The Impact of Exercise on Telomere Length, DNA Methylation and Metabolic Footprints.

Aging as a major risk factor influences the probability of developing cancer, cardiovascular disease and diabetes, amongst others. The underlying mechanisms of disease are still not fully understood, but research suggests that delaying the aging process could ameliorate these pathologies. A key biological process in aging is cellular senescence which is associated with several stressors such as telomere shortening or enhanced DNA methylation. Telomere length as well as DNA methylation levels can be used as biological age predictors which are able to detect excessive acceleration or deceleration of aging. Analytical methods examining aging are often not suitable, expensive, time-consuming or require a high level of technical expertise. Therefore, research focusses on combining analytical methods which have the potential to simultaneously analyse epigenetic, genomic as well as metabolic changes.

Accelerated Biological Aging Secondary to Cardiometabolic Risk Factors Is a Predictor of Cardiovascular Mortality: A Systematic Review and Meta-analysis.

BACKGROUND: Chronological aging is one of the major risk factors of cardiovascular (CV) disease (CVD); however, the effect of biological aging on CVD and outcomes remain poorly understood. Herein, we evaluated the association between leukocyte telomere length (LTL), a marker of biological age, and CV outcomes. METHODS: We searched PubMed, Embase, Ovid Medline, and Web of Science Core Collection for the studies on the association between LTL and myocardial infarction (MI), CV death, and/or CVD risk factors from inception to July 2020. Extracted data were pooled in a random-effects meta-analysis and summarized as risk ratio (RR) and corresponding 95% confidence interval (CI) per LTL tertile. RESULTS: A total of 32 studies (n = 144,610 participants) were included. In a pooled analysis of MI and LTL in a multivariate-adjusted model, the shortest LTL was associated with a 39% higher risk of MI (RR, 1.39; 95% CI, 1.16-1.67; P < 0.001). After adjusting for chronological age and traditional covariance, we showed a 28% increased risk of CV death in the shortest tertile of LTL (RR, 1.28; 95% CI, 1.05-1.56; P = 0.01). Analysis of the studies that investigated the association between CV risk factors and LTL (n = 7) showed that diabetes mellitus is associated with a 46% increased risk of LTL attrition (RR, 1.46; 95% CI, 1.46-2.09; P = 0.039). CONCLUSIONS: This study shows a strong association between LTL, a marker of biological aging, and the risk of MI and CV death. Cardiometabolic risk factors contribute to telomere attrition and therefore accelerates biological aging.

The relationship between telomere length and putative markers of vascular ageing: A systematic review and meta-analysis.

Accelerated biological aging contributes to the evolution of cardiovascular disease. However, its influence on subclinical organ damage remains unclear. Leukocyte telomere length (LTL) is emerging as a marker of biological cardiovascular aging. We performed a systematic review and meta-analysis to assess the association between LTL and measures of end-organ damage. PubMed, Medline, Embase, Cinahl Plus, ClinicalTrials.gov, and grey literature databases were searched for studies that assessed the association of LTL with arterial pulse wave velocity (aPWV), carotid intima-media thickness (cIMT), left ventricular mass (LVM or LVMI), renal outcomes, coronary artery calcium (CAC) and presence of carotid plaques. In a sample of 7256 patients, we found that cIMT (pooled correlation coefficient (r) = -0.249; 95 %CI -0.37, -0.128) and aPWV (pooled r = -0.194; 95 % CI -0.290, -0.100) inversely correlate with LTL. Compared to aPWV, cIMT had a stronger correlation with LTL. Patients without carotid plaques had longer telomeres than patients with carotid plaques. Quantitative analyses documented LTL association with renal outcomes and CAC, but not with LVM/LVMI. Among measures of end-organ damage, cIMT and aPWV provide the most accurate information on the contribution of biological aging to the process of vascular remodeling/damage.

Neuroblastoma-Telomere maintenance, deregulated signaling transduction and beyond.

The childhood malignancy neuroblastoma belongs to the group of embryonal tumors and originates from progenitor cells of the sympathoadrenal lineage. Treatment options for children with high-risk and relapsed disease are still very limited. In recent years, an ever-growing molecular diversity was identified using (epi)-genetic profiling of neuroblastoma tumors, indicating that molecularly targeted therapies could be a promising therapeutic option. In this review article, we summarize the various molecular subtypes and genetic events associated with neuroblastoma and describe recent advances in targeted therapies. We lay a strong emphasis on the importance of telomere maintenance mechanisms for understanding tumor progression and risk classification of neuroblastoma.

Pan-Cancer Analysis of Clinical Relevance via Telomere Maintenance Mechanism.

Understanding the telomere maintenance mechanism (TMM) in immortal cancer cells is vital for TMM-targeted therapies in clinical settings. In this study, we classified four telomere maintenance mechanisms into telomerase, ALT, telomerase + ALT, and non-defined telomere maintenance mechanism (NDTMM) across 31 cancer types using 10,704 transcriptomic datasets from The Cancer Genome Atlas. Our results demonstrated that approximately 50% of the total cohort displayed ALT activity with high telomerase activity in most cancer types. We confirmed significant patient prognoses according to distinct TMMs in six cancer types: adrenocortical carcinoma (ACC), PAAD, HNSC, SARC, GBM, and metastatic cancer. Patients with metastasis had a poor prognosis in the ALT group (p < 0.006) subjected to RAS protein signal transduction. Glioblastoma patients had poor prognosis in NDTMM (p < 0.0043) and showed high levels of myeloid leukocyte activation. Pancreatic adenocarcinoma (p < 0.04) and head and neck squamous cell carcinoma (p < 0.046) patients had a good prognosis in the ALT group with high immune cell activation. Furthermore, we showed that master transcriptional regulators might affect the selection of the TMM pathway and explained why different telomere maintenance mechanisms exist. Furthermore, they can be used to segregate patients and predict responders to different TMM-targeted therapeutics.

SLX4IP N-terminus dictates telomeric localization in ALT-like castration-resistant prostate cancer cell lines.

BACKGROUND: To ensure replicative immortality in cancer, telomeres must be maintained through activation of telomere maintenance mechanisms (TMMs) that are dependent on telomerase or the alternative lengthening of telomeres (ALT) pathway. Although TMM pathways have traditionally been considered to be mutually exclusive, ALT hallmarks have been identified in cancers defined as being telomerase-positive, supporting TMM coexistence. In castration-resistant prostate cancer (CRPC), in vitro models were thought to be universally dependent on telomerase as the primary TMM; however, CRPC models with androgen receptor (AR) loss demonstrate ALT hallmarks with limited telomerase activity and require ALT-associated PML bodies (APBs) for sustained telomere maintenance. The TMM coexistence in AR-negative CRPC is reliant on the ALT regulator protein, SLX4IP. METHODS: To identify the regions of SLX4IP responsible for the induction of APBs and telomere preservation in CRPC models, five 3xFLAG-tagged SLX4IP constructs were designed and stably introduced into parental C4-2B, DU145, and PC-3 cells. Once generated, these cell lines were interrogated for APB abundance and SLX4IP construct localization via immunofluorescence-fluorescence in situ hybridization (IF-FISH) and coimmunoprecipitation experiments for telomeric localization. Similarly, PC-3 cells with endogenous SLX4IP knockdown and SLX4IP construct introduction were interrogated for APB abundance, telomere length preservation, and senescent rescue. RESULTS: Here, we define the N-terminus of SLX4IP as being responsible for the promotion of the ALT-like phenotype of AR-negative CRPC models. Specifically, the N-terminus of SLX4IP was sufficient for promoting APB formation to a similar degree as full-length SLX4IP across CRPC cell lines. Additionally, APB promotion by the N-terminus of SLX4IP rescued telomere shortening and senescent induction triggered by SLX4IP knockdown in AR-negative CRPC cells. Moreover, APB formation and telomere maintenance were dependent on the ability of the N-terminus to direct SLX4IP localization at telomeres and APBs. CONCLUSIONS: These findings identify the role of the uncharacterized ALT regulator SLX4IP in the promotion of TMM coexistence to perpetuate replicative immortality in CRPC in vitro.

Association of accelerated dynamics of telomere sequence loss in peripheral blood leukocytes with incident knee osteoarthritis in Osteoarthritis Initiative cohort.

Osteoarthritis (OA) is a chronic degenerative joint disease, being the main cause of laboral inability. Decreased telomere size in peripheral blood leukocytes (PBL) has been correlated with age-related pathologies, like knee OA. In a dynamic approach, telomere-qPCR was performed to evaluate the relative percentage of PBL telomere loss after a 6-year follow-up, in 281 subjects from the prospective osteoarthritis initiative (OAI) cohort. A radiological Kellgren-Lawrence (KL) grade ≥ 2 was indicative of knee OA. Individuals with knee OA at recruitment (n = 144) showed a higher PBL telomere loss after 6 years than those without knee OA at baseline (n = 137; p = 0.018). Moreover, individuals that developed knee OA during the follow-up (n = 39) exhibited a higher telomere loss compared to those that remained without OA (n = 98; p < 0.001). Logistic regression analysis showed that PBLs telomere loss was not significantly associated with knee OA at recruitment, but behaves as an independent risk factor associated with incidence after follow-up (OR: 1.043; p = 0.041), together with maximum KL grade (OR: 3.627; p = 0.011), body mass index-BMI (OR: 1.252; p < 0.001) and WOMAC-index (OR: 1.247; p = 0.021), at recruitment. The telomere decay in PBLs is faster in individuals with incident knee OA, possibly reflecting a systemic-global accelerated aging that enhances the cartilage degeneration.

Co-expression analysis identifies networks of miRNAs implicated in biological ageing and modulated by short-term interval training.

Exercise training seems to promote healthy biological ageing partly by inducing telomere maintenance, yet the molecular mechanisms are not fully understood. Recent studies have emphasised the importance of microRNAs (miRNAs) in ageing and their ability to mirror pathophysiological alterations associated with age-related diseases. We examined the association between aerobic fitness and leukocyte telomere length before determining the influence of vigorous exercise training on the regulation of leukocyte miRNA networks. Telomere length was positively correlated to aerobic fitness (r = 0.32, p = 0.02). 104 miRNAs were differentially expressed after six weeks of thrice-weekly sprint interval training (SIT) in healthy men (q < 0.05). Gene co-expression analysis (WGCNA) detected biologically meaningful miRNA networks, five of which were significantly correlated with pre-SIT and post-SIT expression profiles (p < 0.001) and telomere length. Enrichment analysis revealed that the immune response, T cell differentiation and lipid metabolism associated miRNAs clusters were significantly down-regulated after SIT. Using data acquired from the Gene Expression Omnibus (GEO), we also identified two co-expressed miRNAs families that were modulated by exercise training in previous investigations. Collectively, our findings highlight the miRNA networks implicated in exercise adaptations and telomere regulation, and suggest that SIT may attenuate biological ageing through the control of the let-7 and miR-320 miRNA families.

Reproductive aging and telomeres: Are women and men equally affected?

Successful reproduction is very important for individuals and for society. Currently, the human health span and lifespan are the object of intense and productive investigation with great achievements, compared to the last century. However, reproduction span does not progress concomitantly with lifespan. Reproductive organs age, decreasing the levels of sexual hormones, which are protectors of health through their action on several organs of the body. Thus, this is the starting point of the organismal decay and infertility. This starting point is easily detected in women. In men, it goes under the surface, undetected, but it goes, nevertheless. Regarding fertility, aging alters the hormonal equilibrium, decreases the potential of reproductive organs, diminishes the quality of the gametes and worsen the reproductive outcomes. All these events happen at a different pace and affecting different organs in women and men. The question is what molecular pathways are involved in reproductive aging and if there is a possible halting or even reversion of the aging events. Answers to all these points will be explained in the present review.

Repeated exposure to challenging environmental conditions influences telomere dynamics across adult life as predicted by changes in mortality risk.

The effects of stress exposure are likely to vary depending on life-stage and stressor. While it has been postulated that mild stress exposure may have beneficial effects, the duration of such effects and the underlying mechanisms are unclear. While the long-term effects of early-life stress are relatively well studied, we know much less about the effects of exposure in adulthood since the early- and adult-life environments are often similar. We previously reported that repeated experimental exposure to a relatively mild stressor in female zebra finches, first experienced in young adulthood, initially had no effect on mortality risk, reduced mortality in middle age, but the apparently beneficial effects disappeared in old age. We show here that this is underpinned by differences between the control and stress-exposed group in the pattern of telomere change, with stress-exposed birds showing reduced telomere loss in middle adulthood. We thereby provide novel experimental evidence that telomere dynamics play a key role linking stress resilience and aging.

Telomere Length Differently Associated to Obesity and Hyperandrogenism in Women With Polycystic Ovary Syndrome.

Background: Polycystic Ovary Syndrome (PCOS) often present metabolic disorders and hyperandrogenism (HA), facts that may influence the telomere length (TL). Aims: To compare the absolute TL (aTL) between women with PCOS and control women, and their association with the presence of obesity and HA parameters. Materials and methods: The PCOS group included 170 unrelated women outpatients and the control group, 64 unrelated donor women. Anthropometric, biochemical-clinical parameters and androgen profile were determined. The PCOS patients were divided accordingly to the presence of obesity and androgenic condition. The aTL was determined from peripheral blood leukocytes by Real Time quantitative PCR. Results: Women with PCOS exhibited a significantly longer aTL than controls after age adjustment (p=0.001). A stepwise multivariate linear regression in PCOS women, showed that WC (waist circumference) contributed negatively (b=-0.17) while testosterone levels contributed positively (b=7.24) to aTL. The non-Obese PCOS (noOB-PCOS) presented the longest aTL when compared to controls (p=0.001). Meanwhile, the aTL was significantly higher in the hyperandrogenic PCOS phenotype (HA-PCOS) than in the controls (p=0.001) and non hyperandrogenic PCOS phenotype (NHA-PCOS) (p=0.04). Interestingly, when considering obesity and HA parameters in PCOS, HA exerts the major effect over the aTL as non-obese HA exhibited the lengthiest aTL (23.9 ± 13.13 Kbp). Conversely, the obese NHA patients showed the shortest aTL (16.5 ± 10.59 Kbp). Conclusions: Whilst a shorter aTL could be related to the presence of obesity, a longer aTL would be associated with HA phenotype. These findings suggest a balance between the effect produced by the different metabolic and hormonal components, in PCOS women.

Biomarkers representing key aging-related biological pathways are associated with subclinical atherosclerosis and all-cause mortality: The Framingham Study.

BACKGROUND: Increased oxidative stress, leukocyte telomere length (LTL) shortening, endothelial dysfunction, and lower insulin-like growth factor (IGF)-1 concentrations reflect key molecular mechanisms of aging. We hypothesized that biomarkers representing these pathways are associated with measures of subclinical atherosclerosis and all-cause mortality. METHODS AND RESULTS: We evaluated up to 2,314 Framingham Offspring Study participants (mean age 61 years, 55% women) with available biomarkers of aging: LTL, circulating concentrations of IGF-1, asymmetrical dimethylarginine (ADMA), and urinary F2-Isoprostanes indexed to urinary creatinine. We evaluated the association of each biomarker with coronary artery calcium [ln (CAC+1)] and carotid intima-media thickness (IMT). In multivariable-adjusted linear regression models, higher ADMA levels were associated with higher CAC values (ßADMA per 1-SD increase 0.25; 95% confidence interval [CI] [0.11, 0.39]). Additionally, shorter LTL and lower IGF-1 values were associated with higher IMT values (ßLTL -0.08, 95%CI -0.14, -0.02, and ßIGF-1 -0.04, 95%CI -0.08, -0.01, respectively). During a median follow-up of 15.5 years, 593 subjects died. In multivariable-adjusted Cox regression models, LTL and IGF-1 values were inversely associated with all-cause mortality (hazard ratios [HR] per SD increase in biomarker, 0.85, 95% CI 0.74-0.99, and 0.90, 95% CI 0.82-0.98 for LTL and IGF-1, respectively). F2-Isoprostanes and ADMA values were positively associated with all-cause mortality (HR per SD increase in biomarker, 1.15, 95% CI, 1.10-1.22, and 1.10, 95% CI, 1.02-1.20, respectively). CONCLUSION: In our prospective community-based study, aging-related biomarkers were associated with measures of subclinical atherosclerosis cross-sectionally and with all-cause mortality prospectively, supporting the concept that these biomarkers may reflect the aging process in community-dwelling adults.

PARP1 modulates telomere sister chromatid exchange and telomere length homeostasis by regulating telomere localization of SLX4 in U2OS cells.

OBJECTIVE: Poly(ADP-ribose) polymerase1 (PARP1) interacts and poly(ADP-ribosyl)ates telomere repeat binding factor 2 (TRF2), which acts as a platform to recruit a large number of proteins at the telomere. Since the discovery of TRF2-SLX4 interaction, SLX4 is becoming the key player in telomere length (TL) maintenance and repair by telomere sister chromatid exchange (T-SCE). Defective TL maintenance pathway results in a spectrum of diseases called telomeropathies like dyskeratosis congenita, aplastic anemia, fanconi anemia, cancer. We aimed to study the role of SLX4 and PARP1 on each other's telomere localization, T-SCE, and TL maintenance in human telomerase-negative osteosarcoma U2OS cells to understand some of the molecular mechanisms of telomere homeostasis. MATERIALS AND METHODS: We checked the role of SLX4 and PARP1 on each other's telomere localization by telomere immunofluorescence. We have cloned full-length wild-type and catalytically inactive mutant PARP1 to understand the role of poly(ADP-ribosyl)ation reaction by PARP1 in telomere length homeostasis. TL of U2OS cells was measured by Q-FISH. T-SCE was measured by Telomere-FISH. KEY FINDINGS: We observed that SLX4 has no role in the telomere localization of PARP1. However, reduced localization of SLX4 at undamaged and damaged telomere upon PARP1 depletion was reversed by overexpression of exogenous wild-type PARP1 but not by overexpression of catalytically inactive mutant PARP1. PARP1 depletion synergized SLX4 depletion-mediated reduction of T-SCE. Furthermore, SLX4 depletion elongated TL, and combined insufficiency of SLX4 with PARP1 further elongated TL. CONCLUSION: So, PARP1 controls SLX4 recruitment at telomere by poly(ADP-ribosyl)ation reaction, thereby regulating SLX4-mediated T-SCE and TL homeostasis.

Number and Replating Capacity of Endothelial Colony-Forming Cells are Telomere Length Dependent: Implication for Human Atherogenesis.

Background Short leukocyte telomere length (TL) is associated with atherosclerotic cardiovascular disease. Endothelial repair plays a key role in the development of atherosclerosis. The objective was to examine associations between TL and proliferative dynamics of endothelial colony-forming cells (ECFCs), which behave as progenitor cells displaying endothelial repair activity. Methods and Results To isolate ECFCs, we performed a clonogenic assay on blood samples from 116 participants (aged 24-94 years) in the TELARTA (Telomere in Arterial Aging) cohort study. We detected no ECFC clones in 29 (group 1), clones with no replating capacity in other 29 (group 2), and clones with replating capacity in the additional 58 (group 3). Leukocyte TL was measured by Southern blotting and ECFCs (ECFC-TL). Age- and sex-adjusted leukocyte TL (mean±SEM) was the shortest in group 1 (6.51±0.13 kb), longer in group 2 (6.69±0.13 kb), and the longest in group 3 (6.78±0.09 kb) (P<0.05). In group 3, ECFC-TL was associated with the number of detected clones (P<0.01). ECFC-TL (7.98±0.13 kb) was longer than leukocyte TL (6.74±0.012 kb) (P<0.0001) and both parameters were strongly correlated (r=0.82; P<0.0001). Conclusions Individuals with longer telomeres display a higher number of self-renewing ECFCs. Our results also indicate that leukocyte TL, as a proxy of TL dynamics in ECFCs, could be used as a surrogate marker of endothelial repair capacity in clinical and laboratory practice because of easy accessibility of leukocytes. Registration URL: https://www.clinicaltrials.gov; Unique identifier: NCT02176941.

Molecular Pathways of Cellular Senescence and Placental Aging in Late Fetal Growth Restriction and Stillbirth.

Abnormally accelerated, premature placental senescence plays a crucial role in the genesis of pregnancy pathologies. Abnormal growth in the third trimester can present as small for gestational age fetuses or fetal growth restriction. One differs from the other by the presence of signs of placental insufficiency and the risk of stillbirth. The majority of stillbirths occur in normally grown fetuses and are classified as "unexplained", which often leads to conclusions that they were unpreventable. The main characteristic of aging is a gradual decline in the function of cells, tissues, and organs. These changes result in the accumulation of senescent cells in mitotic tissues. These cells begin the aging process that disrupts tissues' normal functions by affecting neighboring cells, degrading the extracellular matrix, and reducing tissues' regeneration capacity. Different degrees of abnormal placentation result in the severity of fetal growth restriction and its sequelae, including fetal death. This review aims to present the current knowledge and identify future research directions to understand better placental aging in late fetal growth restriction and unexplained stillbirth. We hypothesized that the final diagnosis of placental insufficiency can be made only using markers of placental senescence.

Nuclear envelope tethering inhibits the formation of ALT-associated PML bodies in ALT cells.

Telomere length homeostasis is essential for maintaining genomic stability and cancer proliferation. Telomerase-negative cancer cells undergo recombination-mediated alternative lengthening of telomeres. Telomeres associate with the nuclear envelope through the shelterin RAP1 and nuclear envelope SUN1 proteins. However, how the associations between telomeres and the nuclear envelope affect the progression of telomere recombination is not understood. Here, we show that telomere anchorage might inhibit telomere-telomere recombination. SUN1 depletion stimulates the formation of alternative lengthening of telomeres-associated promyelocytic leukemia bodies in ALT cells. In contrast, overexpression of a telomere-nuclear envelope-tethering chimera protein, RAP1-SUN1, suppresses APB formation. Moreover, inhibition of this nuclear envelope attachment alleviates the requirement of TOP3α for resolving the supercoiling pressure during telomere recombination. A coimmunoprecipitation assay revealed that the SUN1 N-terminal nucleoplasmic domain interacts with the RAP1 middle coil domain, and phosphorylation-mimetic mutations in RAP1 inhibit this interaction. However, abolishing the RAP1-SUN1 interaction does not hinder APB formation, which hints at the existence of another SUN1-dependent telomere anchorage pathway. In summary, our results reveal an inhibitory role of telomere-nuclear envelope association in telomere-telomere recombination and imply the presence of redundant pathways for the telomere-nuclear envelope association in ALT cells.

The Association Between Adverse Childhood Experiences and Child Telomere Length: Examining Self-Regulation as a Behavioral Mediator.

Childhood adversity is linked to shortened telomere length (TL), but behavioral indicators of telomere attrition remain unclear. This study examined the association between adverse childhood experiences (ACEs) and child TL, and if ACEs were indirectly associated with TL through children's self-regulatory abilities (i.e., effortful control and self-control). Hypotheses were tested using national data from teachers, parents, and their children (N = 2,527; Mage  = 9.35, SD = .36 years). More ACEs were uniquely associated with short TL, and low self-control mediated the association between more ACEs and short TL. While longitudinal studies are needed to strengthen claims of causation, this study identifies a pathway from ACEs to TL that should be explored further.

Telomere length and outcome of treatment for pulmonary tuberculosis in a gold mining community.

Telomere length (TL) is a marker of ageing and mitochondrial DNA (mtDNA) is an early marker of inflammation caused by oxidative stress. We determined TL and mtDNA content among active pulmonary tuberculosis (PTB) patients to assess if these cellular biomarkers differed between artisanal miners and non-miners, and to assess if they were predictive of treatment outcome. We conducted a prospective cohort study from August 2018 to May 2019 involving newly diagnosed PTB patients at three outpatient TB clinics in a rural Democratic Republic of Congo. We measured relative TL and mtDNA content in peripheral blood leukocytes (at inclusion) via qPCR and assessed their association with PTB treatment outcome. We included 129 patients (85 miners and 44 non-miners) with PTB (median age 40 years; range 5-71 years, 22% HIV-coinfected). For each increase in year and HIV-coinfection, TL shortened by - 0.85% (- 0.19 to - 0.52) (p ≤ 0.0001) and - 14% (- 28.22 to - 1.79) (p = 0.02) respectively. Independent of these covariates, patients with longer TL were more likely to have successful TB treatment [adjusted hazard ratio; 95% CI 1.27 for a doubling of leucocyte telomere length at baseline; 1.05-1.44] than patients with a shorter TL. Blood mtDNA content was not predictive for PTB outcome. For a given chronological age, PTB patients with longer telomeres at time of diagnosis were more likely to have successful PTB treatment outcome.

NHP2 downregulation counteracts hTR-mediated activation of the DNA damage response at ALT telomeres.

About 10% of cancer cells employ the "alternative lengthening of telomeres" (ALT) pathway instead of re-activating the hTERT subunit of human telomerase. The hTR RNA subunit is also abnormally silenced in some ALT+ cells not expressing hTERT, suggesting a possible negative non-canonical impact of hTR on ALT. Indeed, we show that ectopically expressed hTR reduces phosphorylation of ssDNA-binding protein RPA (p-RPAS33 ) at ALT telomeres by promoting the hnRNPA1- and DNA-PK-dependent depletion of RPA. The resulting defective ATR checkpoint signaling at telomeres impairs recruitment of the homologous recombination protein, RAD51. This induces ALT telomere fragility, increases POLD3-dependent C-circle production, and promotes the recruitment of the DNA damage marker 53BP1. In ALT+ cells that naturally retain hTR expression, NHP2 H/ACA ribonucleoprotein levels are downregulated, likely in order to restrain DNA damage response (DDR) activation at telomeres through reduced 53BP1 recruitment. This unexpected role of NHP2 is independent from hTR's non-canonical function in modulating telomeric p-RPAS33 . Collectively, our study shines new light on the interference between telomerase- and ALT-dependent pathways and unravels a crucial role for hTR and NHP2 in DDR regulation at ALT telomeres.

Telomerase as a Therapeutic Target in Cardiovascular Disease.

Shortened telomeres have been linked to numerous chronic diseases, most importantly coronary artery disease, but the underlying mechanisms remain ill defined. Loss-of-function mutations and deletions in telomerase both accelerate telomere shortening but do not necessarily lead to a clinical phenotype associated with atherosclerosis, questioning the causal role of telomere length in cardiac pathology. The differential extranuclear functions of the 2 main components of telomerase, telomerase reverse transcriptase and telomerase RNA component, offer important clues about the complex relationship between telomere length and cardiovascular pathology. In this review, we critically discuss relevant preclinical models, genetic disorders, and clinical studies to elucidate the impact of telomerase in cardiovascular disease and its potential role as a therapeutic target. We suggest that the antioxidative function of mitochondrial telomerase reverse transcriptase might be atheroprotective, making it a potential target for clinical trials. Graphic Abstract: A graphic abstract is available for this article.