Telomeres In Cancer Cells Research Articles

ZBTB48 is both a vertebrate telomere‐binding protein and a transcriptional activator

Telomeres constitute the ends of linear chromosomes and together with the shelterin complex form a structure essential for genome maintenance and stability. In addition to the constitutive binding of the shelterin complex, other direct, yet more transient interactions are mediated by the CST complex and HOT1/HMBOX1, while subtelomeric variant repeats are recognized by NR2C/F transcription factors. Recently, the Kruppel‐like zinc finger protein ZBTB48/HKR3/TZAP has been described as a novel telomere‐associated factor in the vertebrate lineage. Here, we show that ZBTB48 binds directly both to telomeric and to subtelomeric variant repeat sequences. ZBTB48 is found at telomeres of human cancer cells regardless of the mode of telomere maintenance and it acts as a negative regulator of telomere length. In addition to its telomeric function, we demonstrate through a combination of RNAseq, ChIPseq and expression proteomics experiments that ZBTB48 acts as a transcriptional activator on a small set of target genes, including mitochondrial fission process 1 (MTFP1). This discovery places ZBTB48 at the interface of telomere length regulation, transcriptional control and mitochondrial metabolism.

Knockdown of homeobox containing 1 increases the radiosensitivity of cervical cancer cells through telomere shortening.

Homeobox containing1 (HMBOX1) modulates telomere length in various types of tumor cells by binding to double‑stranded telomeric DNA. There is a negative correlation between telomere length and radiosensitivity in tumor cells. In the present study, we aimed to investigate the relationship among HMBOX1, telomere and radiosensitivity in cervical cancer cells. Lentivirus-based shRNAs were used to establish stable transfected cell lines in which protein and mRNA levels of HMBOX1 were notably decreased. Knockdown of HMBOX1 increased the radiosensitivity of HeLa and C33A cells. TERT protein was also decreased while HMBOX1 was downregulated. Knockdown of HMBOX1 shortened telomere length in the HeLa cells, while TERT overexpression rescued telomere shortening in the HeLa-HMBOX1 cells. Knockdown of HMBOX1 increased the apoptosis rate, decreased radiation-induced DNA damage foci, and inhibited the expression of ATM, ATR, p-ATM, p-ATR and BRCA1 in the homologous recombination repair pathway. Our data suggest a possible role of HMBOX1 in regulating radiosensitivity in cervical cancer cells. Moreover, HMBOX1 may be a potential factor in the radiotherapy of cervical cancer.

Proteomic Profiling Reveals a Specific Role for Translesion DNA Polymerase η in the Alternative Lengthening of Telomeres

SUMMARYCancer cells rely on the activation of telomerase or the alternative lengthening of telomeres (ALT) pathways for telomere maintenance and survival. ALT involves homologous recombination (HR)-dependent exchange and/or HR-associated synthesis of telomeric DNA. Utilizing proximity-dependent biotinylation (BioID), we sought to determine the proteome of telomeres in cancer cells that employ these distinct telomere elongation mechanisms. Our analysis reveals that multiple DNA repair networks converge at ALT telomeres. These include the specialized translesion DNA synthesis (TLS) proteins FANCJ-RAD18-PCNA and, most notably, DNA polymerase eta (Polη). We observe that the depletion of Polη leads to increased ALT activity and late DNA polymerase δ (Polδ)-dependent synthesis of telomeric DNA in mitosis. We propose that Polη fulfills an important role in managing replicative stress at ALT telomeres, maintaining telomere recombination at tolerable levels and stimulating DNA synthesis by Polδ.

Therapeutic Targeting of Telomerase.

Telomere length and cell function can be preserved by the human reverse transcriptase telomerase (hTERT), which synthesizes the new telomeric DNA from a RNA template, but is normally restricted to cells needing a high proliferative capacity, such as stem cells. Consequently, telomerase-based therapies to elongate short telomeres are developed, some of which have successfully reached the stage I in clinical trials. Telomerase is also permissive for tumorigenesis and 90% of all malignant tumors use telomerase to obtain immortality. Thus, reversal of telomerase upregulation in tumor cells is a potential strategy to treat cancer. Natural and small-molecule telomerase inhibitors, immunotherapeutic approaches, oligonucleotide inhibitors, and telomerase-directed gene therapy are useful treatment strategies. Telomerase is more widely expressed than any other tumor marker. The low expression in normal tissues, together with the longer telomeres in normal stem cells versus cancer cells, provides some degree of specificity with low risk of toxicity. However, long term telomerase inhibition may elicit negative effects in highly-proliferative cells which need telomerase for survival, and it may interfere with telomere-independent physiological functions. Moreover, only a few hTERT molecules are required to overcome senescence in cancer cells, and telomerase inhibition requires proliferating cells over a sufficient number of population doublings to induce tumor suppressive senescence. These limitations may explain the moderate success rates in many clinical studies. Despite extensive studies, only one vaccine and one telomerase antagonist are routinely used in clinical work. For complete eradication of all subpopulations of cancer cells a simultaneous targeting of several mechanisms will likely be needed. Possible technical improvements have been proposed including the development of more specific inhibitors, methods to increase the efficacy of vaccination methods, and personalized approaches. Telomerase activation and cell rejuvenation is successfully used in regenerative medicine for tissue engineering and reconstructive surgery. However, there are also a number of pitfalls in the treatment with telomerase activating procedures for the whole organism and for longer periods of time. Extended cell lifespan may accumulate rare genetic and epigenetic aberrations that can contribute to malignant transformation. Therefore, novel vector systems have been developed for a ‘mild’ integration of telomerase into the host genome and loss of the vector in rapidly-proliferating cells. It is currently unclear if this technique can also be used in human beings to treat chronic diseases, such as atherosclerosis.

Abstract LB-161: Exploiting non-canonical heterochromatin-mediated telomere protection mechanisms in human cells

Abstract Mammalian telomeres are inherently heterochromatic. While enhanced telomere maintenance is evident in malignant cancers, some cancers appear to maintain telomeres by neither the common telomerase nor the alternative telomere repeat recombination mechanisms. Specifically, the roles of epigenetic modifications in telomere protection are largely unknown in human cancers. I have combined newly developed cellular and molecular approaches to show that in some cancer cell types, experimentally enhanced heterochromatinization localized specifically at telomeres reduced damage-induced foci at telomeres, suggesting augmentation of telomere stability. These results lead to the intriguing hypothesis that manipulating the epigenetic status at telomeres may be exploited to elicit damage at the telomeres of cancer cells as a novel approach to fight cancer. My current work in progress focuses on identifying novel chromatin modifiers that weaken telomere protection by modulating telomere compaction. It is especially urgent to understand the plasticity and modes of telomere protection by which some human cancers escape the requirement of telomerase activation to sustain immortalization. Results from this work can potentially advance the development of novel combinatorial telomere-directed cancer treatments. Citation Format: Tracy T. Chow, Elizabeth H. Blackburn. Exploiting non-canonical heterochromatin-mediated telomere protection mechanisms in human cells. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr LB-161.

Telomere-Mitochondrion Links Contribute to Induction of Senescence in MCF-7 Cells after Carbon-Ion Irradiation.

The effects of carbon-ion irradiation on cancer cell telomere function have not been comprehensively studied. In our previous report cancer cells with telomere dysfunction were more sensitive to carbon-ion irradiation, but the underlying mechanisms remained unclear. Here we found that telomerase activity was suppressed by carbon-ion irradiation via hTERT down-regulation. Inhibition of telomere activity by MST-312 further increased cancer cell radiosensitivity to carbon-ion radiation. hTERT suppression caused by either carbon-ion irradiation or MST-312 impaired mitochondrial function, as indicated by decreased membrane potential, mtDNA copy number, mitochondrial mass, total ATP levels and elevated reactive oxygen species (ROS). PGC-1α expression was repressed after carbion-ion irradiation, and hTERT inhibition by MST-312 could further exacerbate this effect. Lowering the mitochondrial ROS level by MitoTEMPO could partially counteract the induction of cellular senescence induced by carbon-ion radiation and MST-312 incubation. Taken together, the current data suggest that telomere-mitochondrion links play a role in the induction of senescence in MCF-7 cells after carbon-ion irradiation.

Abstract 3870: Telomeric non-coding RNA causes genome-wide alteration of gene expression in cancer

Abstract The end replication problem of linear chromosome DNA causes telomere shortening and eventual senescence of normal cells. Cancer cells reactivate telomerase to maintain their telomere length and exhibit an infinite replicative potential. However, telomerase activation does not necessarily increase the average telomere length in cancer cells. Actually, cancer cells often maintain shorter telomeres than the cells in the surrounding normal tissues. We previously demonstrated that forced telomere elongation in cancer cells promotes their differentiation in a xenograft tumor and represses innate immune genes that are upregulated in various human cancers. This implies that short telomeres of cancer cells contribute to their malignancy. However, it is completely unknown how such genetic repression is caused by elongated telomeres. Here, we report that telomeric repeat-containing RNA (TERRA) induces a genome-wide alteration of gene expression in telomere-elongated human cancer cells. Using three different cancer cell lines, including those from prostate, breast and stomach, we found that telomere elongation by hTERT overexpression up-regulates TERRA signals, which were detected by northern blot analysis, and down-regulates innate immune genes such as STAT1, ISG15, and OAS3 in vivo. These changes were caused by telomere elongation and not by enhanced telomerase activity or hTERT overexpression per se. Down-regulation of the innate immune genes was also observed in the telomere-elongated cells under three-dimensional (3D) culture conditions, which mimicked the tumor microenvironment in vivo. Introduction of TERRA oligonucleotides repressed the innate immune genes even in cells with short telomeres under 3D culture conditions. Intriguingly, this gene repression appeared to occur from the action of the non-canonical nucleic acid structure called G-quadruplex (G4) in TERRA, because non-telomeric (i.e., non-G4-forming) control oligonucleotides had no effect whereas another non-telomeric but G4-forming oligonucleotide phenocopied the TERRA oligonucleotides. Transcriptome analysis revealed that telomere elongation and G4-forming oligonucleotides showed similar gene expression signatures. Most of the commonly suppressed genes were involved in the innate immune system and were up-regulated in various human cancers. Taken together, these observations suggest that the telomeric non-coding RNA G4 counteracts cancer malignancy through suppression of innate immune genes, and give a new insight into our understanding of telomere shortening in human cancer. Citation Format: Kyotaro Hirashima, Hiroyuki Seimiya. Telomeric non-coding RNA causes genome-wide alteration of gene expression in cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3870. doi:10.1158/1538-7445.AM2015-3870

Prediagnostic Obesity and Physical Inactivity Are Associated with Shorter Telomere Length in Prostate Stromal Cells

Obesity and inactivity have been associated with advanced-stage prostate cancer, and poor prostate cancer outcomes, though the underlying mechanism(s) is unknown. To determine whether telomere shortening, which has been associated with lethal prostate cancer, may be a potential underlying mechanism, we prospectively evaluated the association between measures of adiposity, physical activity, and telomere length in 596 participants in the Health Professionals Follow-up Study, who were surgically treated for prostate cancer. Using tissue microarrays, we measured telomere length in cancer and benign cells using a telomere-specific FISH assay. Adiposity and activity were assessed via questionnaire within 2 years of diagnosis. Adjusting for age, pathologic stage, and grade, the median and SD of the per cell telomere signals were determined for each man for stromal cells and cancer cells by adiposity and activity categories. Overweight/obese men (54%) were similar to normal weight men on most factors, but had higher Gleason sum and lower activity levels. Overweight/obese men had 7.4% shorter telomeres in stromal cells than normal weight men (P = 0.06). The least active men had shorter telomeres in stromal cells than more active men (Ptrend = 0.002). Men who were overweight/obese and the least active had the shortest telomeres in stromal cells (20.7% shorter; P = 0.0005) compared with normal weight men who were the most active. Cancer cell telomere length and telomere length variability did not differ by measures of adiposity or activity. Telomere shortening in prostate cells may be one mechanism through which lifestyle influences prostate cancer risk and outcomes.

Telomere shortening in breast cancer correlates with the pathological features of tumor progression.

Telomeres are involved in the maintenance of genomic stability. Telomere alteration has been observed in most human cancer types, and is known to be a feature of malignancy. The aim of the present study was to evaluate whether the telomere length of breast cancer cells correlates with TNM stage and several pathological features. We investigated a total of 44 breast cancers, including 17 scirrhous, 15 papillotubular and 12 solid-tubular carcinomas. Telomere lengths were determined by tissue quantitative fluorescence in situ hybridization (Q-FISH), and compared according to the TNM stage, histological tumor size, lymph node metastases, vascular invasion and immunohistochemical status (ER, PR, HER2 status and Ki67 labeling index). In all histological types, telomeres of cancer cells were significantly shorter than those of normal epithelial cells. Mean telomere length was significantly less in patients with TNM stage III, and in those with large tumors, lymph node metastases and vascular invasion. Our results suggest that the telomere length of cancer cells is strongly correlated with the degree of cancer progression.

Prostate stromal cell telomere shortening is associated with risk of prostate cancer in the placebo arm of the Prostate Cancer Prevention Trial

Telomeres are repetitive nucleoproteins that help maintain chromosomal stability by inhibiting exonucleolytic degradation, prohibiting inappropriate homologous recombination, and preventing chromosomal fusions by suppressing double-strand break signals. We recently observed that men treated for clinically localized prostate cancer with shorter telomeres in their cancer-associated stromal cells, in combination with greater variation in cancer cell telomere lengths, were significantly more likely to progress to distant metastases, and die from their disease. Here, we hypothesized that shorter stromal cell telomere length would be associated with prostate cancer risk at time of biopsy. Telomere-specific fluorescence in situ hybridization (FISH) analysis was performed in normal-appearing stromal, basal epithelial, and luminal epithelial cells in biopsies from men randomized to the placebo arm of the Prostate Cancer Prevention Trial. Prostate cancer cases (N = 32) were either detected on a biopsy performed for cause or at the end of the study per trial protocol, and controls (N = 50), defined as negative for cancer on an end-of-study biopsy performed per trial protocol (e.g., irrespective of indication), were sampled. Logistic regression was used to estimate the association between mean telomere length of the particular cell populations, cell-to-cell telomere length variability, and risk of prostate cancer. Men with short stromal cell telomere lengths (below median) had 2.66 (95% CI 1.04-3.06; P = 0.04) times the odds of prostate cancer compared with men who had longer lengths (at or above median). Conversely, we did not observe statistically significant associations for short telomere lengths in normal-appearing basal (OR = 2.15, 95% CI 0.86-5.39; P= 0 .10) or luminal (OR = 1.15, 95% CI 0.47-2.80; P = 0.77) cells. These findings suggest that telomere shortening in normal stromal cells is associated with prostate cancer risk. It is essential to extend and validate these findings, while also identifying the cellular milieu that comprises the subset of cells with short telomeres within the prostate tumor microenvironment.

Telomeric repeat-containing RNA/G-quadruplex-forming sequences cause genome-wide alteration of gene expression in human cancer cells in vivo.

Telomere erosion causes cell mortality, suggesting that longer telomeres enable more cell divisions. In telomerase-positive human cancer cells, however, telomeres are often kept shorter than those of surrounding normal tissues. Recently, we showed that cancer cell telomere elongation represses innate immune genes and promotes their differentiation in vivo. This implies that short telomeres contribute to cancer malignancy, but it is unclear how such genetic repression is caused by elongated telomeres. Here, we report that telomeric repeat-containing RNA (TERRA) induces a genome-wide alteration of gene expression in telomere-elongated cancer cells. Using three different cell lines, we found that telomere elongation up-regulates TERRA signal and down-regulates innate immune genes such as STAT1, ISG15 and OAS3 in vivo. Ectopic TERRA oligonucleotides repressed these genes even in cells with short telomeres under three-dimensional culture conditions. This appeared to occur from the action of G-quadruplexes (G4) in TERRA, because control oligonucleotides had no effect and a nontelomeric G4-forming oligonucleotide phenocopied the TERRA oligonucleotide. Telomere elongation and G4-forming oligonucleotides showed similar gene expression signatures. Most of the commonly suppressed genes were involved in the innate immune system and were up-regulated in various cancers. We propose that TERRA G4 counteracts cancer malignancy by suppressing innate immune genes.

The effect of chemotherapeutic agents on telomere length maintenance in breast cancer cell lines.

Mammalian telomeric DNA consists of tandem repeats of the sequence TTAGGG associated with a specialized set of proteins, known collectively as Shelterin. These telosomal proteins protect the ends of chromosomes against end-to-end fusion and degradation. Short telomeres in breast cancer cells confer telomere dysfunction and this can be related to Shelterin proteins and their level of expression in breast cancer cell lines. This study investigates whether expression of Shelterin and Shelterin-associated proteins are altered, and influence the protection and maintenance of telomeres, in breast cancer cells. 5-aza-2′-deoxycytidine (5-aza-CdR) and trichostatin A (TSA) were used in an attempt to reactivate the expression of silenced genes. Our studies have shown that Shelterin and Shelterin-associated genes were down-regulated in breast cancer cell lines; this may be due to epigenetic modification of DNA as the promoter region of POT1 was found to be partially methylated. Shelterin genes expression was up-regulated upon treatment of 21NT breast cancer cells with 5-aza-CdR and TSA. The telomere length of treated 21NT cells was measured by q-PCR showed an increase in telomere length at different time points. Our studies have shown that down-regulation of Shelterin genes is partially due to methylation in some epithelial breast cancer cell lines. Removal of epigenetic silencing results in up-regulation of Shelterin and Shelterin-associated genes which can then lead to telomere length elongation and stability.Electronic supplementary materialThe online version of this article (doi:10.1007/s10549-014-2975-x) contains supplementary material, which is available to authorized users.

G-四链体DNA稳定剂的研究进展

Human telomere consists of a series of DNA sequences rich in Guanine, which may form G-quadruplex structure. The formation and stabilization of the specific structure by small molecular compounds can not only inhibit the telomerase and disrupt the mechanism of the maintenance of telomere in cancer cells but also work as a probe for G-quedruplex on the research of the biological functions of G-quadruplex and diagnosis of relevant diseases. So the designing of stabilizers of G-quadruplex has been one of the focuses of chemical biology in recent years. The G-quadruplex stabilizers can be divided in two types up to now: small organic molecular compounds and metal complexes. This review focuses on the latest progress of the two parts, especially the latter one.

Abstract PR06: Prediagnostic obesity and inactivity are associated with shorter telomere length in prostate cancer-associated stromal cells

Abstract Background: Meta-analyses and some cohort studies support that obese men are more likely to develop high-grade and advanced stage prostate cancer, recur after treatment, and die of this cancer. Physical activity, independent of body mass index, has been inversely associated with the development of advanced prostate cancer in some prospective studies. Mechanisms underlying these observations remain unknown. Previously, we showed that shorter telomere length in prostate-cancer associated stromal cells, especially in combination with greater variability in telomere length in cancer cells, was associated with a significantly increased risk of prostate cancer death after prostatectomy. Given this finding and because telomeres shorten with each round of replication, we hypothesized that obesity-associated growth factors influence telomere length, and might explain the more aggressive prostate cancer phenotype in obese men. Methods: We conducted a cross-sectional study of 596 men surgically treated for prostate cancer who participated in the Health Professionals Follow-up Study. Tissue microarrays containing areas of adenocarcinoma and benign tissue were stained using a telomere-specific FISH probe and using DAPI for total nuclear DNA. Image analysis was used to quantify telomeric signals in individual cancer and benign cells. Body mass index (BMI) and total physical activity were collected via questionnaire 2-years before the diagnosis. BMI was categorized into normal (≥25 kg/m2) and overweight/obese (>25 kg/m2). Total physical activity was categorized into tertiles. Adjusting for pathologic stage, grade and other factors, median and standard deviation of the telomere signal normalized to DAPI were determined for each man for cancer cells and cancer-associated stromal cells by categories of BMI and physical activity. Results: Overweight/obese men (54%) were similar to normal weight men on demographics and pathologic stage, but had higher Gleason sum and were less active than normal weight men. Overweight/obese men had shorter mean telomere length in cancer-associated stromal cells (7.5% shorter) than normal weight men (p=0.055). The least active men also had shorter mean telomere length in cancer-associated stromal cells than more active men (p-trend=0.004). Men who were overweight/obese and the least active had the shortest mean telomere length in cancer-associated stromal cells (20.7% shorter) compared to normal weight men who were the most active, who had the longest mean telomere length (p=0.0005). Cancer cell telomere length variability did not differ by BMI or activity level. Conclusions: Increased weight and decreased activity were associated with shorter mean telomere length in cancer-associated stromal cells. Telomere shortening in prostate cells may be one mechanism through which lifestyle influences prostate cancer risk and outcomes. Funding: Prostate Cancer Foundation, DOD, NIH/NCI, Seraph Foundation This abstract is also presented as Poster B43. Citation Format: Corinne E. Joshu, Sarah B. Peskoe, Christopher M. Heaphy, Stacey A. Kenfield, Lorelei A. Mucci, Edward Giovanucci, Meir J. Stampfer, Ghil Suk Yoon, Thomas Lee, Jessica L. Hicks, Angelo M. De Marzo, Alan K. Meeker, Elizabeth A. Platz. Prediagnostic obesity and inactivity are associated with shorter telomere length in prostate cancer-associated stromal cells. [abstract]. In: Proceedings of the Twelfth Annual AACR International Conference on Frontiers in Cancer Prevention Research; 2013 Oct 27-30; National Harbor, MD. Philadelphia (PA): AACR; Can Prev Res 2013;6(11 Suppl): Abstract nr PR06.

Telomeric impact of conventional chemotherapy

The increased level of chromosome instability in cancer cells, leading to aneuploidy and gross chromosomal rearrangements, is not only a driving force for oncogenesis but also can be the Achille's heel of the disease since many chemotherapies (CT) kill cells by inducing a non-tolerable rate of DNA damage. A wealth of published evidence showed that telomere stability can be more affected than the bulk of the genome by several conventional antineoplasic drugs. These results raise the interesting possibility that CT with genotoxic drugs preferentially target telomeres. In agreement with this view, accelerated shortening of telomere length has been described in blood lineage cells following high-dose CT (stem cell transplantation) or non-myeloablative CT. However, almost nothing is known on the consequences of this shortening in terms of telomere stability, senescence and on the development of second cancers or post-treatment aging-like syndromes in cancer survivors (cognitive defect, fertility impairment, etc.). In this article, we propose: (1) telomeres of cancer cells are preferential genomic targets of chemotherapies altering chromosome maintenance; (2) telomere functional parameters can be a surrogate marker of chemotherapy sensitivity and toxicity; (3) the use of anti-telomere molecule could greatly enhance the sensitivity to standards chemotherapies.

Prostate Cancer Cell Telomere Length Variability and Stromal Cell Telomere Length as Prognostic Markers for Metastasis and Death

Current prognostic indicators are imperfect predictors of outcome in men with clinically localized prostate cancer. Thus, tissue-based markers are urgently needed to improve treatment and surveillance decision-making. Given that shortened telomeres enhance chromosomal instability and such instability is a hallmark of metastatic lesions, we hypothesized that alterations in telomere length in the primary cancer would predict risk of progression to metastasis and prostate cancer death. To test this hypothesis, we conducted a prospective cohort study of 596 surgically treated men who participated in the ongoing Health Professionals Follow-up Study. Men who had the combination of more variable telomere length among prostate cancer cells (cell-to-cell) and shorter telomere length in prostate cancer-associated stromal (CAS) cells were substantially more likely to progress to metastasis or die of their prostate cancer. These findings point to the translational potential of this telomere biomarker for prognostication and risk stratification for individualized therapeutic and surveillance strategies. In this prospective study, the combination of more variable telomere length among cancer cells and shorter telomere length in CAS cells was strongly associated with progression to metastasis and prostate cancer death, pointing to the translational potential for prognostication and risk stratifi cation for individualized therapeutic and surveillance strategies.

Telomere Length Influences Cancer Cell Differentiation In Vivo

Limitless reproductive potential is one of the hallmarks of cancer cells. This ability is due to the maintenance of telomeres, erosion of which causes cellular senescence or death. While most cancer cells activate telomerase, a telomere-elongating enzyme, it remains elusive as to why cancer cells often maintain shorter telomeres than the cells in the surrounding normal tissues. Here, we show that forced telomere elongation in cancer cells promotes their differentiation in vivo. We elongated the telomeres of human prostate cancer cells that possess short telomeres by enhancing their telomerase activity. The resulting cells had long telomeres and retained the ability to form tumors in nude mice. Strikingly, these tumors exhibited many duct-like structures and reduced N-cadherin expression, reminiscent of well-differentiated adenocarcinoma. These changes were caused by telomere elongation and not by enhanced telomerase activity. Gene expression profiling revealed that tumor formation was accompanied by the expression of innate immune system-related genes, which have been implicated in maintaining tumor cells in an undifferentiated state and poor-prognosis cancers. In tumors derived from the telomere-elongated cells, upregulation of such gene sets is not observed. Our observations suggest a functional contribution of short telomeres to tumor malignancy by regulation of cancer cell differentiation.

PML bodies provide an important platform for the maintenance of telomeric chromatin integrity in embryonic stem cells

We have previously shown that α-thalassemia mental retardation X-linked (ATRX) and histone H3.3 are key regulators of telomeric chromatin in mouse embryonic stem cells. The function of ATRX and H3.3 in the maintenance of telomere chromatin integrity is further demonstrated by recent studies that show the strong association of ATRX/H3.3 mutations with alternative lengthening of telomeres in telomerase-negative human cancer cells. Here, we demonstrate that ATRX and H3.3 co-localize with the telomeric DNA and associated proteins within the promyelocytic leukemia (PML) bodies in mouse ES cells. The assembly of these telomere-associated PML bodies is most prominent at S phase. RNA interference (RNAi)-mediated knockdown of PML expression induces the disassembly of these nuclear bodies and a telomere dysfunction phenotype in mouse ES cells. Loss of function of PML bodies in mouse ES cells also disrupts binding of ATRX/H3.3 and proper establishment of histone methylation pattern at the telomere. Our study demonstrates that PML bodies act as epigenetic regulators by serving as platforms for the assembly of the telomeric chromatin to ensure a faithful inheritance of epigenetic information at the telomere.

Structural and functional association of androgen receptor with telomeres in prostate cancer cells

Telomeres protect the ends of linear chromosomes from being recognized as damaged DNA, and telomere stability is required for genome stability. Here we demonstrate that telomere stability in androgen receptor (AR)-positive LNCaP human prostate cancer cells is dependent on AR and androgen, as AR inactivation by AR antagonist bicalutamide (Casodex), AR-knockdown, or androgen-depletion caused telomere dysfunction, and the effect of androgen-depletion or Casodex was blocked by the addition of androgen. Notably, neither actinomycin D nor cycloheximide blocked the DNA damage response to Casodex, indicating that the role of AR in telomere stability is independent of its role in transcription. We also demonstrate that AR is a component of telomeres, as AR-bound chromatin contains telomeric DNA, and telomeric chromatin contains AR. Importantly, AR inactivation by Casodex caused telomere aberrations, including multiple abnormal telomere signals, remindful of a fragile telomere phenotype that has been described previously to result from defective telomere DNA replication. We suggest that AR plays an important role in telomere stability and replication of telomere DNA in prostate cancer cells, and that AR inactivation-mediated telomere dysfunction may contribute to genomic instability and progression of prostate cancer cells.

Acacetin and Chrysin, Two Polyphenolic Compounds, Alleviate Telomeric Position Effect in Human Cells

We took advantage of the ability of human telomeres to silence neighboring genes (telomere position effect or TPE) to design a high-throughput screening assay for drugs altering telomeres. We identified, for the first time, that two dietary flavones, acacetin and chrysin, are able to specifically alleviate TPE in human cells. We further investigated their influence on telomere integrity and showed that both drugs drastically deprotect telomeres against DNA damage response. However, telomere deprotection triggered by shelterin dysfunction does not affect TPE, indicating that acacetin and chrysin target several functions of telomeres. These results show that TPE-based screening assays represent valuable methods to discover new compounds targeting telomeres.