Dissertations / Theses: 'Telomeren'

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Genest, Paul-André Joseph Jean. "Analysis of the modified DNA base J and the J-binding proteins in Leishmania." Amsterdam : Amsterdam : Nederlands Kanker Instituut / Antoni Van Leeuwenhoekziekenhuis ; Universiteit van Amsterdam [Host], 2007. http://dare.uva.nl/document/47968.

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Nijjar, Tarlochan Singh. "Molecular characterization of steps involved in immortal transformation of human mammary epithelial cells." [S.l. : Amsterdam : s.n.] ; Universiteit van Amsterdam [Host], 2003. http://dare.uva.nl/document/87095.

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Shakirov, Yevgeniy Vitalievich. "Telomeres and telomere binding proteins in Arabidopsis thaliana." Diss., Texas A&M University, 2004. http://hdl.handle.net/1969.1/422.

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Telomeres are important protein-DNA structures at the ends of linear eukaryotic chromosomes that are necessary to prevent chromosome fusions and exonuclease attack. We found that telomere tracts in Arabidopsis are fairly uniformly distributed throughout a size range of 2-9kb. Unexpectedly, telomeres in WS plants displayed a bimodal size distribution with some individuals exhibiting 4-8 kb telomeres and others 2-5 kb telomeres. We also examined the dynamics of telomere tracts on individual chromosome ends. Following the fate of telomeres in plants through successive generations, we found that the shortest telomeres were typically elongated in the subsequent generation, while the longest telomeres were usually shortened. Thus, telomere length homoeostasis is achieved through intermittent telomerase action on shorter telomeres to attain an optimal size.Single-strand telomere binding proteins were also analyzed. Four major telomere binding protein complexes from cauliflower were identified and their DNA-binding properties characterized. The DNA-binding component of one of the complexes was purified and analyzed by mass-spectrometry. Peptide mass data was used to search for putative protein candidates from the Arabidopsis thaliana database. Additionally, two Arabidopsis genes, AtPot1 and AtPot2, were identified and characterized. The genes encode two single-strand telomeric DNA binding proteins. AtPot1 and AtPot2 proteins can homo- and heterodimerize in vitro. Pot1 protein predominantly localizes to the nucleolus, whereas Pot2 is exclusively nuclear. Plants over-expressing full-length Pot1 and Pot2 proteins had no obvious phenotype, while over-expression of P2DBD and P1∆DBD caused moderate telomere shortening. Plants over-expressing P2DBD had severe morphological and reproductive defects, multiple chromosome abnormalities and aneuploidy. Over-expression of a chimeric protein DBD-P1∆DBD led to rapid telomere shortening, confirming the involvement of Arabidopsis Pot proteins in telomere length maintenance. Intriguingly, telomerase in DBD-P1∆DBD-EYFP plants is inactivated, suggesting that Pot proteins are also involved in regulation of telomerase activity. The analysis of Arabidopsis telomeres and telomere binding proteins will provide additional information towards understanding the role of the telomeric nucleoprotein complex in eukaryotic chromosome biology.

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Schulze, Franziska. "Die Telomerlänge als Prognosefaktor in MYCN nicht-amplifizierten Neuroblastomen." Doctoral thesis, Universitätsbibliothek Leipzig, 2016. http://nbn-resolving.de/urn:nbn:de:bsz:15-qucosa-200943.

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Eines der charakteristischen Merkmale des Neuroblastoms stellt seine einzigartige biologische Heterogenität dar, die eine genaue Ausage des weiteren klinischen Verlaufes stark erschwert. Bestimmte prognostisch wirksame klinische, molekularbiologische und genetische Faktoren, wie zum Beispiel Alter bei Erstdiagnose, Tumorstadium, MYCN-Amplifikation und 1p Deletion, werden seit längerem zur Risikostratifizierung genutzt. Bereits in anderen Tumorerkrankungen konnte nun der Einfluß einer Telomerlängenveränderung auf das Gesamtüberleben von Patienten nachgewiesen werden. Telomere sichern die genomische Integrität und bestimmen maßgeblich die proliferative Kapazität jeder somatischen Zelle. Aktuelle Forschungsergebnisse legen die Vermutung nahe, dass Veränderungen der Telomerlänge auch in Neuroblastomen einen prognostischen Effekt auf das Gesamtüberleben haben. In diesem Kontext untersucht die vorliegende Arbeit den Zusammenhang zwischen Telomerlänge und Gesamtüberleben in 420 MYCN nicht-amplifizierten primären Neuroblastomen mit Erstdiagnosen von 1983-2001. Hierfür wurden die relativen Telomerlängen mithilfe einer neu etablierten monochromen multiplex q-RT-PCR ermittelt. Anschließend wurden diese sowohl mit ausgesuchten klinischen Variablen (Alter bei Erstdiagnose, Tumorstadium, Primärlokalisation des Tumors, Histologie, Geschlecht und Rezidivauftreten) korreliert als auch auf ihren Einfluß auf das Gesamt- und ereignisfreie Überleben untersucht. In Korrelation mit den klinischen Parametern konnte zwischen Alter bei Erstdiagnose und Telomerlänge ein eindeutiger Zusammenhang nachgewiesen werden. Je älter die Patienten bei Erstdiagnose, desto höher war sowohl der Anteil verlängerter Telomere als auch der extremer Telomerlängenveränderungen. Neuroblastome mit verlängerten Telomeren zeigten in der gleichen Altersgruppe ein verringertes Gesamtüberleben der betroffenen Patienten verglichen mit Neuroblastomen mit verkürzten Telomeren. Somit könnte eine Telomerlängenveränderung, insbesondere verlängerte Telomere, im klinischen Alltag als Hinweis auf einen prognostisch ungünstigen Verlauf genutzt werden.

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Marzec, Paulina. "NR2C/F telomeric association drives telomere-genome rearrangements in ALT cells." Thesis, Montpellier 2, 2013. http://www.theses.fr/2013MON20179.

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L'immortalité cellulaire est toujours accompagnée par l'activation du mécanisme de maintien des télomères. Dans la plupart des cancers humains, ce rôle est assuré par l'enzyme télomérase. Cependant, dans 15 % des tumeurs, la télomérase n'est pas activée et les télomères sont maintenus par l'allongement alternatif des télomères (ALT), voie qui implique la recombinaison des télomères. ALT est plus fréquent dans les tumeurs provenant de tissus mésenchymateux (sarcomes), representant 40-60 % des cas, que dans les tumeurs épithéliales. Comprendre le mécanisme ALT est primordial dans les thérapies anti-cancéreuses puisque certaines drogues inhibant la télomérase conduisent souvent à l'activation de l'ALT.La voie ALT est définie par de caractéristiques typiques des télomères. Dans les cellules ALT, les recombinaisons aberrantes d'ADN ne se limitent pas aux télomères puisque les génomes sont souvent fortement réarrangés. Les liens de ces caractéristiques génomiques anormales et la maintenance des télomères atypique ne sont pas connues, mais l'instabilité du génome contribue certainement à la transformation. Notre équipe a montré que les récepteurs orphelins appartenant aux familles NR2C/F ont été trouvés enrichies dans les télomères des lignées cellulaires ALT. Nous avons proposé que ces facteurs puissent être recrutés aux télomères par liaison directe à la séquence répétée GGGTCA, un site de liaison à haute affinité pour ces protéines. Mon projet vise à comprendre (i) leur mécanisme de liaison et (ii) leur rôle, dans le processus d'ALT.Dans cette étude nous montrons que dans les sarcomes primaires humains, les télomères d'ALT sont souvent liés par des récepteurs nucléaires orphelins des sous-familles NR2C/F, en particulier dans les tumeurs au stade avancé. Ceci suggère un rôle actif de ces facteurs dans la progression tumorale ALT. En utilisant la technique de ChIP-sequencing, nous avons montré que les protéines NR2C/F se lient à une répétition directe amplifiée (DR0) aux télomères, et pas de manière significative à toute autre combinaison de motif GGGTCA. Nous avons également analysé la distribution sur tout le génome de NR2C2/F2 et TRF2, une protéine de liaison des télomères, dans des cellules ALT (-) et ALT (+). Bien qu'il n'y ait que peu de sites génomiques liés par TRF2 dans les cellules ALT (-), nous avons été surpris d'identifier plusieurs centaines de régions liées par TRF2 dans les cellules ALT (+). Plus surprenant, la grande majorité de ces régions spécifiques TRF2 ALT chevauche des sites endogènes de NR2C2/F2. Étant donné que ces sites ne contiennent généralement pas les répétitions des télomères, TRF2 est probablement recruté de façon indirecte. Conformément à cette interprétation, nous montrons que les facteurs NR2C/F entrainent un rapprochement des loci et sont responsables du regroupement atypique des télomeres dans ALT. De plus, un sous-ensemble de ces régions génomiques uniques a des additions hétérogènes des séquences télomeriques ALT, suggérant un rôle dans le recrutement des télomères par des protéines NR2C/F mais aussi une fonction de ciblage de recombinaison génomique. Systématiquement, nous trouvons que ces réarrangements des télomères/génome sont situés à proximité des motifs GGGTCA endogènes. Le caryotype spectral des lignées cellulaires ATL montre que les sites télomériques interstitielles sont fréquemment localisés aux niveaux des sites de translocations/réarrangements entre deux ou plusieurs chromosomes, ce qui est également observé dans les données de ChIPseq. Ces résultats suggèrent que les réarrangements entres les télomères et le génome pourraient participer à la formation d'un caryotype complexe ce qui caractérise environ 50% des sarcomes. De plus, l'addition de sites télomériques interstitielles dans le génome est spécifique des cellules ALT et est favorisée par les dommages de l'ADN
Cellular immortality is always accompanied by the activation of telomere maintenance mechanism. In most human cancers this role is fulfilled by the telomerase enzyme. However in 15% of tumors, telomerase is not activated and telomeres are maintained by an Alternative Lengthening of Telomeres (ALT) pathway that involves telomere-telomere recombination. Interestingly ALT is more prevalent in tumors originating from mesenchymal tissues (sarcomas), where it is present in 40-60% of cases, than in epithelial tumors. Understanding ALT maintenance is critical since inhibiting telomerase in tumors leads to the activation of ALT. The ALT pathway is operationally defined by typical telomere hallmarks. In ALT cells, aberrant DNA transactions are not restricted to telomeres since genomes are often highly rearranged. Whether these abnormal genomic features are linked to atypical telomere maintenance is not known, but genome instability is certainly contributing to transformation. We have previously shown that orphan receptors of the NR2C/F families were enriched at telomeres in ALT cell lines. We proposed that these factors could be recruited to telomeres through direct binding to the GGGTCA variant repeat, a high affinity binding site for these proteins. My project is aimed at understanding (i) their mechanism of binding and (ii) their role, if any, in the ALT process.We show that in human primary sarcomas, ALT telomeres are often bound by orphan nuclear receptors of the NR2C/F subfamilies, particularly in more advanced-stage tumors. This suggests an active role for these factors in ALT tumor progression. Using ChIP-sequencing, we show that NR2C/F proteins bind to an amplified direct repeat (DR0) at telomeres, and not significantly to any other GGGTCA motif combination. We also analyzed the genome wide distribution of NR2C2/F2 and TRF2, a telomere binding protein, in ALT(-) and in ALT(+) cells. While there are only few genomic sites bound by TRF2 in ALT(-) cells, we were surprised to identify several hundred regions bound by TRF2 in ALT(+) cells. More surprisingly, the great majority of these ALT specific TRF2 regions overlap with endogenous NR2C2/F2 sites. Since these sites usually do not contain telomere repeats, TRF2 is likely indirectly recruited. Consistent with this interpretation, we show that NR2C/F factors drive locus proximity. Moreover, a subset of these unique genomic regions harbor heterogeneous ALT telomere sequence additions, not only suggesting a telomere recruitment role for NR2C/F proteins but also a recombination targeting function in the genome. Consistently, we find these telomere/genome rearrangements are located close to endogenous GGGTCA motifs. Next, we wanted to evaluate a role of these rearrangements in formation of complex karyotype which characterize approximately 50% of sarcomas. We found by spectral karyotyping that interstitial telomeric sites are frequently located at translocation/ rearrangements sites between two or more chromosomes, which we could also observe in our ChIPseq data. Furthermore, we demonstrate that addition of interstitial telomeric sites to the genome is enhanced by DNA damage and specific for ALT genome. Therefore we conclude that NR2C/F factors target telomere proximity to defined NR2C/F regions which enables telomere-genome rearrangements under DNA damage condition. This contributes not only to efficient telomere recombination, but also it drives further genomic instability at selected NR2C/F sites.We believe we identified a new mechanism of telomere dysfunction potentially driving targeted genome instability and mediated by NR2C/F proteins in ALT cells which probably underlie complexity of sarcomas genome. Understanding the ALT mechanism allows designing NR2C/F-targeted therapies in treatment of ALT tumors and therapies for patients treated with anti-telomerase drugs to prevent ALT appearance

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Henson, Jeremy D. "The role of Alternative Lengthening of Telomeres in human cancer." University of Sydney, 2006. http://hdl.handle.net/2123/1533.

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Doctor of Philosophy
Activation of a telomere maintenance mechanism is a vital step in the development of most cancers and provides a target for the selective killing of cancer cells. Cancers can use either telomerase or Alternative Lengthening of Telomeres (ALT) to maintain their telomeres and inhibition of either telomere maintenance mechanism can cause cancer cells to undergo senescence or apoptosis. Although telomerase inhibitors are undergoing clinical trials, on commencing this study very little was known about the role of ALT in cancer, what proteins were involved in its mechanism and regulation and how it could be targeted clinically. The primary aim of this thesis was to develop an assay for ALT suitable for examining archived tumour specimens and to begin using it to examine the prevalence and clinical significance of ALT in cancer. This assay and gene expression analysis was also used to identify genes that are involved in or associated with the activation of the ALT mechanism, to contribute towards the overall goal of an ALT cancer therapy. The ALT mechanism involves recombination mediated replication and ALT cells have a marked increase in a range of recombinational events specifically at their telomeres. Presumably, as a consequence of this the telomere lengths of ALT cells are very heterogeneous and on average long. This can be detected by terminal restriction fragment (TRF) Southern analysis, which has been used previously as the definitive test for ALT activity. However, TRF analysis requires intact genomic DNA and is unsuitable for tumour specimens which are commonly archived by paraffin embedding. Another hallmark of ALT is ALT-associated PML bodies (APBs) which are the subset of PML bodies that contain telomeric DNA. Work done in this study to consolidate APBs as a hallmark of ALT, combined with published data, showed 29/31 ALT[+], 3/31 telomerase[+] and 0/10 mortal cell lines/strains are APB[+]. The three APB[+]/telomerase[+] cell lines identified here had an order of magnitude lower frequency of APB[+] nuclei than the ALT[+] cell lines. APBs may be functionally linked to the ALT mechanism and contain the recombination proteins that are thought to be involved in the ALT mechanism. This study, in collaboration with Dr W-Q Jiang, strengthened this functional link by demonstrating that loss of ALT activity (as determined by TRF analysis) coincided with the disruption of APBs. The detection of APBs was developed into a robust assay for ALT in archived tumour specimens using a technique of combined immunofluorescence and telomere fluorescence in situ hybridisation. It was demonstrated that the APB assay concurred exactly with the standard assay for ALT (TRF analysis) in 60 tumours for which TRF analysis gave unequivocal results. The APB assay may be a more appropriate technique in the case of tumour specimen heterogeneity, which may explain why the APB assay was able to give definitive results when TRF analysis was equivocal. We demonstrated that intratumoral heterogeneity for ALT does exist and this could explain why about 3% of tumours in this study were APB[+] but with more than a ten-fold reduction in the frequency of APB[+] nuclei. This study also made the novel discovery of single stranded C-rich telomeric DNA inside APBs which potentially could be used to make the APB assay more suitable for routine pathology laboratory use. The APB assay was used to show that ALT is a significant concern for oncology. ALT was utilised in approximately one quarter of glioblastoma multiforme (GBM), one third of soft tissue sarcomas (STS) including three quarters of malignant fibrous histiocytomas (MFH), half of osteosarcomas and one tenth of non-small cell lung carcinomas (NSCLC). Furthermore, the patients with these ALT[+] tumours had poor survival; median survivals were 2 years for ALT[+] GBM, 4 years for ALT[+] STS including 3.5 years for ALT[+] MFH and 5 years for ALT[+] osteosarcoma. ALT[+] STS and osteosarcomas were also just as aggressive as their ALT[-] counterparts in terms of grade and patient outcome. ALT status was not found to be associated with response to chemotherapy in osteosarcomas or survival in STS. ALT was however, less prevalent in metastatic STS. The APB assay was a prognostic indicator for GBM and was correlated with three fold increased median survival in GBM (although this survival was still poor). ALT was more common in lower grade astrocytomas (88% ALT[+]) than GBM (24% ALT[+]) and ALT[+] GBM had an identical median age at diagnosis to that reported for secondary GBM. It is discussed that these data indicate that ALT was indirectly associated with secondary GBM and is possibly an early event in its progression from lower grade astrocytoma. This is relevant because secondary GBM have distinct genetic alterations that may facilitate activation of the ALT mechanism. Putative repressors of ALT could explain why this study found that ALT varied among the different STS subtypes. ALT was common in MFH (77%), leiomyosarcoma (62%) and liposarcoma (33%) but rare in rhabdomyosarcoma (6%) and synovial sarcoma (9%). ALT was not found in colorectal carcinoma (0/31) or thyroid papillary carcinoma (0/17) which have a high prevalence of telomerase activity and a reduced need for a telomere maintenance mechanism (low cell turnover), respectively. A yeast model of ALT predicts that one of the five human RecQ helicases may be required for ALT. Using the APB assay to test for the presence of ALT in tumours from patients with known mutations in either WRN or RECQL4 it was demonstrated that neither of these RecQ helicases is essential for ALT. Although p53 and mismatch repair (MMR) proteins have been suggested to be possible repressors of ALT, there was no apparent increase in the frequency of ALT in tumours from patients with a germline mutation in p53 codon 273 or in colorectal carcinomas that had microsatellite instability and thus MMR deficiency. Also contrary to being a repressor of ALT but consistent with its ability to interact with a protein involved in the ALT mechanism, the MMR protein MLH1, was demonstrated to be present in the APBs of an ALT[+] cell line. To further test for genes that may be involved in the ALT mechanism or associated with its activation, RNA microarray was used to compare the gene expression of 12 ALT[+] with 12 matched telomerase[+] cell lines; 240 genes were identified that were significantly differentially expressed (p<0.005) between the ALT[+] and telomerase[+] cell lines. Only DRG2 and SFNX4 were significantly differentially expressed after adjusting for the estimated false positive rate. Overall, DRG2, MGMT and SATB1 were identified as most likely to be relevant to the ALT[+] tumours and Western analysis indicated that DRG2 and MGMT levels were down-regulated after activation of ALT and up-regulated after activation of telomerase, whereas SATB1 protein levels appeared to be up-regulated after immortalisation but to a higher degree with activation of ALT compared to telomerase. Since lack of MGMT is known to be a determinant of temozolomide sensitivity in GBM, the possibility that ALT and the APB assay could be used to predict temozolomide sensitivity is discussed. The microarray data was consistent with MGMT expression being suppressed by EGF (p < 0.05), indicating that caution may be needed with combining EGFR inhibitors with temozolomide in ALT cancers. One ALT[+] cell line which did not express MGMT had TTAA sequence in its telomeres. This could possibly have resulted from mutations due to lack of MGMT expression and a possible role for MGMT in the ALT mechanism is discussed. Further analysis of the microarray data identified two groups of co-regulated genes (p < 5x10-5): CEBPA, TACC2, SFXN4, HNRPK and MGMT, and SIGIRR, LEF1, NSBP1 and SATB1. Two thirds of differentially expressed genes were down-regulated in ALT. Chromosomes 10 and 15 had a bias towards genes with lower expression in ALT while chromosomes 1, 4, 14 and X had a bias towards genes with higher expression levels in ALT. This work has developed a robust assay for ALT in tumour specimens which was then used to show the significance of ALT in sarcomas, astrocytomas and NSCLC. It has also identified genes that could possibly be molecular targets for the treatment of ALT[+] cancers.

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Elchinova, Elena Georgieva. "Analysis of the levels of monocyte subsets in patients with heart failure." Doctoral thesis, Universitat Autònoma de Barcelona, 2019. http://hdl.handle.net/10803/667760.

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La insuficiencia cardíaca es un síndrome, caracterizado por diferentes signos y síntomas clínicos debidos a una anomalía estructural o funcional del corazón. Es una de las cardiopatías más predominantes en los países desarrollados, tanto desde el punto de vista epidemiológico como de sus manifestaciones clínicas. La insuficiencia cardíaca es un problema médico creciente relacionado con una alta tasa de hospitalización e importante mortalidad y un pronóstico desfavorable con coste socioeconómico muy elevado en todo el mundo. Los monocitos son una población heterogénea de células efectoras con funciones clave en el mantenimiento y la restauración de la integridad del tejido y del sistema inmunológico. Mediante citometria de flujo se pueden separar tres subpoblaciones de monocitos humanos distintos: clásicos (CD14 ++ / CD16–), intermedios (CD14 ++ / CD16 +) y no clásicos (CD14– / CD16 +). Poco se sabe acerca de la importancia, la relación entre los niveles de los monocitos circulantes en sangre periférica y su distribución en la insuficiencia cardíaca, incluso menos se conoce si estos parámetros podrían usarse como marcadores predictores de la progresión de la enfermedad. El objetivo principal del proyecto actual fue evaluar la relación entre los niveles y la distribución de las diferentes subpoblaciones monocitarias y la longitud de sus telómeros en pacientes con insuficiencia cardíaca y los eventos adversos, comomortalidad y hospitalización por insuficiencia cardíaca. La tesis doctoral actual describe tres estudios, respectivamente de 28, 400 y 101 pacientes ambulatorios, tratados consecutivamente en una unidad multidisciplinaria de insuficiencia cardíaca desde diciembre de 2013 hasta mayo de 2015. Todos los procedimientos del estudio se realizaron de acuerdo con todos los estándares éticos y todos los participantes proporcionaron un consentimiento informado por escrito. Se extrajeron muestras de sangre periférica de todos los pacientes para su posterior análisis mediante citometría de flujo. Las muestras se incubaron directamente con anticuerpos monoclonales con fluorocromos contra antígenos de superficie específicos de monocitos, tipo CD86 (o HLA -DR), CD14 y CD 16, y en paralelo (en 101 muestras) se analizaron marcadores genéticos (telómeros) mediante un citómetro de flujo. (BD LSRFortessa) en el Departamento de Citolatría de la IGTP. Se analizó la distribución porcentual de cada subconjunto de monocitos y también se determinó cuantitativamente su recuento de células absoluto (U/mL). Durante nuestro proyecto pudimos establecer un nuevo método de análisis conjunta de subpoblaciones de monocitos y con determinación de la longitud relativa de los telómeros, que resulto rápido, preciso y mucho más barato que otros métodos utilizados previamente. En nuestro estudio, la subpoblación intermedia se asoció de forma independiente en el análisis multivariable con mortalidad por todas las causas y con la variable compuesta (mortalidad por todas las causas o ingreso por insuficiencia cardiaca). La determinación cuantitativa del recuento de células absoluto de cada subpoblación de monocitos expresado en U/mL fue superior desde el punto de vista pronóstico al porcentaje de estas subpoblaciones. Se observó una reducción de aproximadamente el 22% en la longitud de los telómeros durante un año en los monocitos de nuestros pacientes, aunque la longitud relativa y el cambio en la longitud de los telómeros no se asociaron significativamente con los resultados. Por lo tanto, no es probable que el cambio en la longitud de los telómeros sea un biomarcador útil de la progresión de la insuficiencia cardíaca. Los monocitos y las subpoblaciones de monocitos podrían usarse en el futuro no solo como un factor predictor, sino que también podrían tomarse en consideración como parte de una terapia de inmunomodulación para los pacientes con insuficiencia cardíaca.
Heart failure is a disorder characterized by different clinical signs and symptoms due to a structural or functional anomaly of the heart. It is the most predominant heart disease in developed countries, both from epidemiological point of view and clinical implications. Indeed, it is a growing medical problem related to major hospitalization needs and high mortality, with significant economic and population burden worldwide. Established prognostic factors, such as age, sex, aetiology, comorbidities, New York Heart Association functional class, left ventricle ejection fraction, and routine laboratory markers might fail to completely and individually predict disease progression and mortality. A good risk stratification strategy is crucial as risk might be refined using several biological biomarkers of different pathophysiological processes that the former mortality risk factors do not necessarily directly reflect. That is why efficient and reliable new prognostic predictor markers are of upmost importance and relevance for the future management of the disease. Monocytes are a heterogeneous population of effector cells with key roles in the maintenance and restoration of tissue integrity. Three distinct human monocyte subsets can be identified by flow cytometry: classical (CD14++/CD16–), intermediate (CD14++/CD16+) and non-classical (CD14–/CD16+). Little is known about the importance, relationship between the levels of the circulating monocytes and their distribution in heart failure, even less if these parameters could be used as a predictor markers for the progression of the disease. The main objective of the current project was to assess the relationship between the levels and distribution of the different circulating monocyte subsets and the length of its telomeres in outpatients with heart failure with adverse events, namely mortality and heart failure hospitalizations. Three cohorts of respectively 28, 400 and 101 ambulatory patients, consecutively treated at a multidisciplinary heart failure Clinic from December 2013 to May 2015 were included in the studies described in this doctoral thesis, independently of the data of their entry into the heart failure Clinic program. All study procedures were performed in accordance with all ethical standards and all participants provided written informed consent. Peripheral blood samples of all patients were extracted for subsequent analysis by flow cytometry. The samples were incubated directly by means of monoclonal antibodies with fluorocromes against monocyte specific surface antigens, type CD86 (or HLA -DR), CD14 and CD 16 and in parallel (in 100 samples) genetic markers (telomeres) were subsequently analyzed by flow cytometer (BD LSRFortessa) in the Department of Citolatry of the IGTP. The percentage distribution of each monocyte subset was analyzed and their absolute cell count (U/mL) was also determined quantitatively. We were able to establish an innovating, accurate and much less expensive method than established ones for simultaneously measuring the different monocyte subsets and the its relative telomere length. In our study, the intermediate subset was independently associated with all-cause death and the composite end-point of all-cause death or heart failure hospitalization, in multivariable analyses. The quantitative determination of the absolute cell count of each monocyte subset expressed by U/mL was superior from the prognostic point of view than the percentage of these monocyte subsets in outpatients with Heart failure. We observed about 22% reduction in telomere length over 1 year in the monocytes of our patients, being the baseline telomere length and change in telomere length not significantly associated with outcomes. Therefore, the change in telomere length is not likely to be a useful biomarker of heart failure progression. The monocytes and monocyte subsets could be used not only as a predictor factor but also might be taken into consideration as part of an immuno-modulation therapy in the future for the heart failure patients.

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Garg, Aggarwal Mansi. "Characterization of the role of SUMO in telomere length homeostasis and overhang processing at yeast telomeres." Thesis, University of Sussex, 2017. http://sro.sussex.ac.uk/id/eprint/68661/.

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Nanavaty, Vishal P. "Function of Telomere Protein RAP1 and Telomeric Transcript in Antigenic Variation in Trypanosoma Brucei." Cleveland State University / OhioLINK, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=csu1485424039406009.

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Wiley, Emily A. "Yeast telomere structure : genetic analysis implicating a novel terminus-specific factor in telomeric silencing /." Thesis, Connect to this title online; UW restricted, 1996. http://hdl.handle.net/1773/6359.

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Östlund-Lagerström, Lina. "Effect of long-term ultra-endurance training on telomere length and telomere regulatory protein expressions in vastus lateralis of healthy humans." Thesis, Örebro universitet, Hälsoakademin, 2010. http://urn.kb.se/resolve?urn=urn:nbn:se:oru:diva-15859.

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Lee, Hyemin [Verfasser]. "mEst1A (mouse ever shorter telomeres 1A) regulates telomere length and RNA quality control in murine stem cells / Hyemin Lee." Ulm : Universität Ulm. Fakultät für Naturwissenschaften, 2012. http://d-nb.info/1024534235/34.

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Sherwood, Rebecca. "The Effect of the Copy Number of the Telomerase RNA Gene on the Elongation of Telomeres in Saccharomyces cerevisiae." Thesis, Boston College, 2008. http://hdl.handle.net/2345/532.

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Thesis advisor: Clare O'Connor
Telomeres are repeated sequences at the ends of chromosomes, which promote chromosome stability by preventing the loss of necessary nucleotides from the DNA with successive rounds of replication. Telomeres are elongated by the enzyme telomerase, which has both a protein component and an RNA component. In the yeast Saccharomyces cerevisiae, the TLC1 gene encodes the RNA component of the enzyme. Telomerase RNA interacts with several proteins to perform its function, including the Ku protein, which binds to the end of the DNA and helps to recruit telomerase to the chromosome thereby facilitating the lengthening of chromosome ends. Ku interacts with telomerase RNA at the site of a 48-nucleotide stem-loop on the RNA's structure. Previous experiments have shown that yeast strains engineered to carry two copies of the TLCI gene exhibit higher levels of telomerase RNA than those that have only one copy of the gene. Also, a yeast strain carrying a copy of the mutant tlc1Δ48 gene, which contains a deletion of the 48-nucleotide stem-loop, contains lower levels of telomerase RNA than a strain with the wild type TLC1 gene. This series of experiments is investigating whether the copy number of the telomerase RNA gene affects the elongation of telomeres in S. cerevisiae. In order to determine this effect, the de novo telomere addition of four strains was examined, as were the native telomere lengths of these strains. The assay indicated that the efficiency of telomere elongation was unchanged by increasing the copy number of the wild type gene but was increased upon increasing the copy number of the mutant gene. Analysis of the native telomere lengths showed that increasing the copy number of either the wild type or the mutant gene allowed the cells to maintain their telomeres at a longer length
Thesis (BS) — Boston College, 2008
Submitted to: Boston College. College of Arts and Sciences
Discipline: Biology
Discipline: College Honors Program

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14

Wikgren, Mikael. "Telomeres and the brain : an investigation into the relationships of leukocyte telomere length with functional and structural attributes of the brain." Doctoral thesis, Umeå universitet, Psykiatri, 2011. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-50634.

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Telomeres are the outermost parts of linear chromosomes. They consist of tandemly repeated non-coding short nucleotide sequences (TTAGGG in all vertebrates), in humans spanning over the last 2 to 15 kilobase pairs of the chromosome. Due to the end-replication problem, telomeres shorten with each cellular division. A critically short telomere will trigger the cell to enter a state of cellular senescence or to apoptose. The rate of telomere shortening can be accelerated by factors such as oxidative stress and inflammation. Taken together, this contributed to making telomere length a candidate biomarker of health and aging. Studies have shown that leukocyte telomere length progressively shortens with age, and that it independent of age is associated with age-related morbidity, lifestyle factors, and mortality. This thesis was aimed at exploring the relationships of leukocyte telomere length with various functional and structural attributes of the brain. In Paper I, telomere length was shown to be longer among non-demented carriers of the apolipoprotein E (APOE) ε4 allele, a well-established risk factor for Alzheimer’s disease. However, the rate of telomere shortening was greater among the ε4 carriers, possibly due to the higher levels of oxidative stress and inflammation associated with this allele. Furthermore, performance on episodic memory tests was inversely related to telomere length among ε4 carriers. The results may contribute to a better understanding of the pathophysiology related to the APOE ε4 allele. The volume of the hippocampus, a structure in the brain critical for episodic memory function, was in Paper II found to be inversely related to telomere length among non-demented APOE ε3/ε3 carriers. No correlation between hippocampal volume and telomere length was discernible among ε4 carriers, but they fit the pattern exhibited by the ε3/ε3 carriers as they tended to have smaller hippocampi and longer telomere length compared with the ε3/ε3 carriers. The results are possibly explained by a low proliferative activity among subjects with smaller hippocampi, which might also explain the inverse association between telomere length and episodic memory performance in Paper I. In Paper III, we describe results corroborating earlier findings of shorter telomere length among individuals suffering from depression. Moreover, we found that the shorter telomere length among the patients to a large extent could be linked to a hypocortisolemic state; a state which has been associated with chronic stress. The findings corroborate the link between telomere length and stress, and underline the role of stress in depressive illness. Two prominent manifestations of the aging brain are atrophy and white matter hyperintensities. In Paper IV, we report that white matter hyperintensities and cerebral subcortical atrophy were associated with shorter telomere length in aged non-demented individuals. Cortical atrophy was not associated with telomere length. Inflammation may be the underlying cause of the associations, as it is linked to telomere attrition, subcortical atrophy, and white matter hyperintensities. Taken together, these results show that leukocyte telomere length has the potential of being used as a biomarker for structural and functional attributes of the brain. Furthermore, the findings can provide new insights into mechanisms of disease and aging of the brain

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15

Anjomani, Virmouni Sara. "Genotype and phenotype characterisation of Friedreich ataxia mouse models and cells." Thesis, Brunel University, 2013. http://bura.brunel.ac.uk/handle/2438/7831.

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Friedreich ataxia (FRDA) is an autosomal recessive neurodegenerative disorder, caused by a GAA repeat expansion mutation within intron 1 of the FXN gene, resulting in reduced level of frataxin protein. Normal individuals have 5 to 40 GAA repeat sequences, whereas affected individuals have approximately 70 to more than 1000 GAA triplets. Frataxin is a mitochondrial protein involved in iron-sulphur cluster and heme biosynthesis. The reduction in frataxin expression leads to oxidative stress, mitochondrial iron accumulation and consequential cell death with the primary sites of neurons of the dorsal root ganglia and the dentate nucleus of the cerebellum. FRDA, which is the most common inherited ataxia, affecting 1:50,000 Caucasians, is characterised by neurodegeneration, cardiomyopathy, diabetes mellitus and skeletal deformities. To investigate FRDA molecular disease mechanisms and therapy, several human FXN YAC transgenic mouse models have been established: Y47R, containing normal-sized (GAA)9 repeats; YG8R and YG22R, which initially contained expanded GAA repeats of 90-190 units and 190 units, respectively, but which have subsequently been bred to now contain expanded GAA repeats of 120-220 units and 170-260 units, respectively, and YG8sR (YG8R with a small GAA band) that was recently generated from YG8R breeding. To determine the FXN transgene copy number in the enhanced GAA repeat expansion-based FRDA mouse lines, a TaqMan qPCR assay was developed. The results demonstrated that the YG22R and Y47R lines had a single copy of the FXN transgene while the YG8R line had two copies. The YG8s lines showed less than one copy of the target gene, suggesting potential deletion of the FXN gene. Single integration sites of all transgenes were confirmed by fluorescence in situ hybridisation (FISH) analysis of metaphase and interphase chromosomes. However, in the YG8s line, at least 25% of the YG8s cells had no signals, while the remaining cells showed one signal corresponding to the transgenic FXN gene. In addition, the analysis of FXN exons in YG8s rescue mice by PCR confirmed the presence of all FXN exons in these lines, suggesting the incidence of somatic mosaicism in these lines. Extended functional analysis was carried out on these mice from 4 to 12 months of age. Coordination ability of YG8R, YG8sR and YG22R ‘FRDA-like’ mice, together with Y47R and C57BL6/J wild-type control mice, was assessed using accelerating rotarod analysis. The results indicated a progressive decrease in the motor coordination of YG8R, YG22R and YG8sR mice compared to Y47R or C57BL6/J controls. Locomotor activity was also assessed using an open field beam-breaker apparatus followed by four additional functional analyses including beam-walk, hang wire, grip strength and foot print tests. The results indicated significant functional deficits in the FRDA mouse models. Glucose and insulin tolerance tests were also conducted in the FRDA mouse models, indicating glucose intolerance and insulin hypersensitivity in the aforementioned lines. To investigate the correlation between the FRDA-like pathological phenotype and frataxin deficiency in the FRDA mouse models, frataxin mRNA and protein levels as well as somatic GAA repeat instability were examined. The results indicated that somatic GAA repeats increased in the cerebellum and brain of YG22R, YG8R and YG8sR mice, together with significantly reduced levels of FXN mRNA and protein in the liver of YG8R and YG22R compared to Y47R. However, YG8sR lines showed a significant decrease in FXN mRNA in all of the examined tissues compared to Y47R human FXN and C57BL6/J mouse Fxn mRNA. Protein expression levels were also considerably reduced in all the tissues of YG8sR mice compared to Y47R. Subsequently, the telomere length of human and mouse FRDA and control fibroblasts was assessed using qPCR and Q-FISH. The results indicated that the FRDA cells had chromosomes with relatively longer telomeric repeats in comparison to the controls. FRDA cells were screened for expression of telomerase activity using the TRAP assay and a quantitative assay for hTERT mRNA expression using TaqMan qRT-PCR. The results indicated that telomerase activity was not present in the FRDA cells. To investigate whether FRDA cells maintained their telomeres by ALT associated PML bodies (APBs), co-localisation of PML bodies with telomeres was assessed in these cells using combined immunofluorescence to PML and Q-FISH for telomere detection. The results demonstrated that the FRDA cells had significantly higher co-localised PML foci with telomeric DNA compared to the normal cells. Moreover, telomere sister chromatid exchange (T-SCE) frequencies were analysed in the human FRDA cell lines using chromosome orientation FISH (CO-FISH). The results indicated a significant increase in T-SCE levels of the FRDA cell lines relative to the controls. Furthermore, growth curve and population doubling analysis of the human FRDA and control fibroblasts was carried out. The results showed that the FRDA fibroblast cell cultures underwent growth arrest with higher cumulative population doubling compared to the controls. Though, further analysis of telomere length at different passage numbers revealed that the FRDA cells lost telomeres faster than the controls. Finally, the telomere dysfunction-induced foci (TIF) assay was performed to detect DNA damage in the human FRDA fibroblast cells using an antibody against DNA damage marker γ-H2AX and a synthetic PNA probe for telomeres. The frequency of γ-H2AX foci was significantly higher in the FRDA cells compared to the controls. Similarly, the FRDA cells had greater frequencies of TIFs in comparison to the controls, suggesting induced telomere dysfunction in the FRDA cells.

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16

Santos, Gabriel Arantes Galvão Dias dos. "Caracterização molecular da atividade de interação da proteína RPA-1 com os telômeros de Leishmania spp." Universidade Estadual Paulista (UNESP), 2018. http://hdl.handle.net/11449/153962.

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Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)
Entre as espécies do gênero Leishmania estão os protozoários que causam leishmaniose, uma doença tropical negligenciada endêmica em muitos países, incluindo o Brasil. Métodos de controle e tratamento ainda são ineficientes e a resistência a drogas é um desafio. Por isso, pesquisas para entender melhor a biologia molecular desses parasitos são encorajadas. Uma possível estratégia para isso, é o estudo dos telômeros, estrutura fundamental para a homeostase do genoma. Os telômeros são estruturalmente diferentes do resto do cromossomo, e contam com proteínas específicas que realizam sua manutenção. A Replication Protein A subunit 1 (RPA-1) é uma proteína que interage de DNA de simples fita que tem diversas funções relacionadas com o metabolismo do DNA eucarioto, incluindo os telômeros. A RPA-1 é parte de um complexo heterotrimérico conservado nos eucariotos, incluindo Leishmania spp.. Recentemente nós mostramos por modelagem molecular que a estrutura terciária da LaRPA-1 difere dos seus ortólogos em humanos e leveduras, além de mostrar interações específicas nos telômeros dos parasitos, que na ausência de homólogos canônicos para telomere-end binding protein (TEP) elegem a LaRPA-1 como um potencial candidato para essa função. Neste trabalho, avaliamos a capacidade da LaRPA-1 como uma TEP, cujo papel principal é proteger a extremidade 3' dos telômeros de ataques por exonucleases. Uma busca estrutural por proteínas que compartilham com as TEP domínios de interação proteína-DNA, mostrou que no genoma de Leishmania spp. não existem homólogos estruturais para as mesmas. Aqui mostramos por diferentes abordagens que a LaRPA-1 tem a capacidade de interagir com no mínimo uma repetição telomérica e também é capaz de proteger in vitro a simples fita telomérica rica em G (5’ TTAGGG 3’) da digestão por Exonuclease I bacteriana cuja atividade é no sentido 3’-5’. Somando esses dados, com dados anteriores que mostram que a LaRPA-1 tem preferência pela fita telomérica rica em G e o fato dela ter sido co-purificada com a atividade de telomerase sugerem fortemente que ela está diretamente relacionada com a manutenção da maquinaria telomérica, podendo inclusive ser considerada a principal ligante de simples fita telomérica rica em G (3’ G-overhang) em Leishmania spp.
Among the protozoa parasites of the Leishmania genus are the causative agents of leishmaniasis, a neglected tropical disease endemic in many countries, including Brazil. Disease control and treatment are still inefficient and parasite drug resistance is a challenge. Therefore, efforts for the establishment of intensive research to better understand the molecular biology of these parasites are encouraged. One possible strategy is to study parasite telomeres, a vital chromosome structure important to maintain genome homeostasis. Telomeres are significantly different from the rest of the chromosome and are associated with proteins involved in their maintenance. Replication Protein A subunit 1 (RPA1), a single-stranded DNA-binding protein that plays multiple roles in eukaryotic DNA metabolism, including telomeres, is part of a conserved heterotrimeric complex which is present in most eukaryotes including Leishmania spp. Recently, using molecular dynamics simulations we have shown that the tertiary structure of LaRPA-1 differs from human and yeast RPA-1 and that it also shows parasitespecific interactions with telomeric DNA. In the absence of real homologues to telomere-end binding proteins, LaRPA-1 could be considered a potential candidate. If LaRPA-1 is a telomere-end binding protein, one of its main role would be to protect the telomeric 3`-end termini from nuclease attack. A structural search for proteins that share with the TEP domains of protein-DNA interaction, showed that in the genome of Leishmania spp. there are no structural homologues for them. In this work, we show by different methods, that in vitro LaRPA-1 can bind at least one telomeric repeat and it can also protect the telomeric G-rich sequence (5’ TTAGGG 3’) from the bacterial 3’-5’Exonuclease I digestion. These data compiled to previous data showing that LaRPA-1 preferentially binds the G-rich telomeric DNA and that it co-purifies with telomerase activity strongly suggest that LaRPA-1 is directly involved with parasite telomere maintenance and, possibly, is the main G-rich single-stranded (3’ G-overhang) telomere-binding protein in Leishmania spp.

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17

Foote, Christopher Graham. "Avian telomere dynamics." Thesis, University of Glasgow, 2009. http://theses.gla.ac.uk/539/.

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Telomeres, the repetitive DNA sequences that cap eukaryotic chromosomes, are thought to play an important role in linking life conditions and senescence. In vertebrate somatic cells, telomeres shorten at each cell division, and the rate at which they do so has been linked to cellular and organismal senescence. Although telomeres generally shorten with age in vertebrates, in most species studied there is considerable variation between same age individuals. In this thesis, I examined the telomere dynamics of various avian species, investigating both the causes of variation in telomere length among individuals and what effect this variation has on attributes such as survival rates. Previous studies have shown that most telomere loss occurs in young individuals and it thus makes sense that early life conditions are responsible for much of the inter-individual variation in telomere length. I investigated this idea by studying chick telomere dynamics in a wild population of lesser black-backed gulls Larus fuscus. There was considerable variation in hatching telomere length among individuals and much of this variation was related to circumstances during embryonic growth. Larger hatchlings had shorter telomere lengths, suggesting that embryonic growth rate could have affected telomere attrition. Independent of this trend, males had longer telomeres at hatching than females. Although telomere length did decrease with age post-hatching, these initial variations remained consistent during the initial post-hatching period. The relationship between early life conditions and telomere length was investigated further with a longitudinal study of telomere length in chicks of the European shag Phalacrocorax aritotelis. A previous study on this population of birds had shown that telomere length declines with age within individuals over a period of several years. However no change in telomere length was detected over a period of 11-13 days during the chick period. Body size had no effect on telomere length, but males did have longer telomere than females. These initial chapters investigate telomere length in chicks; however there are very few studies that investigate telomere length over the entire lifespan of long-lived species. I thus next examined the telomere dynamics of two species of long-lived seabird, the northern and southern giant petrels (Macronectes spp.). In both giant petrel species, telomeres were shorter in adults than chicks, but there was no trend for adult telomere length to decrease with age. In southern giant petrels, there was a significant relationship (independent of age and sex) between an individuals telomere length and whether it was still alive 8 years after it was initially sampled. This relationship was not present in northern giant petrels, possibly due to a smaller sample size. The results thus support both the idea that most telomere loss occurs in young individuals and that telomere length may be an indicator of life expectancy. Various methods exist to measure telomeres. As the number of taxa whose telomere dynamics are being studied increases, it becomes increasingly important to know which methods are the best to use and to what extent these methods are applicable across species. These questions were investigated in relation to work conducted on the telomere dynamics of the blue-footed booby Sula nebouxxi. Both the TRF and qPCR techniques were used to measure booby telomeres, but problems arose with both methods. It is possible that these problems occurred because blue-footed boobies have a particularly large amount of interstitial telomeric DNA, although a more detailed analysis of booby telomeres would be necessary to determine this. These findings suggest that standardised methods to measure telomeres cannot necessarily be applied to every new species whose telomere dynamics are studied. The evidence presented here suggests that the study of telomere dynamics can be a very powerful tool for behavioural ecologists. It now seems possible that telomeres might provide both a way of measuring the long-term costs of early life-conditions and a way to measure the quality of an individual. However, further research is still needed to fill in the considerable gaps in our knowledge and fully exploit the potential telomeres have for behavioural ecology.

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18

Hidalgo, Bravo Alberto. "Human telomeres and recombination." Thesis, University of Leicester, 2013. http://hdl.handle.net/2381/27809.

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Telomeres are DNA-protein complexes that help protecting the end of linear chromosomes. They consist of repetitive DNA, in mammals the repeat unit is the hexanucleotide TTAGGG, these repeats span 5-20 kb. Under normal conditions in somatic cells, telomeres get shorter with every population doubling until they reach a critical length and then, the cell enters a checkpoint called senescence or M1 where it stops dividing. If the cell escapes senescence and continues dividing with further telomere shortening, it reaches a second checkpoint called crisis or M2. Crisis is characterized by telomere dysfunction leading to genomic instability that can end with cell death. However, some cells achieve to maintain telomere length by activating a telomere maintenance mechanism (TMM). The presence of a TMM is a hallmark of cancer cells. Two TMM have been described in human cells, one is the through the enzyme telomerase, which is active in 85% of cancers, and the second is a homologous recombination (HR) based mechanism called Alternative Lengthening of Telomeres (ALT) active in 15% of cancers. The evidence that the ALT pathway relies in HR was the observation that sequences can be copied from one telomere to another in ALT+ but not in telomerase+ cells and that several genes involved in HR are necessary for ALT progression. The ALT pathway is not the only event involving HR at telomeres. It has been shown that the human herpesvirus 6 (HHV-6) can integrate into human telomeres. Interestingly, HHV-6 possesses perfect telomeric repeats within its genome. The proposed mechanism for integration if through HR between the telomeric repeats present in the virus with the human telomere repeats. The aim of this work is to unravel the molecular mechanism underlying the ALT pathway and HHV-6 integration. The data obtained will contribute to the understanding of HR in human telomeres.

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19

Musetti, Caterina Livia. "Heterocyclic Cations as Potential Anticancer Agents: An Approach that Targets G-quadruplex with Different Binding Modes." Digital Archive @ GSU, 2010. http://digitalarchive.gsu.edu/chemistry_theses/26.

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G-quadruplex structures are found in important regions of the eukaryotic genome, such as telomeres and regulatory sequences of genes, and are likely to play important roles in regulation of biological events. The significant structural differences with duplex DNA make quadruplex DNA a very attractive target for anticancer drug design. The purpose of this study is to explore conformational space in a series of heterocyclic cations to discover novel structural motifs that can selectively bind and stabilize specific G-quadruplex arrangements. A variety of biophysical techniques such as thermal melting experiments, biosensor surface plasmon resonance, circular dichroism, fluorescence displacement assay and mass spectrometry were employed to evaluate the affinity of the compounds and their recognition properties. The screening of the molecules allowed the identification of not only selective G-quadruplex ligands but also potential quadruplex groove binders. These results can be useful for the development of new efficient telomerase inhibitors which are endowed with pharmacological activity.

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20

Martinez, Alaina R. "Variant requirements for DNA repair proteins in cancer cell lines that use alternative lengthening of telomere mechanisms of elongation." The Ohio State University, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=osu1479924417740462.

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21

Silva, Filipa Isabel Serra e. "Mistranslation and telomere stability." Master's thesis, Universidade de Aveiro, 2011. http://hdl.handle.net/10773/7648.

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Mestrado em Biologia Aplicada
The regulation of a stable proteome is crucial for the cell homeostasis. The translation process from the nucleotide sequence of a gene into the aminoacid sequence of a protein is associated with a basal error of 10-4 which the cell deals with through quality control mechanisms. The misincorporation of aminoacids into de novo synthesized proteins tends to rise when the cell is exposed to stressful conditions. The increase of dysfunctional proteins produced by mistranslation may induce expression of genes related to stress response and genome destabilization. In this work we used yeast as a model to study the impact of high mistranslation rates in telomere stability, since the telomeric and adjacent sub-telomeric regions are key elements on the preservation of genome integrity. Results shown that some types of induced mistranslation may in fact have an impact on telomere length, and also that some proteins’ activity, such as Pnc1 and Sir2, is important regarding telomeric DNA stability. These results shed a new light over the importance of controlling mRNA mistranslation rates in eukaryotic cells.
A manuten¸c˜ao de um proteoma est´avel ´e crucial para a homeostase celular. Ao processo de tradu¸c˜ao da sequˆencia de nucle´otidos de um gene para a sequˆencia de amino´acidos de uma prote´ına est´a associada uma taxa de erro basal de cerca de 10 −4, com a qual a c´elula lida atrav´es de mecanismos de controlo de qualidade das prote´ınas. A incorpora¸c˜ao incorrecta de amino´acidos nas prote´ınas sintetizadas de novo tende a aumentar quando as c´elulas est˜ao expostas a condi¸c˜oes de stress. Por sua vez, o aumento de prote´ınas disfuncionais provocado por erros de tradu¸c˜ao pode induzir a express˜ao de genes de resposta ao stress e destabiliza¸c˜ao do genoma. Neste trabalho, utilizou-se a levedura como modelo para o estudo do impacto de uma elevada taxa de erros de tradu¸c˜ao ao n´ıvel da estabilidade dos tel´omeros, uma vez que esta regi˜ao, juntamente com a regi˜ao subtelom´erica, representa um elemento-chave na preserva¸c˜ao da integridade do genoma. Metodologias de engenheria de tRNA foram utilizadas para induzir erros de tradu¸c˜ao, sendo a t´ecnica de Southern blot escolhida para an´alise do padr˜ao de migra¸c˜ao electrofor´etica de fragmentos de ADN correspondentes `a regi˜ao subtelom´erica. Os resultados obtidos demonstraram que os erros de tradu¸c˜ao podem, de facto, ter impacto a n´ıvel do comprimento dos tel´omeros, dependendo do tipo de erro de tradu¸c˜ao induzido, permitindo ainda confirmar a importˆancia da actividade de prote´ınas como Pnc1 e Sir2 no controlo da estabilidade do ADN telom´erico, lan¸cando uma nova luz sobre a importˆancia do controlo da taxa de erros de tradu¸c˜ao nas c´elulas eucari´oticas.

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22

Schluth-Bolard, Caroline. "Influence des séquences subtélomériques sur la régulation des télomères : exemple du locus de la Dystrophie Facio-Scapulo-Humérale en 4q35 et implication en pathologie." Thesis, Lyon, École normale supérieure, 2011. http://www.theses.fr/2011ENSL0623/document.

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Les subtélomères forment la transition entre les séquences spécifiques des chromosomes et les répétitions télomériques terminales. Ils semblent capables d’influencer les fonctions télomériques mais les connaissances sur les mécanismes mis en jeu sont encore limitées. Les subtélomères sont pourtant associés à de nombreuses pathologies comme la myopathie facio-scapulo-humérale (FSHD), une dystrophie musculaire secondaire à la contraction de répétitions macrosatellites D4Z4 dans la région subtélomérique 4q35. Afin d’étudier les propriétés de la séquence subtélomérique D4Z4, nous avons créé des constructions reproduisant l’organisation génomique au locus 4q35. Nous avons montré que D4Z4 est capable d’adresser un télomère à la périphérie du noyau. Cette activité est couplée à une activité insulatrice au niveau d’une séquence proximale de 80 pb et est dépendante de CTCF et des Lamines A. De plus, la relocalisation périphérique d’un télomère par D4Z4 s’accompagne d’une réplication plus tardive de celui-ci. Par ailleurs, la recherche de séquences capables de s’opposer à l’effet de position télomérique (TPE) a identifié un élément de 30 pb contenant un site CTCF dans la séquence insulatrice proximale de D4Z4. De même, l’introduction d’un signal de poly-adénylation entre un gène rapporteur et les répétitions télomériques interfère avec le TPE et est accompagnée d’une diminution d’un transcrit hybride contenant le gène rapporteur et des répétitions télomériques, suggérant un rôle des transcrits télomériques TERRAs dans la régulation du TPE. En conclusion, ce travail a permis de caractériser l’implication de séquences subtélomériques, et notamment D4Z4, dans la régulation des télomères, leur compartimentalisation nucléaire, la réplication ou l’effet de position télomérique. De plus, il apporte un éclairage nouveau sur la physiopathologie de la FSHD et ouvre des perspectives dans la compréhension d’autres pathologies liées aux subtélomères
Subtelomeres form the transition between chromosome specific sequences and terminal telomeric repeats. They might influence telomeric functions but underlying mechanisms are still unclear. Nevertheless, subtelomeres are associated with a number of human pathologies such as facioscapulohumeral muscular dystrophy (FSHD), an autosomal dominant disease secondary to the contraction of an array of D4Z4 macrosatellite repeats in the subtelomeric region 4q35. In order to study the biological function of the D4Z4 sequence, we created contructs that mimic the genomic organization of the 4q35 locus. We showed that D4Z4 is able to localize a telomere at the nuclear periphery. This perinuclear activity was dependant on interactions with CTCF and A type lamins and lied within a 80 bp proximal sequence that harbors an insulator activity. Moreover, the peripheral positionning of a telomere by D4Z4 is accompanied by a late replication timing of the telomere. We also searched for sequences able to counteract telomeric position effect (TPE) and identified a 30 bp element containing a CTCF binding site in the proximal region of D4Z4. In another construct, the introduction of a poly-adenylation signal between a reporter gene and telomeric repeats counteracted TPE. This effect is accompanied by the production of a hybrid transcript encompassing the reporter gene and telomeric repeats, suggesting a role for the TERRAs telomeric transcripts in TPE regulation. This work contibuted to characterize the role of subtelomeric sequences, especially the D4Z4 macrosatellite, in telomere regulation, their nuclear compartimentalization, their replication or the telomeric position effect. We will discuss the implications in the understanding of the pathophysiology of FSHD and other subtelomeric diseases

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23

Thomson, Philippa. "Isolation and characterisation of telomere and telomere-related sequences in the chicken genome." Thesis, University of Leicester, 1997. http://hdl.handle.net/2381/29768.

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Telomeres are the physical ends of eukaryotic chromosomes, and are essential for chromosome stability and complete replication. Chicken telomere and telomere-related sequences were characterised using a variety of approaches including: cosmid libraries, anchored PCR strategies, and fluorescence in situ hybridisation (FISH). The results suggested that chicken chromosomes terminate in 6.7-14.0 kb tandem arrays of the vertebrate telomere repeat, (TTAGGG)n. The proximal boundaries of these arrays are degraded, and include many variant repeat types which differ from the telomere repeat by one base change. Southern analysis suggested that the degraded repeats can be digested with restriction enzymes, increasing the probability of locating telomere-related sequences in standard cosmid libraries. Complex repeats associated with the terminal arrays and the degraded repeats were isolated, and shown to be located proterminally on metaphase chromosomes. Hybridisation of a telomere repeat probe to genomic DNA separated by pulse field gel electrophoresis (PFGE) suggested that complex subtelomeric repeats may form large satellite blocks at the ends of a subset of chromosomes, resulting in terminal MboI restriction fragments of up to 620kb. Fluorescence in situ hybridisation (FISH) experiments indicated that the distribution of subterminal and terminal sequences within the genome were correlated with distance from the chromosome end. Sequences located towards the ends of chromosomes (i.e. interspersed telomere repeats) were found on more chromosomes than those from more proximal locations (i.e. complex repeats). Screening of a large insert cosmid library (Stratagene) with a telomere repeat probe resulted in the isolation of 59 clones. Single-locus restriction fragment length polymorphisms (RFLPs) were identified from ten of these clones, and the inserts were localised by FISH on metaphase chromosomes. The cosmid probes showed no bias towards location on macrochromosomes. The results suggested that some of the clones may contain DNA from subtelomeric locations, but that others clearly identify interstitial loci.

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24

Kamnert, Iréne. "Classes of DNA associated with telomeres in the chironomids C. pallidivittatus and C. tentans." Lund : Dept. of Genetics, Lund University, 1997. http://catalog.hathitrust.org/api/volumes/oclc/39009480.html.

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25

Brouilette, Scott Wayne. "Telomeres and coronary heart disease." Thesis, University of Leicester, 2004. http://hdl.handle.net/2381/29899.

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Using mean telomere length as a marker of biological age, I show that: 1. Subjects with premature myocardial infarction (MI) have significantly shorter telomeres than age-sex matched, healthy, controls. The mean telomere length in MI subjects was similar to controls almost 11 years older. 2. Healthy young adult children of families with a strong history of premature MI have shorter telomeres than age matched children of families without such a history. 3. Shorter telomere lengths are associated with increase risk of subsequent CHD events in a prospective study. This analysis was carried out on samples collected in the West of Scotland Coronary Prevention Study (WOSCOPS). This randomised blinded trial was designated to examine the benefits of statin treatment on preventing CHD and showed a 30% reduction of events in those treated with pravastatin. Interestingly, my analysis showed that this benefit of statin is only seen in those subjects at higher risk of CHD based on their telomere length.;As the final part of the thesis I carried out a quantitative linkage trait (QTL) analysis in sib-pairs in an attempt to identify genetic loci regulating telomere length. I report the mapping of a major QTL on chromosome 12 that determines almost 50% of the inter-individual variation in mean telomere length.;These findings support a novel "telomere" hypothesis of CHD. They indicate that telomere biology is intimately linked to the genetic aetiology and pathogenesis of CHD. Specifically, the findings suggest that (i) those individuals born with shorter telomeres may be at increased risk of CHD (ii) rather than individual genes, a more global structural property of the genetic material may explain the familial basis of CHD (iii) variation in telomere length may explain, in part, the variable age of onset of CHD. The findings provide several new avenues for future research.

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26

Revie, John. "Identification and characterisation of telomere regulatory and signalling pathways after induction of telomere dysfunction." Thesis, University of Glasgow, 2016. http://theses.gla.ac.uk/7358/.

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Telomeres are DNA-protein complexes which cap the ends of eukaryotic linear chromosomes. In normal somatic cells telomeres shorten and become dysfunctional during ageing due to the DNA end replication problem. This leads to activation of signalling pathways that lead to cellular senescence and apoptosis. However, cancer cells typically bypass this barrier to immortalisation in order to proliferate indefinitely. Therefore enhancing our understanding of telomere dysfunction and pathways involved in regulation of the process is essential. However, the pathways involved are highly complex and involve interaction between a wide range of biological processes. Therefore understanding how telomerase dysfunction is regulated is a challenging task and requires a systems biology approach. In this study I have developed a novel methodology for visualisation and analysis of gene lists focusing on the network level rather than individual or small lists of genes. Application of this methodology to an expression data set and a gene methylation data set allowed me to enhance my understanding of the biology underlying a senescence inducing drug and the process of immortalisation respectively. I then used the methodology to compare the effect of genetic background on induction of telomere uncapping. Telomere uncapping was induced in HCT116 WT, p21-/- and p53-/- cells using a viral vector expressing a mutant variant of hTR, the telomerase RNA template. p21-/- cells showed enhanced sensitivity to telomere uncapping. Analysis of a candidate pathway, Mismatch Repair, revealed a role for the process in response to telomere uncapping and that induction of the pathway was p21 dependent. The methodology was then applied to analysis of the telomerase inhibitor GRN163L and synergistic effects of hypoglycaemia with this drug. HCT116 cells were resistant to GRN163L treatment. However, under hypoglycaemic conditions the dose required for ablation of telomerase activity was reduced significantly and telomere shortening was enhanced. Overall this new methodology has allowed our group and collaborators to identify new biology and improve our understanding of processes regulating telomere dysfunction.

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Gerasimopoulos, Efthalia. "Topoisomerase-mediated poxviral telomere resolution." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 2000. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape2/PQDD_0021/MQ47325.pdf.

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28

Qi, Qi. "Mathematical modelling of telomere dynamics." Thesis, University of Nottingham, 2011. http://eprints.nottingham.ac.uk/12258/.

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Telomeres are repetitive elements of DNA which are located at the ends of chromosomes. During cell division, telomeres on daughter chromomeres shorten until the telomere length falls below a critical level. This shortening restricts the number of cell divisions. In this thesis, we use mathematical modelling to study dynamics of telomere length in a cell in order to understand normal ageing (telomere shortening),Werner’s syndrome (a disease of accelerated ageing) and the immortality of cells caused by telomerase (telomere constant length maintenance). In the mathematical models we compared four possible mechanisms for telomere shortening. The simplest model assumes that a fixed amount of telomere is lost on each replication; the second supposes that telomere loss depends on telomere length; for the third case the amount of telomeres loss per division is fixed but the probability of dividing depends on telomere length; the fourth cases has both telomere loss and the probability of division dependent on telomere length. We start by developing Monte Carlo simulations of normal ageing using these four cases. Then we generalize the Monte Carlo simulations to consider Werner’s syndrome, where the extra telomeres are lost during replication accelerate the ageing process. In order to investigate how the distribution of telomere length varies with time, we derive, from the discrete model, continuum models for the four different cases. Results from the Monte Carlo simulations and the deterministic models are shown to be in good agreement. In addition to telomere loss, we also consider increases in telomere length caused by the enzyme telomerase, by appropriately extending the earlier Monte Carlo simulations and continuum models. Results from the Monte Carlo simulations and the deterministic models are shown to be in good agreement. We also show that the concentration of telomerase in cells can control their proliferative potential.

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Richter, Torsten. "Mechanisms of telomere-dependent senescence." Thesis, University of Newcastle Upon Tyne, 2006. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.435572.

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Kedziora, Sylwia Maria. "How Rif1 controls telomere length." Thesis, University of Aberdeen, 2017. http://digitool.abdn.ac.uk:80/webclient/DeliveryManager?pid=235993.

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The activation of replication origins is temporally regulated in S phase, with some origins activating early and some late. The molecular events controlling the temporal programme are not well understood, but in S. cerevisiae there is a close relationship between telomere length and nearby origin activation time. In the first part of this thesis I explore how the initiation time of origins near to telomeres is regulated by telomere length in a manner dependent on the Tel1 kinase. I demonstrate that an induced short telomere drives early activation of a nearby origin, but that in the absence of Tel1 the same origin activates late. In the second, major part of this thesis I focus on how the Rif1 protein negatively regulates length of the terminal TG1-3 repeats. While Rif1 has long been known to control telomere length, the mechanism through which Rif1 prevents telomere over-extension has remained unclear. Recently Rif1 was discovered to act in DNA replication control as a Protein Phosphatase 1-targeting subunit, directing Protein Phosphatase 1 (PP1) to dephosphorylate the MCM replicative helicase complex. I therefore investigated whether Rif1 also controls telomere length through PP1 interaction. I examine the effects of a mutant Rif1 with its PP1 interaction sites mutated to ablate PP1 binding. I found the mutant Rif1 binds normally to telomeres but causes a long telomere phenotype, similar to that in ∆rif1 cells, implicating Rif1-PP1 interaction in telomere length control. In further experiments I show that tethered PP1 can partially substitute for Rif1 in telomere length control. I also establish that the effect of Rif1-PP1 on telomere length does not operate indirectly through replication timing control, but rather appears to act through a direct pathway controlling telomerase recruitment. I discuss potential dephosphorylation targets, and the mechanism through which Rif1 and PP1 may control telomere length homeostasis. To summarise, my PhD research demonstrates that S. cerevisiae Rif1 acts with PP1 to repress telomerase-mediated TG1-3 repeat extension.

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Pooley, Karen Anne. "Genetic factors in telomere length." Thesis, University of Cambridge, 2011. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.609670.

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Zhao, Y. "Pot1 phosphorylation regulates telomere function." Thesis, University College London (University of London), 2012. http://discovery.ucl.ac.uk/1380712/.

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The telomere is a conserved nucleoprotein structure at the ends of eukaryotic chromosomes. It is essential for maintenance of genomic stability: on the one hand, it suppresses DNA damage response and protects the natural chromosome ends from repair activities; on the other hand, it recruits telomerase, the specialized reverse transcriptase, to counteract the end-replication problem. The telomeric G-strand ssDNA-binding protein Pot1 plays a crucial role in both of these functions. In fission yeast S. pombe, inhibition of Pot1 induces rampant 5’ resection and loss of telomere signal in a single cell cycle. It was recently shown that spPot1 interacts with, and is phosphorylated by, the master cell cycle regulator DDK. Alleles of a V5-tagged version of pot1+ were constructed with mutations at the putative phosphorylation sites, which reside in the N-terminal OB-fold DNA binding domain of Pot1 {Kuznetsov, 2008 #6380}. The goal of this study was to determine the molecular mechanism by which phosphorylation of Pot1 regulates telomere function. We found that the phospho-deficient mutants of Pot1 induce telomere elongation, checkpoint activation, and deregulation of ssDNA generation, suggesting reduced association with the ssDNA. Our data point to a model in which cell cycle-regulated Pot1 phosphorylation coordinates telomere replication and telomerase activity in different cell cycle phases. Furthermore, we showed that the C-terminal V5-tagging of Pot1 also affects its functions, suggesting an additional layer of complexity governing Pot1 function.

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Bernal, Martínez Aina. "Telomere deprotection and the maintenance of genome integrity: discrepancy between telomere shortening and shelterin dysfunction." Doctoral thesis, Universitat Autònoma de Barcelona, 2019. http://hdl.handle.net/10803/666888.

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Els telòmers són estructures nucleoprotèiques que segellen l’extrem cromosòmic i el protegeixen de la reparació il·legítima mitjançant la formació d’un llaç telomèric anomenat t-loop que està modulat per la proteïna TRF2. En les últimes dècades, s’ha demostrat que la disfunció telomèrica és un mecanisme capaç d’originar inestabilitat cromosòmica (CIN) en cèl·lules humanes i de ratolí i promoure la tumorigènesis en models murins. L’objectiu d’aquesta tesi és immortalitzar cèl·lules telomèricament inestables i avaluar el seu potencial tumorigènic mitjançant la desprotecció telomèrica a través de l’escurçament telomèric o de la disfunció de la proteïna TRF2. En el Treball I, cèl·lules epitelials mamàries humanes deficients per p16INK4a (vHMECs) i deficients o no per p53 (p53+/+) foren cariotipades a diferents temps de doblatge per avaluar la presència d’anomalies cromosòmiques i l’evolució del cariotip. En l’absència de la telomerasa, els telòmers s’escurçaren de forma progressiva fins esdevenir desprotegits, i la subseqüent reparació il·legítma dels extrems afavorí l’entrada en cicles de ruptura-fusió-pont (BFB) i disparà la CIN. Tanmateix, les cèl·lules inestables finalment moren independentment de la funcionalitat de p53. Per contra, la sobreexpressió d’hTERT en les cèl·lules vHMEC-p53+/+ afavoreix la proliferació indefinida amb un cariotip gairebé estable, mentre que la immortalització de les cèl·lules vHMEC-shp53 amb signes de CIN és permissiva amb la proliferació cel·lular i l’evolució del cariotip. Al Treball II i al Treball III, la desprotecció telomèrica aguda va ser induïda mitjançant l’expressió transitòria d’un dominant negatiu de TRF2 (TRF2BM). El dany telomèric agut fou avaluat en la línia cel·lular epitelial mamària MCF-10A (Treball II) i en cèl·lules epitelials mamàries (HMECs) immortalitzades derivades de pacients sanes (Treball III). Al Treball II, la desprotecció telomèrica mitjançant TRF2BM generà un increment de TIFs, fusions telomèriques i ponts anafàsics. Els ponts anafàsics es consideren els inductors dels cicles (BFB) i de la posterior reorganització cariotípica. No obstant, no s’observaren reorganitzacions cromosòmiques derivades dels cicles BFB després de l’expressió transitòria del TRF2BM, independentment de la funcionalitat de les proteïnes p53 i pRb. Després de successius cicles de desprotecció telomèrica, s’observà un increment en el nombre de cèl·lules diploides, tot suggerint que l’excessiu dany telomèric evitaria la proliferació d’aquelles cèl·lules que podrien esdevenir altament reorganitzades. Al Treball III s’expressà TRF2BM en HMECs immortalitzades mitjançant hTERT i l’antigen SV40LT (HMEC-TO). L’expressió transitòria del TRF2BM induí l’increment de ponts anafàsics, però les cèl·lules tampoc presentaren reorganitzacions pròpies dels cicles BFB en marxa. A diferència del Treball II, les cèl·lules poliploides incrementaren arrel del procés d’immortalització. Independentment de la causa d’aquest increment en les cèl·lules poliploides, les cèl·lules exposades als cicles d’expressió de TRF2BM no presentaven un fenotip associat a la disfunció telomèrica, ni tampoc un potencial tumorigènic, tot suggerint que l’expressió del mutant TRF2BM provoca un efecte deleteri sobre la viabilitat cel·lular i l’inici de la CIN. En resum, la present tesi evidencia que el grau de dany telomèric és una eina de doble fil en el manteniment de la integritat genòmica. Per una banda, l’escurçament telomèric indueix un dany cel·lular progressiu i lleu, compatible amb la viabilitat cel·lular. Quan el dany telomèric assoleix un cert llindar, la cèl·lula activa mecanismes dependents i independents de p53 que indueixen la seva mort. Pel contrari, la disfunció de TRF2 afecta a un nombre molt elevat de telòmers i indueix una exagerada resposta cel·lular que la fa incompatible amb la viabilitat cel·lular i la reorganització del cariotip. Aquesta tesi demostra que la desprotecció simultània d’un nombre elevat de telòmers pot ser una eina útil per generar un dany al DNA molt elevat i mantenir la integritat genòmica.
Telomeres are nucleoprotein structures that cap the end of chromosomes and protect them from illegitimate recombination through a lariat conformation or t-loop that is mainly promoted by TRF2 protein. Dysfunctional telomeres have been proved to be a mechanism capable of originating chromosome instability (CIN) in mouse and human cells, and promote tumorigenesis in mouse models. This dissertation thesis aims to generate immortalised but unstable cells due to telomere deprotection through progressive telomere shortening and by TRF2 depletion, and to evaluate their tumorigenic potential. In Work I, p16INK4a-deficient human mammary epithelial cells (vHMECs) lacking or not for p53 function through specific short-hairpin RNA inactivation, were karyotyped at different population doublings to evaluate chromosomal abnormalities and their evolution. In the absence of telomerase, vHMECs progressively shortened their telomeres and subsequent end-to-end fusions initiated breakage-fusion-bridge (BFB) cycles and promoted CIN. However, these unstable cells finally succumbed to cell cycle arrest, independently of their p53 checkpoint status. In contrast, hTERT overexpression in p53-proficient vHMECs resulted in cells able to proliferate indefinitely with a nearly stable karyotype, while immortalised p53-deficient cells showed signs of CIN that could be permissive with an evolving karyotype. In Work II and Work III, acute telomere deprotection was induced by t-loop disassembly through transient expression of the dominant negative form of TRF2 (TRF2BM) in the mammary cell line MCF-10A and in immortalised HMEC derived from cosmetic reductions of four healthy donors, respectively. In Work II, acute telomere deprotection phenotype was reflected by the presence of TIFs and by an increase of end-to-end fusions and anaphase bridges after TRF2BM induction. Anaphase bridges are considered the prelude to breakage-fusion-bridge cycles and subsequent karyotype reorganisations. However, no scars of BFB cycles or highly reorganised cells have been observed after transient expression of TRF2BM, independently of the status of p53 and pRb proteins. Instead, diploid cells were enriched after successive cycles of telomere deprotection induction, thus suggesting that excessive telomere deprotection could be detrimental for the origin of cells with highly reorganised karyotypes. According with these results, in Work III, immortalised HMEC through hTERT and SV40LT overexpression transiently expressing TRF2BM (HMEC-TO) exhibited an increase of anaphase bridges. But after a minimum of five cycles of telomere protection and deprotection, TRF2BM expressing cells did not display scars of telomere deprotection and ongoing BFB cycles. In contrast to MCF-10A derived cell lines, the HMEC-TO cell lines exhibited a progressive increase of polyploid cells as a consequence of SV40LT immortalisation process. Independently of the cause of polyploidy increase, cells exposed to TRF2BM expression cycles did not exhibit a telomere dysfunction phenotype or either a tumorigenic potential, thus suggesting that TRF2BM expression provoked a deleterious effect over TRF2BM expressing cells and prevented CIN emergence. In conclusion, the present dissertation provides evidence that telomere dysfunction acts as a double sword mechanism for genome integrity. On the one hand, telomere shortening induces a mild and progressive DNA damage that firstly is compatible with cell viability, until damage is high enough to induce cell death. On the contrary, shelterin dysfunction affects widely to all chromosomes inducing an exacerbated cell response that is deleterious for cell viability and karyotype reorganisation. This Thesis illustrate that acute telomere deprotection through shelterin dysfunction could be a useful tool to impinge an exacerbated DNA damage and maintain genome integrity in human mammary cells.

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34

Hirsch, Erica. "Telomerase activity and telomere lengths in fibroblast cells treated with ependymin peptide mimetics." Link to electronic thesis, 2005. http://www.wpi.edu/Pubs/ETD/Available/etd-050505-134911/.

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Sousa, Rute Inês Silva e. 1983. "The Epigenetic regulation of Drosophila telomeres." Doctoral thesis, Universitat Pompeu Fabra, 2012. http://hdl.handle.net/10803/96908.

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Drosophila telomere maintenance depends on the transposition of three specialized retrotransposons – HeT-A, TART and TAHRE (HTT). Controlling the activation and silencing of these elements is crucial to maintain telomere length homeostasis without compromising genomic instability. In this thesis, I have identified the role of different chromosomal proteins involved in creating the correct chromatin environment to achieve telomere length homeostasis and stability. JIL-1, together with HP1a and Z4, act as a boundary at the telomere-subtelomere frontier. The interplay of these proteins leads to an equilibrium in the activation/repression state of the telomere retrotransposons. Additionally, I have contributed to the finding that the HeT-A Gag protein is a key component targeting different protein complexes to the telomeres and guaranteeing genome stability. I have also been able to demonstrate that the Z4 partners DREF, TRF2 and KEN are also involved in the silencing of HTT, probably by mediating chromatin remodeling. Finally, I have identified a special subtelomere domain at the 4R telomere with different chromatin characteristics and demonstrated that SETDB1, HP1a and POF are involved in the regulation of the telomeric retrotransposons in the 4th chromosome. These results provide important insights to better understand how in Drosophila the telomere retrotransposons are orchestrated to achieve a telomere function analogous to telomerase telomeres in other eukaryotes.
El manteniment dels telòmers de Drosophila depèn de la transposició especialitzada de tres retrotransposons, HeT-A, TART i TAHRE (HTT). El control de l’activació i la repressió d’aquests elements és crucial a l’hora de mantenir la llargada telomèrica sense comprometre l’estabilitat genòmica. En aquesta tesi jo he pogut identificar el paper de diferents proteïnes cromosòmiques involucrades en crear un estat de la cromatina adient per mantenir la longitud i l’estabilitat telomèrica. JIL-1 juntament amb HP1a i Z4 ajuden a crear el llindar entre la frontera dels domini telomèric i subtelomèric. L’actuació conjunta d’aquestes proteïnes aconsegueix un estat d’equilibri activació/repressió dels retrotransposons telomèrics. A més a més, he contribuït a la descoberta de la implicació de la proteïna HeT-A Gag en el reclutament de diferents complexes proteics als telomèrs de Drosophila per poder garantir l’estabilitat telomèrica. També he pogut demostrar que altres membres dels complexes on participa Z4, com ara: DREF, TRF2 i KEN, estan també implicats en el silenciament dels retrotransposons telomèrics segurament per mitjà de la remodelació de la cromatina. Finalment he pogut demostrar que el domini subtelomèric del telòmer 4R, té una estructura cromatínica diferent a la resta dels dominis subtelomèrics dels altres cromosomes i he pogut demostrar que les proteïnes SETDB1, HP1a i POF estan implicades en la regulació de l’HTT del cromosoma 4. Els resultats d’aquesta tesi ajuden de manera substancial a comprendre com els retrotransposons telomèrics estan orquestrats per tal de poder fer una funció anàloga als telòmers de telomerasa en altres eucariotes.

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Wang, Zhuo. "Extracellular Inflammatory Signaling from Dysfunctional Telomeres." Thesis, University of the Sciences in Philadelphia, 2018. http://pqdtopen.proquest.com/#viewpdf?dispub=10692989.

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Telomere dysfunction describes the catastrophic damage at telomeres, which often leads to genomic instability at the cellular level. There is rising evidence showing that telomere dysfunction also influences the extracellular environment with the inflammatory response. However, little is known about the molecular mechanism of this dysfunctional telomere-associated inflammation. In this dissertation, we identified extracellular forms of Telomeric repeat-containing RNA (TERRA), and demonstrated it might play a role in mediating the crosstalk of telomere dysfunction and inflammation. We found this cell-free TERRA (cfTERRA) is present in mouse tumor and embryonic brain tissue, as well as in human tissue culture cell lines using RNA in situ hybridization. RNA-seq analyses revealed TERRA to be among the most highly represented transcripts in extracellular fractions derived from both normal and cancer patient blood plasma. By characterizing extracellular fractions of the human lymphoblastoid cell line (LCL) culture media, cfTERRA is shown as a shorter form (∼200 nt) of cellular TERRA and co-purifies with CD63- and CD81-positive exosome vesicles that could be visualized by cryo-electron microscopy. Mass spectrometry and extracellular chromatin immunoprecipitation (ChIP) assays revealed that regular cfTERRA was physically interacting with histones and telomeric DNA. Incubation of cfTERRA-containing exosomes with peripheral blood mononuclear cells (PBMCs) stimulated transcription of several inflammatory cytokine genes, including TNFα, IL6, and C-X-C chemokine 10 (CXCL10). Exosomes engineered with elevated TERRA or liposomes with synthetic TERRA further stimulated inflammatory cytokines, suggesting that exosome-associated TERRA augments innate immune signaling. The levels of cfTERRA and DNA damage marker γH2AX were increasingly incorporated into the exosomes during telomere dysfunction. These dysfunctional telomere-derived exosomes activated a more robust transcription of inflammatory cytokines in PBMCs. These findings imply a previously unknown extrinsic function of TERRA and a potentially molecular mechanism of communication between telomeres and innate immune signaling in tissue and tumor microenvironments.

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Cross, Sally H. "Isolation and characterisation of human telomeres." Thesis, University of Edinburgh, 1989. http://hdl.handle.net/1842/13500.

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Cavazos, David Antonio. "Inefficient repair of double-strand breaks at telomeres in Werner syndrome : a dissertation /." San Antonio : UTHSC, 2007. http://proquest.umi.com/pqdweb?did=1400951191&sid=1&Fmt=2&clientId=70986&RQT=309&VName=PQD.

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39

Surovtseva, Yulia V. "Telomere-associated proteins in Arabidopsis thaliana." [College Station, Tex. : Texas A&M University, 2008. http://hdl.handle.net/1969.1/ETD-TAMU-2656.

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Gylfadottir, Valgerdur. "Telomere dynamics in hematopoietic stem cells." Thesis, University of British Columbia, 2007. http://hdl.handle.net/2429/31905.

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Hematopoietic stem cells (HSCs) are known to possess an exceptionally high replicative potential (Szilvassy SJ 2003). However, although they express detectable levels of telomerase, previous murine studies have suggested that HSCs still experience telomere shortening and are not immortal (Allsopp RC 2003). To further clarify the role of aging and clonal exhaustion in HSCs we used murine models to investigate the relationship between the number of cells transplanted, telomerase status, the length of their telomeres and their long term exhaustion. We initiated our studies by investigating the replicative potential of titrated numbers of highly purified HSCs. Results showed that bone marrow stemming from a single HSC could only be serially transplanted twice whereas bone marrow stemming from 10 HSCs could be serially transplanted at least three times. In order to better understand HSC replicative potential, we modified Flow-FISH for murine telomere length measurements. Our Flow-FISH protocol enables us to measure telomere lengths at different timepoints within individual mice, providing us with a unique insight into the telomere length status. With the primary goal of clarifying the role of aging and clonal exhaustion in HSC we aimed to investigate the relationship between telomerase status and HSC exhaustion. We set out to characterize the HSCs of TERT-KO mice that lack the mTERT gene encoding the telomerase enzyme (Erdmann N 2004). A comparison of the frequency of Kit⁺Sca⁺Lineage ̄ cells between the TERT-KO and its wild type counterpart revealed no difference. To our knowledge no studies have explored the rate of telomere shortening of HSC within individual mice over time. Using our optimised Flow-FISH based method we studied the rate of telomere shortening within primary BM transplant recipients, in relation to telomerase status. We find that telomere length is maintained over a nine month period post transplantation, regardless of telomerase status. Furthermore, serially transplanting titrated numbers of wild type and telomerase deficient WBM does not result in telomere shortening. However, despite the apparent maintenance of telomere length, reconstitution levels decrease with each transplantation regardless of transplant dose and telomerase status. These findings suggest the presence of telomere independent barriers, HSC dilution (Iscove NN 1999), alternative lengthening of telomeres or low HSC turnover following WBM transplantation.
Medicine, Faculty of
Medicine, Department of
Experimental Medicine, Division of
Graduate

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Poon, Steven Sui-Sang. "Telomere length measurements using fluorescence microscopy." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1998. http://www.collectionscanada.ca/obj/s4/f2/dsk3/ftp04/nq27227.pdf.

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42

Barnett, Michael A. "Telomere directed breakage of mammalian chromosomes." Thesis, University of Oxford, 1992. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.314882.

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Konrad, Jonathan Paul. "Telomere replication and regulation in vertebrates." Thesis, University of Cambridge, 1999. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.624117.

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Hyatt, Sam. "Examining telomere dysfunction in multiple myeloma." Thesis, Cardiff University, 2017. http://orca.cf.ac.uk/106360/.

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Telomeres are repetitive nucleotide sequences of TTAGGG that cap the ends of linear eukaryotic chromosomes. Short dysfunctional telomeres have previously been identified as a driving force in cancer, resulting in chromosomal fusion and rearrangement that acts to facilitate progression of the malignancy. As it has recently been demonstrated that telomere length is an accurate predictor of clinical outcome in patients with chronic lymphocytic leukaemia (CLL), we aimed to determine whether a similar relationship existed in multiple myeloma (MM). Having used single telomere length analysis (STELA) to measure the mean XpYp telomere length of whole bone marrow aspirates from 141 MM patients, a telomere length threshold of 3.92kb was identified which could be used to stratify patients as either low- or high-risk. Incorporation of this threshold into the international staging system (ISS) for MM increased its prognostic resolution, allowing each prognostic subset to be further risk-stratified. Having demonstrated that a shorter mean XpYp telomere length (< 3.92kb) was associated with inferior patient outcome in MM, we next sought to identify a potential cause for this observation. Using clonal populations of the JJN-3 cell line, each expressing DN-hTERT, we observed that telomeric shortening resulted in a greater frequency of fusion and the initiation of a telomere-driven crisis. Cells were eventually able to escape crisis, driven by the spontaneous reactivation of telomerase which led to increasing telomere length and decreasing fusion frequency. Finally, we sought use the PARP inhibitors Rucaparib and Olaparib to prevent the escape of these clonal JJN-3 populations from a telomere-driven crisis. It was thought that PARP inhibition would interfere with the DNA repair pathways that are known to be responsible for processing telomere-deficient chromosomes. It was established that treating JJN-3 cells with either 7.50μM Rucaparib or 3.75μM Olaparib prevented their escape from a telomere-driven crisis.

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Johnston, Jeffrey Scott. "Combination therapy targeting telomere and telomerase /." The Ohio State University, 2002. http://rave.ohiolink.edu/etdc/view?acc_num=osu1486462067841929.

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Depcrynski, Amy. "Chaperone Association with Telomere Binding Proteins." VCU Scholars Compass, 2009. http://scholarscompass.vcu.edu/etd/1949.

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The Hsp90 chaperone complex associates with the telomerase enzyme, facilitating the assembly of the ribonucleoprotein complex. While previous data from our laboratory indicate that Hsp90 and p23 remain stably associated with (functionally active) telomerase, more recent experiments suggest that these chaperones associate with telomeres independent of telomerase, presumably through a specific interaction with telomere binding proteins. The current study examines the novel interactions between TRF2, TRF1, TIN2 and TPP1 and molecular chaperones (Hsp90, Hsp70, p23). In vitro and in cell experiments have shown an interaction between TRF1 and TRF2 and the molecular chaperones Hsp90 and Hsp70. Inhibition of Hsp90 using drugs that specifically block ATPase activity results in an increased association of TRF1 and TRF2 with Hsp90 to presumably stabilize the telomere associated proteins to the telomere. A definitive explanation as to the mechanisms underlying the chaperone/telomere associated protein interaction has yet to be determined and further studies examining chaperones’ contribution to telomere structure and function are underway. A better understanding of the telomeric proteins and Hsp90 and their roles in nuclear events is important, as both have extremely important functions in the cell. Our current working hypothesis is that chaperone proteins associate with TRF2, TRF1, TIN2 and TPP1 to facilitate telomeric protein-protein interactions and protein-telomere binding in both cancer and normal cells. The interaction between chaperones and telomere binding proteins may eventually provide a better understanding of telomeric structure and function. Defining the mechanisms of telomeric protein regulation is important in the development of new therapeutic approaches for targeting telomeres to induce dysfunction. Clinical trials are underway employing drugs targeting Hsp90 in cancer cells and given the results here, these Hsp90 compounds likely cause telomere alterations.

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Heyse, Serena R. "Functional divergence between Tetrahymena telomere proteins: Potential role for POT1b in chromosome breakage and new telomere synthesis." University of Cincinnati / OhioLINK, 2011. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1299168952.

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Mangosh, Tawna L. "SLX4 Interacting Protein (SLX4IP): A Vital Primer for Alternative Lengthening of Telomere (ALT)-like Processes Promoting Replicative Immortality in Castration-resistant Prostate Cancer with Androgen Receptor Loss." Case Western Reserve University School of Graduate Studies / OhioLINK, 2021. http://rave.ohiolink.edu/etdc/view?acc_num=case1623255136624147.

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Baker, Asmaa M. "New Roles For TRF2 In Chromatin Architecture." Scholarly Repository, 2008. http://scholarlyrepository.miami.edu/oa_dissertations/164.

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Telomeres are specialized nucleoprotein structures found at the end of eukaryotic chromosomes. The telomere DNA in humans is composed of the sequence "5'-TTAGGG-3'" tandemly repeated in a stretch of 5-30kb of double stranded DNA. TTAGGG Repeat Factor 2 (TRF2) is a telomere DNA binding protein that has a critical role in telomere end protection. The current model for telomere protection by TRF2 is through its ability to remodel telomeres into looped higher-order structures, called the t-loop, which sequesters the end from DNA damage sensors. Since telomeres are known to be comprised of nucleosomal chromatin, it is important to determine how TRF2 binds to and affects the structure of nucleosomal arrays. The ability of TRF2 to bind to unusual DNA structures such as the t-loop and the single stranded/double (ss/ds) stranded telomere DNA junction may facilitate its binding to DNA in the form of nucleosomal arrays and promote higher-order chromatin structures. In this study, we have reconstituted a 2kb DNA fragment containing 550bp of telomere DNA into nucleosomal arrays and tested the binding of full-lengthTRF2 and four truncation mutants to telomeric nucleosomal arrays. Our data indicates that TRF2 and its truncation mutants bind to telomere nucleosomal arrays as well as it binds to telomere DNA. We used a novel electrophoretic technique, Analytical Agarose Gel Electrophoresis (AAGE), to measure changes in surface charge density, hydrodynamic radius, and conformational flexibility of DNA and nucleosomal arrays upon protein binding. Our results indicate that the C-terminal DNA binding Myb/SANT domain of TRF2 might be rearranging nucleosomal structure through either nucleosome sliding, unwrapping, or changing the arrangement of the linker DNA, while the N-terminal basic DNA binding region is causing nucleosomal arrays compaction. Instead of significant compaction, histone-free DNA undergoes DNA condensation and self-association. This activity is observed with the full-length protein and all regions of the protein, with the exception of TRF2-DBD, participate in the process. We speculate that the ability of TRF2-DBD to rearrange nucleosomal structure and N-terminal basic region to cause nucleosomal fiber compaction may allow TRF2 to promote t-loop formation in the context of chromatin. We propose that TRF2, possessing all the features, has a new role at telomeres as a chromatin architectural protein.

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Johansson, Linnéa. "Kan fysisk aktivitet påverka längden av telomerer?" Thesis, Linnéuniversitetet, Institutionen för kemi och biomedicin (KOB), 2014. http://urn.kb.se/resolve?urn=urn:nbn:se:lnu:diva-38477.

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Telomerer består av specifika DNA- sekvenser samt proteiner i de yttersta ändarna på våra kromosomer, och skyddar arvsmassan som kromosomerna innehåller. Telomerer har fått mycket uppmärksamhet i studier kring åldrande på senare tid. Detta eftersom telomerer förkortas för varje celldelning, vilket kan leda till att cellen slutar dela sig vid en viss kritisk längd. Korta telomerer har kopplats till åldersrelaterade sjukdomar som exempelvis hjärtsvikt. Fysisk aktivitet i sin tur minskar risken att drabbas av åldersrelaterade sjukdomar. Syftet med arbetet var att undersöka om det finns belägg i den vetenskapliga litteraturen för ett samband mellan fysisk aktiviet och telomerlängd hos människor. Artiklar söktes i databasen PubMed. Inklusionskriterier var att artiklarna skulle undersöka effekter på vita blodkroppar (leukocyter), samt undersöka långtidseffekter av fysisk aktivitet. Totalt nio artiklar inkluderades till den tänkta undersökningen. Åtta studiers resultat tyder på att det finns ett samband mellan fysisk aktivitet och telomerlängd, medan en av studierna inte påvisar något samband. Fysisk aktivitet av måttlig intensitet verkar bevara telomerlängd, medan inaktivitet verkar förkorta telomererna i snabbare takt. Det råder en oenighet huruvida fysisk aktivitet av tyngre intensitet påverkar telomerlängd. Mer forskning krävs för att mer konkreta slutsatser ska kunna dras.
Telomeres consist of specific sequences of DNA and proteins at the ends of our chromosomes, and protects the genetic data they hold. Lately, attention has been focused on telomeres in research related to the aging process. Telomeres shorten during every cell division, and might reach a critical level when the cell stops dividing. Short telomeres are associated with several age- related diseases, for example cardiovascular disease. Physical activity reduces the risk of many age- related diseases. The purpose of this thesis was to investigate if physical activity has an effect on human telomeres. This was done by a review of the scientific literature. The database PubMed was used to search for articles. Articles in the database were included in the study if they investigated the long term effects of physical activity on telomere length in white blood cells (leukocytes). Nine articles were found that meet the inclusion criteria. The results from eight of them seems to point towards an association between psysical activity and telomere length, while one of the studies sees no such associasion. Physical activity by moderate intensity seems to protect telomeres from shortening, while inactivity seems to speed the telomere shortening process. There is some inconsistency about how high intensity of physical activity affects telomeres. More research is necessary to be able to draw further conclusions about the relationship between physical activity and telomere length.

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Dissertations / Theses on the topic 'Telomeren'

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