCR and inserted into the plasmid pEGFP-N1 to generate Zbtb34-EGFP fusion protein. The upregulation of Zbtb34 in mouse embryonic stem cells promoted telomere elongation and increased cell proliferation. In order to understand the above phenomena, the telomere co-immunoprecipitation technique was employed to investigate the relationship between Zbtb34 and telomeres. The results indicated that Zbtb34 could bind to the DNA sequences of the telomeres. Alanine substitution of the third zinc finger abolished such binding. Since Pot1 is the only protein binding to the single-stranded DNA at the end of the telomeres, we further investigated the relationship between Zbtb34 and Pot1. The results revealed that the upregulation of Zbtb34 decreased the binding of Pot1b to the telomeres. Through the upregulation of Pot1b, the binding of Zbtb34 to the telomeres was also reduced. In conclusion, we showed that the main biological function of Zbtb34 was to bind telomere DNA via its third ZnF, competing with Pot1b for the binding sites, resulting in telomere elongation and cell proliferation.

ConspectusDNA G-quadruplex secondary structures formed in guanine-rich human telomeres and oncogene promoters are functionally important and have emerged as a promising new class of cancer-specific drug targets. These globular intramolecular structures are stabilized by K

Over the last decades, selection in cattle has mainly been based on milk production rather than on reproductive efficiency. While, when applied, focus on reproduction has involved females, attention has barely been paid to males and, if so, it has only looked at classical sperm quality parameters. In effect, variables such as telomere length have been missed, despite the fact that longer telomeres have been suggested to be linked to male fertility in humans. For this reason, the present study aimed to determine the length of telomeres in bovine sperm and their relationship with a) sperm quality evaluated through the conventional spermiogram and flow cytometry, and b) bull reproductive performance. For this purpose, 29 bulls were involved in this study. Sperm telomere length was evaluated through quantitative Fluorescent In Situ Hybridization (qFISH), and sperm quality was determined at 0 h and 4 h post-thaw. Bull fertility was assessed as non-return to estrus rates after 90 days of artificial insemination. Although the mean telomere length in bovine sperm was 12.06 ± 2.75 kb, the intra-individual variability in length led us to observe three different groups of telomeres in each sperm cell: short telomeres (7.14% ± 5.79% of telomeres; 8.29 ± 2.34 kb), medium telomeres (31.03% ± 12.92% of telomeres; 16.00 ± 2.72 kb) and long telomeres (61.93% ± 18.11% of telomeres; 30.13 ± 11.35 kb). Moreover, whereas reactive oxygen species (ROS) were found to be correlated to sperm telomere length (Rs = −0.492; P= 0.007), no correlation with other sperm quality parameters was found (P > 0.05). Reproductive performance after artificial insemination was not seen to be correlated to sperm telomere length (Rs = 0.123; P= 0.520). In conclusion, this study determined, for the first time, the mean telomere length in bovine sperm and also reported that there is a high variability within each sperm cell. Yet, while telomere length was found to be correlated to ROS generation, it was not related to bull reproductive performance The present study was funded by the Ministry of Science and Innovation, Spain (AGL2017-88329-R and PID2020-113320RB-I00), the Regional Government of Catalonia, Spain (2017-SGR-1229 and 2020-FI-B-00412), the European Commission (H2020 Research and Innovation program under the Marie Skłodowska-Curie grant agreement No 801342; Tecniospring INDUSTRY, TECSPR-19-1- 0003), and La Marato de TV3 Foundation (Grant: 214/857-202039) Open Access funding provided thanks to the CRUE-CSIC agreement with Elsevier

This investigation was designed to study the relationship between weight change over 10 years and leukocyte telomere length (LTL) in a large sample of 3070 randomly selected U.S. adults, 36–70 years old. The National Health and Nutrition Examination Survey (NHANES) data were used to examine the relationship between percent weight change and LTL. Potential mediating variables were controlled using partial correlation. After adjusting for age, race, year, and housing status, the association between percent weight change over 10 years and LTL was significant in women (F = 6.9, p = 0.0138). Adjusting for the demographic and several other covariates weakened the relationship slightly (F = 4.7, p = 0.0392). With all the covariates controlled, for each one percentage point increase in weight over the previous 10 years, telomeres were, on average, 3.48 base pairs (bp) shorter in women. Given that each one-year increase in age was associated with telomeres that were 15.0 bp shorter in women, the median weight change in U.S. women over the previous 10 years (an increase of 10.4%) was predictive of LTLs that were 36 bp shorter, on average, or an increase of 2.4 years of biological aging. Percent weight change over 10 years was not associated with LTL in U.S. men. Percent weight change over 10 years is a strong predictor of biological aging in U.S. women, but not in men.

COPD, one of world’s leading contributors to morbidity and mortality, is characterized by airflow limitation and heterogeneous clinical features. Three main phenotypes are proposed: overlapping asthma/COPD (ACO), exacerbator, and emphysema. Disease severity can be classified as mild, moderate, severe, and very severe. The molecular basis of inflammatory amplification, cellular aging, and immune response are critical to COPD pathogenesis. Our aim was to investigate EP300 (histone acetylase, HAT), HDAC 2 (histone deacetylase), HDAC3, and HDAC4 gene expression, telomere length, and differentiation ability to M1/M2 macrophages. For this investigation, 105 COPD patients, 42 smokers, and 73 non-smoker controls were evaluated. We identified a reduced HDAC2 expression in patients with mild, moderate, and severe severity; a reduced HDAC3 expression in patients with moderate and severe severity; an increased HDAC4 expression in patients with mild severity; and a reduced EP300 expression in patients with severe severity. Additionally, HDAC2 expression was reduced in patients with emphysema and exacerbator, along with a reduced HDAC3 expression in patients with emphysema. Surprisingly, smokers and all COPD patients showed telomere shortening. COPD patients showed a higher tendency toward M2 markers. Our data implicate genetic changes in COPD phenotypes and severity, in addition to M2 prevalence, that might influence future treatments and personalized therapies.

Telomere length (TL) is a marker of biological aging, and shorter telomeres have been associated with several medical and psychiatric disorders, including cardiometabolic dysregulation and Major Depressive Disorder (MDD). In addition, studies have shown shorter TL to be associated with poorer response to certain psychotropic medications, and our previous work suggested shorter TL and higher telomerase activity (TA) predicts poorer response to Selective Serotonin Reuptake Inhibitor (SSRI) treatment. Using a new group of unmedicated medically healthy individuals with MDD (n = 48), we sought to replicate our prior findings demonstrating that peripheral blood mononuclear cell (PBMC) TL and TA predict response to SSRI treatment and to identify associations between TL and TA with biological stress mediators and cardiometabolic risk indices. Our results demonstrate that longer pre-treatment TL was associated with betterresponse to SSRI treatment (β = .407 p = .007). Additionally, we observed that TL had a negative relationship with allostatic load (β = - .320 p = .017) and a cardiometabolic risk score (β = - .300 p = .025). Our results suggest that PBMC TL reflects, in part, the cumulative effects of physiological stress and cardiovascular risk in MDD and may be a biomarker for predicting SSRI response.

BackgroundTheMUC5Bpromoter variant (rs35705950) and telomere length are linked to pulmonary fibrosis and CT-based qualitative assessments of interstitial abnormalities, but their associations with longitudinal quantitative changes of the lung interstitium among community-dwelling adults are unknown.MethodsWe used data from participants in the Multi-Ethnic Study of Atherosclerosis with high-attenuation areas (HAAs, Examinations 1–6 (2000–2018)) andMUC5Bgenotype (n=4552) and telomere length (n=4488) assessments. HAA was defined as the per cent of imaged lung with attenuation of −600 to −250 Hounsfield units. We used linear mixed-effects models to examine associations ofMUC5Brisk allele (T) and telomere length with longitudinal changes in HAAs. Joint models were used to examine associations of longitudinal changes in HAAs with death and interstitial lung disease (ILD).ResultsTheMUC5Brisk allele (T) was associated with an absolute change in HAAs of 2.60% (95% CI 0.36% to 4.86%) per 10 years overall. This association was stronger among those with a telomere length below an age-adjusted percentile of 5% (p value for interaction=0.008). A 1% increase in HAAs per year was associated with 7% increase in mortality risk (rate ratio (RR)=1.07, 95% CI 1.02 to 1.12) for overall death and 34% increase in ILD (RR=1.34, 95% CI 1.20 to 1.50). Longer baseline telomere length was cross-sectionally associated with less HAAs from baseline scans, but not with longitudinal changes in HAAs.ConclusionsLongitudinal increases in HAAs were associated with theMUC5Brisk allele and a higher risk of death and ILD.

Telomere biology was first studied in maize, ciliates, yeast, and mice, and in recent decades, it has informed understanding of common disease mechanisms with broad implications for patient care. Short telomere syndromes are the most prevalent premature aging disorders, with prominent phenotypes affecting the lung and hematopoietic system. Less understood are a newly recognized group of cancer-prone syndromes that are associated with mutations that lengthen telomeres. A large body of new data from Mendelian genetics and epidemiology now provides an opportunity to reconsider paradigms related to the role of telomeres in human aging and cancer, and in some cases, the findings diverge from what was interpreted from model systems. For example, short telomeres have been considered potent drivers of genome instability, but age-associated solid tumors are rare in individuals with short telomere syndromes, and T cell immunodeficiency explains their spectrum. More commonly, short telomeres promote clonal hematopoiesis, including somatic reversion, providing a new leukemogenesis paradigm that is independent of genome instability. Long telomeres, on the other hand, which extend the cellular life span in vitro, are now appreciated to be the most common shared germline risk factor for cancer in population studies. Through this contemporary lens, I revisit here the role of telomeres in human aging, focusing on how short and long telomeres drive cancer evolution but through distinct mechanisms.

The work deals with molecular dynamics (MD) simulations of protonated, human telomeric i-motif deoxyribonucleic acid (DNA) with functionalized graphene. We studied three different graphene sheets: unmodified graphene with hydrogen atoms attached to their edges and two functionalized ones. The functionalization of graphene edge consists in attaching partially protonated or dissociated amine and carboxyl groups. We found that in all cases the protonated i-motif adsorbs strongly on the graphene surface. The biased MD simulations showed that the work necessary to drag the i-motif out from amine-doped graphene is about twice larger than that in other cases. In general, the system i-motif/amine-doped graphene stands out from the rest, e.g., in this case, the i-motif adsorbs its side with 3' and 5' ends oriented in the opposite to surface direction. In other cases, the DNA fragment is adsorbed to graphene by 3' and 5' ends. In all cases, the adsorption on graphene influences the i-motif internal structure by changing the distances between i-motif strands as well as stretching or shortening the DNA chain, but only in the case of amine-doped graphene the adsorption affects internal H-bonds formed between nucleotides inside the i-motif structure.

Mutations in DKC1 (encoding dyskerin) cause telomere diseases including dyskeratosis congenita (DC) by decreasing steady-state levels of TERC, the non-coding RNA component of telomerase. How DKC1 mutations variably impact numerous other snoRNAs remains unclear, which is a barrier to understanding disease mechanisms in DC beyond impaired telomere maintenance. Here, using DC patient iPSCs, we show that mutations in the dyskerin N-terminal extension domain (NTE) dysregulate scaRNA13. In iPSCs carrying the del37L NTE mutation or engineered to carry NTE mutations via CRISPR/Cas9, but not in those with C-terminal mutations, we found scaRNA13 transcripts with aberrant 3′ extensions, as seen when the exoribonuclease PARN is mutated in DC. Biogenesis of scaRNA13 was rescued by repair of the del37L DKC1 mutation by genome-editing, or genetic or pharmacological inactivation of the polymerase PAPD5, which counteracts PARN. Inspection of the human telomerase cryo-EM structure revealed that in addition to mediating intermolecular dyskerin interactions, the NTE interacts with terminal residues of the associated snoRNA, indicating a role for this domain in 3′ end definition. Our results provide mechanistic insights into the interplay of dyskerin and the PARN/PAPD5 axis in the biogenesis and accumulation of snoRNAs beyond TERC, broadening our understanding of ncRNA dysregulation in human diseases.

Telomere maintenance is essential for maintaining genome integrity in both normal and cancer cells. Without functional telomeres, chromosomes lose their protective structure and undergo fusion and breakage events that drive further genome instability, including cell arrest or death. One means by which this loss can be overcome in stem cells and cancer cells is via re-addition of G-rich telomeric repeats by the telomerase reverse transcriptase (TERT). During aging of somatic tissues, however, insufficient telomerase expression leads to a proliferative arrest called replicative senescence, which is triggered when telomeres reach a critically short threshold that induces a DNA damage response. Cancer cells express telomerase but do not entirely escape telomere instability as they often possess short telomeres; hence there is often selection for genetic alterations in the TERT promoter that result in increased telomerase expression. In this review, we discuss our current understanding of the consequences of telomere instability in cancer and aging, and outline the opportunities and challenges that lie ahead in exploiting the reliance of cells on telomere maintenance for preserving genome stability.

A subset of cancers across multiple histologies with predominantly poor outcomes use the alternative lengthening of telomeres (ALT) mechanism to maintain telomere length, which can be identified with robust biomarkers. ALT has been reported to be prevalent in high-risk neuroblastoma and certain sarcomas, and ALT cancers are a major clinical challenge that lack targeted therapeutic approaches. Here, we found ALT in a variety of pediatric and adult cancer histologies, including carcinomas. Patient-derived ALT cancer cell lines from neuroblastomas, sarcomas, and carcinomas were hypersensitive to the p53 reactivator eprenetapopt (APR-246) relative to telomerase-positive (TA+) models. Constitutive telomere damage signaling in ALT cells activated ataxia-telangiectasia mutated (ATM) kinase to phosphorylate p53, which resulted in selective ALT sensitivity to APR-246. Treatment with APR-246 combined with irinotecan achieved complete responses in mice xenografted with ALT neuroblastoma, rhabdomyosarcoma, and breast cancer and delayed tumor growth in ALT colon cancer xenografts, while the combination had limited efficacy in TA+ tumor models. A large number of adult and pediatric cancers present with the ALT phenotype, which confers a uniquely high sensitivity to reactivation of p53. These data support clinical evaluation of a combinatorial approach using APR-246 and irinotecan in ALT patients with cancer. Significance: This work demonstrates that constitutive activation of ATM in chemotherapy-refractory ALT cancer cells renders them hypersensitive to reactivation of p53 function by APR-246, indicating a potential strategy to overcome therapeutic resistance.

Telomere dynamics are linked with both cellular and organismal senescence, and life history, individual quality and health. Telomere dynamics, particularly telomere length, have therefore garnered much research interest in evolutionary biology. To examine the evolution of telomere length, it is important to quantify its heritability, the proportion of total variation explained by additive genetic effects. Many studies have quantified telomere length heritability, but estimates are varied, and no general conclusion has been drawn. Additionally, it is unclear whether biological and methodological factors influence telomere length heritability estimates. We present the first meta-analysis of telomere length heritability, using 104 estimates from 43 studies over 18 vertebrate species. We calculated an overall mean heritability and examined how estimates varied by study, phylogeny, species-specific ecology, environmental setting, age at sampling, laboratory methods, statistical methods, sex and repeated measurements. Overall heritability was moderate (44.9%, 95% CI: 25.2-64.7%), and there was considerable heterogeneity in heritability estimates, in particular among studies and estimates. Laboratory method influenced heritability estimates, with in-gel hybridization TRF yielding higher heritabilities than qPCR and Southern blot TRF. There was also an effect from statistical method, with twin-based and SNP-based estimates lower than correlation-based or pedigree-based estimates. Our results highlight an overall heritable basis of telomere length, and we recommend future research on a wider range of taxa, and the use of variance-partitioning methods with relatedness or SNP data over correlation methods to minimize heritability estimation bias.

Dyskeratosis congenita (DC) is a multisystem syndrome characterized by mucocutaneous abnormalities, bone marrow failure, and predisposition to cancer. Studies over the last 25 years have led to the identification of 18 disease genes. These have a principal role in telomere maintenance, and patients usually have very short/abnormal telomeres. The advances have also led to the unification of DC with a number of other diseases, now collectively referred to as the telomeropathies or telomere biology disorders.Clinical features, genetics, and biology of the different subtypes. Expert view on diagnosis, treatment of the hematological complications and future.As these are very pleotropic disorders affecting multiple organs, a high index of suspicion is necessary to make the diagnosis. Telomere length measurement and genetic analysis of the disease genes have become useful diagnostic tools. Although hematological defects can respond to danazol/oxymetholone, the only current curative treatment for these is hematopoietic stem cell transplantation (HSCT) using fludarabine-based conditioning protocols. New therapies are needed where danazol/oxymetholone is ineffective and HSCT is not feasible.

Telomeric repeat-containing RNA (TERRA) has been suggested to participate in telomere maintenance. TERRA consisting of UUAGGG repeats is capable of forming an intermolecular G-quadruplex (GQ) with single-stranded TTAGGG-repeat DNA in the telomere 3' overhang. To explore the structural features and potential functions of this DNA-RNA hybrid GQ (HGQ), we used single-molecule FRET to study the folding patterns of DNA with four to seven telomeric tandem repeats annealed with a short RNA consisting of two or five telomeric repeats. Our data highlight that RNA prefers to form DNA-RNA HGQ near the 3' end of telomeric DNA. Furthermore, the unfolding of secondary structures by a complementary C-rich sequence was observed for DNA GQ but not for DNA-RNA HGQ, which demonstrated the enhanced stability of the telomere 3' end via hybridization with RNA. These conformational and physical properties of telomeric DNA-RNA HGQ suggest that TERRA might limit access to the 3' end of the telomeric DNA overhang, which is known to be critical for the interaction with telomerase and other telomere-associated proteins.

Telomeres are the physical ends of linear chromosomes. They are composed of short repeating sequences (such as TTGGGG in the G-strand for Tetrahymena thermophila) of double-stranded DNA with a single-strand 3'overhang of the G-strand and, in humans, the six shelterin proteins: TPP1, POT1, TRF1, TRF2, RAP1 and TIN21,2. TPP1 and POT1 associate with the 3'overhang, with POT1 binding the G-strand3 and TPP1 (in complex with TIN24) recruiting telomerase via interaction with telomerase reverse transcriptase5 (TERT). The telomere DNA ends are replicated and maintained by telomerase6, for the G-strand, and subsequently DNA polymerase α-primase7,8 (PolαPrim), for the C-strand9. PolαPrim activity is stimulated by the heterotrimeric complex CTC1-STN1-TEN110-12 (CST), but the structural basis of the recruitment of PolαPrim and CST to telomere ends remains unknown. Here we report cryo-electron microscopy (cryo-EM)structures of Tetrahymena CST in the context of the telomerase holoenzyme, in boththe absence and thepresence of PolαPrim, and of PolαPrim alone. Tetrahymena Ctc1 binds telomerase subunit p50, a TPP1 orthologue, on a flexible Ctc1 binding motif revealed by cryo-EM and NMR spectroscopy. The PolαPrim polymerase subunit POLA1 binds Ctc1 and Stn1, and its interface with Ctc1 forms an entry port for G-strand DNA to the POLA1 active site. We thus provide a snapshot of four keycomponents that are required for telomeric DNA synthesis in a single active complex-telomerase-core ribonucleoprotein, p50, CST and PolαPrim-that provides insights into the recruitment of CST and PolαPrim and the handoff between G-strand and C-strand synthesis.

The mammalian DNA polymerase-α–primase (Polα–primase) complex is essential for DNA metabolism, providing the de novo RNA–DNA primer for several DNA replication pathways(1–4) such as lagging-strand synthesis and telomere C-strand fill-in. The physical mechanism underlying how Polα–primase, alone or in partnership with accessory proteins, performs its complicated multistep primer synthesis function is unknown. Here we show that CST, a single-stranded DNA-binding accessory protein complex for Polα–primase, physically organizes the enzyme for efficient primer synthesis. Cryogenic electron microscopy structures of the CST-Polα–primase preinitiation complex (PIC) bound to various types of telomere overhang reveal that template-bound CST partitions the DNA and RNA catalytic centres of Polα–primase into two separate domains and effectively arranges them in RNA–DNA synthesis order. The architecture of the PIC provides a single solution for the multiple structural requirements for the synthesis of RNA–DNA primers by Polα–primase. Several insights into the template-binding specificity of CST, template requirement for assembly of the CST-Polα–primase PIC and activation are also revealed in this study.

AIMS: Aging is accompanied by telomere shortening. Increased telomere shortening is considered a marker of premature aging. Cardiac aging results in the development of cardiac pathologies. Electrocardiogram (ECG) measures reflect cardiac excitation, conduction, and repolarization. ECG measures also prolong with aging and are associated with cardiac pathologies including atrial fibrillation. As premature prolongation of ECG measures is observed, we hypothesized that such prolongation may be associated with telomere length. METHODS AND RESULTS: We studied the large, community-based KORA F4 Study. Of 3,080 participants enrolled between 2006 and 2007 with detailed information on demographic, anthropometric, clinical, and ECG characteristics, 2,575 presented with available data on leukocyte telomere length. Telomere length was determined by real-time quantitative PCR and expressed relative to a single copy gene. We fitted multivariable adjusted linear regression models to associate the ECG measures RR-interval, PR-interval, QRS-duration, and heart rate corrected QTc with telomere length. In our cohort, the mean age was 54.9±12.9 years and 46.6% were men. Increased age was associated with shorter telomere length (p

Telomere length (TL) is a biomarker of cellular proliferative history. In healthy individuals, leukocyte TL shortens with age and associates with the lifespan of men and women. However, most of studies had used linear regression models to address the association of the TL attrition, aging and sex.We evaluated the association between the TL, aging and sex in a cohort of 180 healthy subjects by quantile regression. The TL of nucleated blood cells was measured by fluorescent in situ hypridization (flow-FISH) in a cohort of 89 men, 81 women, and 10 umbilical cord samples. The results were validated by quantitative polymerase chain reaction (qPCR) and compared to a linear regression analysis.By quantile regression, telomere dynamics slightly differed between sexes with aging: women had longer telomeres at birth and slower attrition rate than men until the sixth decade of life; after that, TL eroded faster and became shorter than that in men. These differences were not observed by linear regression analysis, as the overall telomere attrition rates in women and men were similar (42 pb per year, p 0.0001 vs. 45 pb kb per year, p 0.0001). Also, qPCR did not recapitulate flow-FISH findings, as the telomere dynamics by qPCR followed a linear model.The quantile regression analysis accurately reproduced a third-order polynomial TL attrition rate in both women and men, but it depended on the technique applied to measure TL. The Flow-FISH reproduced the expected telomere dynamics through life and, differently from the qPCR, was able to detect the subtle TL variations associated with sex and aging.

G-quadruplex regulates a wide spectrum of biological processes, including telomere maintenance, DNA replication and transcription. The development of small molecules to selectively target G-quadruplex and their application remain hotspots in cancer therapy. Here, we explored the biological effect of G-quadruplexes stabilizer Tetra-Pt(bpy) in telomerase-positive cancer cells. Telomere maintenance was evaluated by telomerase repeat amplification protocol, chromosome orientation fluorescence in situ hybridization and telomere restriction fragment assays. We found that Tetra-Pt(bpy) accelerates telomere shortening through dual inhibition of telomerase activity and telomere sister chromatin exchanges mediated by telomeric G-quadruplexes. Consequently, Tetra-Pt(bpy)-treated cancer cells became enriched with extremely short telomeres and produced a strong telomeric DNA damage response following long-term treatment, leading to cell proliferation inhibition and senescence. Experimental evidence from RNA seq and cell migration-related assays showed that Tetra-Pt(bpy) decreased cell-matrix adhesion and inhibited the migration of non-senescent tumor cells. Mechanistically, Tetra-Pt(bpy) induced the formation of G-quadruplexes in focal adhesion kinase (FAK)-encoding gene PTK2, resulting in FAK transcription inhibition. Tetra-Pt(bpy) reduced xenograft tumor formation and inhibited tumor cell growth and migration in mice. This study further elucidates the function of G-quadruplexes in the human genome and reveals the potential of Tetra-Pt(bpy) as a novel chemotherapeutic agent for targeting telomerase-positive cancer cells.

Short telomeres are recognized as risk factor for idiopathic pulmonary fibrosis (IPF). We aimed to assess the role of telomere length (TL) in fibrotic-Interstitial Lung Diseases (f-ILDs) associated with a usual interstitial pneumonia (UIP) pattern as well as in IPF acute exacerbation (IPF-AE).TL was measured from peripheral white blood cells using a multiplex quantitative polymerase chain reaction in consecutive patients with f-ILDs, all presenting UIP pattern in the high-resolution chest-computed-tomography and compared to age-matched healthy controls.Seventy-nine individuals were included (mean age 69.77 ± 0.72 years); 24 stable IPF, 18 IPF-AE, 10 combined pulmonary fibrosis and emphysema, 7 Rheumatoid arthritis-UIP-ILDs and 20 controls. TL in all patients was significantly shorter compared to controls [mean T/S ratio (SE) 0.77 (±0.05) vs 2.26 (±0.36), p 0.001] as well as separately in each one of f-ILD subgroups. IPF-AE patients presented significantly shorter TL compared to stable IPF (p = 0.029). Patients with IPF and shorter than the median TL (0-0.72) showed reduced overall survival (p = 0.004). T/S 0.72 was associated with increased risk for IPF-AE (OR = 30.787, 95% CI: 2.153, 440.183, p = 0.012) independent of age, gender, smoking and lung function impairment. A protective effect of TL was observed, as it was inversely associated with risk of death both in UIP-f-ILDs (HR = 0.174, 95%CI: 0.036, 0.846, p = 0.030) and IPF patients (HR = 0.096, 95%CI: 0.011, 0.849, p = 0.035).Shorter TL characterizes different UIP f-ILDs. Although no difference was observed in TL among diverse UIP subgroups, IPF-AE presented shorter TL compared to stable IPF. Reduced overall survival and higher hazard ratio of death are associated with shorter TL in IPF.

It is challenging to preoperatively distinguish malignant and benign forms of intraductal papillary mucinous neoplasms (IPMNs) of the pancreas. The aims of this study were to investigate whether telomere length is associated with pathological grade of IPMNs and age and to clarify the utility of telomere length as a marker to identify malignant IPMNs.Pancreas tissue was obtained from 28 patients after resection. We measured the telomere lengths of tumor cells in IPMNs and normal duct cells by quantitative fluorescence in situ hybridization. The association of normalized telomere-centromere ratio (NTCR) to pathological grade of IPMNs and age were determined.The NTCR showed a gradual decrease with increasing pathological grade of IPMNs. The NTCR in intermediate- and high-grade dysplasia and adenocarcinoma lesions was significantly shorter than in normal pancreatic ducts (Plt; 0.05). In multivariate analysis, telomere length was most associated with carcinogenesis. When the cutoff value of NTCR was set to 0.74, the sensitivity for detection of high-grade dysplasia and adenocarcinoma was 82.8%, with a specificity of 87.5%.Telomere shortening occurs with carcinogenesis and aging. A significant reduction of telomere length in IPMNs may be useful for surgical decision making.

2021 marked the 80th anniversary of Barbara McClintock's pioneering article on the breakage-fusion-bridge (BFB) cycle. Of the three steps of the BFB cycle, breakage remains the least understood despite its major contribution to mutagenesis. We discuss recent findings shedding light on how chromatin bridges break in yeast and animal cells.