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1.
Nucleic Acids Res ; 51(22): 12242-12260, 2023 Dec 11.
Artículo en Inglés | MEDLINE | ID: mdl-37930826

RESUMEN

Telomeric repeat-containing RNA (TERRA) and its formation of RNA:DNA hybrids (or TERRA R-loops), influence telomere maintenance, particularly in human cancer cells that use homologous recombination-mediated alternative lengthening of telomeres. Here, we report that the RNA-binding motif protein 14 (RBM14) is associated with telomeres in human cancer cells. RBM14 negatively regulates TERRA expression. It also binds to TERRA and inhibits it from forming TERRA R-loops at telomeres. RBM14 depletion has several effects, including elevated TERRA levels, telomeric R-loops, telomere dysfunction-induced DNA damage foci formation, particularly in the presence of DNA replication stress, pRPA32 accumulation at telomeres and telomere signal-free ends. Thus, RBM14 protects telomere integrity via modulating TERRA levels and its R-loop formation at telomeres.


Asunto(s)
Homeostasis del Telómero , Telómero , Humanos , ADN/genética , Estructuras R-Loop , ARN/genética , ARN/metabolismo , ARN Largo no Codificante/genética , Motivos de Unión al ARN , Telómero/genética , Telómero/metabolismo , Neoplasias/genética
2.
PLoS Genet ; 18(11): e1010506, 2022 11.
Artículo en Inglés | MEDLINE | ID: mdl-36441670

RESUMEN

Short telomeres induce a DNA damage response (DDR) that evokes apoptosis and senescence in human cells. An extant question is the contribution of telomere dysfunction-induced DDR to the phenotypes observed in aging and telomere biology disorders. One candidate is RAP1, a telomere-associated protein that also controls transcription at extratelomeric regions. To distinguish these roles, we generated a knockin mouse carrying a mutated Rap1, which was incapable of binding telomeres and did not result in eroded telomeres or a DDR. Primary Rap1 knockin embryonic fibroblasts showed decreased RAP1 expression and re-localization away from telomeres, with an increased cytosolic distribution akin to that observed in human fibroblasts undergoing telomere erosion. Rap1 knockin mice were viable, but exhibited transcriptomic alterations, proinflammatory cytokine/chemokine signaling, reduced lifespan, and decreased healthspan with increased body weight/fasting blood glucose levels, spontaneous tumor incidence, and behavioral deficits. Taken together, our data present mechanisms distinct from telomere-induced DDR that underlie age-related phenotypes.


Asunto(s)
Complejo Shelterina , Telómero , Animales , Humanos , Ratones , Longevidad , Fenotipo , Telómero/genética , Acortamiento del Telómero
3.
EMBO J ; 39(21): e103420, 2020 11 02.
Artículo en Inglés | MEDLINE | ID: mdl-32935380

RESUMEN

Short telomeres are a principal defining feature of telomere biology disorders, such as dyskeratosis congenita (DC), for which there are no effective treatments. Here, we report that primary fibroblasts from DC patients and late generation telomerase knockout mice display lower nicotinamide adenine dinucleotide (NAD) levels, and an imbalance in the NAD metabolome that includes elevated CD38 NADase and reduced poly(ADP-ribose) polymerase and SIRT1 activities, respectively, affecting many associated biological pathways. Supplementation with the NAD precursor, nicotinamide riboside, and CD38 inhibition improved NAD homeostasis, thereby alleviating telomere damage, defective mitochondrial biosynthesis and clearance, cell growth retardation, and cellular senescence of DC fibroblasts. These findings reveal a direct, underlying role of NAD dysregulation when telomeres are short and underscore its relevance to the pathophysiology and interventions of human telomere-driven diseases.


Asunto(s)
Disqueratosis Congénita/genética , Disqueratosis Congénita/metabolismo , Fibroblastos/metabolismo , NAD/metabolismo , Telomerasa/genética , Telómero/metabolismo , ADP-Ribosil Ciclasa 1/genética , Animales , Encéfalo/patología , Línea Celular , Senescencia Celular , Disqueratosis Congénita/patología , Femenino , Homeostasis , Humanos , Glicoproteínas de Membrana/genética , Ratones , Ratones Noqueados , Mitocondrias/metabolismo , Niacinamida/análogos & derivados , Niacinamida/metabolismo , Fenotipo , Poli(ADP-Ribosa) Polimerasa-1/metabolismo , Compuestos de Piridinio/metabolismo , Telomerasa/metabolismo
4.
Nucleic Acids Res ; 46(11): 5664-5677, 2018 06 20.
Artículo en Inglés | MEDLINE | ID: mdl-29750271

RESUMEN

Base excision repair (BER) handles many forms of endogenous DNA damage, and apurinic/apyrimidinic endonuclease 1 (APE1) is central to this process. Deletion of both alleles of APE1 (a.k.a. Apex1) in mice leads to embryonic lethality, and deficiency in cells can promote cell death. Unlike most other BER proteins, APE1 expression is inversely correlated with cellular senescence in primary human fibroblasts. Depletion of APE1 via shRNA induced senescence in normal human BJ fibroblasts, a phenotype that was not seen in counterpart cells expressing telomerase. APE1 knock-down in primary fibroblasts resulted in global DNA damage accumulation, and the induction of p16INK4a and p21WAF1 stress response pathways; the DNA damage response, as assessed by γ-H2AX, was particularly pronounced at telomeres. Conditional knock-out of Apex1 in mice at post-natal day 7/12 resulted in impaired growth, reduced organ size, and increased cellular senescence. The effect of Apex1 deletion at post-natal week 6 was less obvious, other than cellular senescence, until ∼8-months of age, when premature aging characteristics, such as hair loss and impaired wound healing, were seen. Low APE1 expression in patient cancer tissue also correlated with increased senescence. Our results point to a key role for APE1 in regulating cellular senescence and aging features, with telomere status apparently affecting the outcome.


Asunto(s)
Senescencia Celular , ADN-(Sitio Apurínico o Apirimidínico) Liasa/metabolismo , Envejecimiento Prematuro/genética , Animales , Células Cultivadas , Daño del ADN , Reparación del ADN , ADN-(Sitio Apurínico o Apirimidínico) Liasa/deficiencia , ADN-(Sitio Apurínico o Apirimidínico) Liasa/genética , Fibroblastos/metabolismo , Ratones Noqueados , Telomerasa/metabolismo , Telómero/metabolismo
5.
Br J Haematol ; 181(3): 372-377, 2018 05.
Artículo en Inglés | MEDLINE | ID: mdl-29693246

RESUMEN

In a previous whole exome sequencing of patients from 41 families with Hodgkin lymphoma, we identified two families with distinct heterozygous rare coding variants in POT1 (D224N and Y36H), both in a highly conserved region of the gene. POT1 D224N mutant did not bind to a single-stranded telomere oligonucleotide in vitro suggesting the mutation perturbs POT1's ability to bind to the telomeric G-rich overhang. Human HT1080 cells expressing POT1 D224N and lymphoblastoid cells carrying Y36H both showed increased telomere length and fragility in comparison to wild type cells. This strongly suggests that mutant POT1 causes chromosome instability and may play a role in lymphomagenesis in these families.


Asunto(s)
Inestabilidad Cromosómica , Familia , Mutación de Línea Germinal , Enfermedad de Hodgkin , Mutación Missense , Proteínas de Unión a Telómeros , Sustitución de Aminoácidos , Línea Celular Tumoral , Femenino , Enfermedad de Hodgkin/genética , Enfermedad de Hodgkin/metabolismo , Enfermedad de Hodgkin/patología , Humanos , Masculino , Complejo Shelterina , Telómero/genética , Telómero/metabolismo , Proteínas de Unión a Telómeros/genética , Proteínas de Unión a Telómeros/metabolismo
6.
Nucleic Acids Res ; 44(10): 4871-80, 2016 06 02.
Artículo en Inglés | MEDLINE | ID: mdl-27131364

RESUMEN

The Fanconi anemia protein SLX4 assembles a genome and telomere maintenance toolkit, consisting of the nucleases SLX1, MUS81 and XPF. Although it is known that SLX4 acts as a scaffold for building this complex, the molecular basis underlying this function of SLX4 remains unclear. Here, we report that functioning of SLX4 is dependent on its dimerization via an oligomerization motif called the BTB domain. We solved the crystal structure of the SLX4BTB dimer, identifying key contacts (F681 and F708) that mediate dimerization. Disruption of BTB dimerization abrogates nuclear foci formation and telomeric localization of not only SLX4 but also of its associated nucleases. Furthermore, dimerization-deficient SLX4 mutants cause defective cellular response to DNA interstrand crosslinking agent and telomere maintenance, underscoring the contribution of BTB domain-mediated dimerization of SLX4 in genome and telomere maintenance.


Asunto(s)
Endonucleasas/metabolismo , Recombinasas/química , Línea Celular , Interacciones Hidrofóbicas e Hidrofílicas , Mitomicina/toxicidad , Dominios Proteicos , Multimerización de Proteína , Recombinasas/metabolismo , Telómero/enzimología , Telómero/ultraestructura
7.
Nucleic Acids Res ; 43(12): 5912-23, 2015 Jul 13.
Artículo en Inglés | MEDLINE | ID: mdl-25990736

RESUMEN

SLX4 assembles a toolkit of endonucleases SLX1, MUS81 and XPF, which is recruited to telomeres via direct interaction of SLX4 with TRF2. Telomeres present an inherent obstacle for DNA replication and repair due to their high propensity to form branched DNA intermediates. Here we provide novel insight into the mechanism and regulation of the SLX4 complex in telomere preservation. SLX4 associates with telomeres throughout the cell cycle, peaking in late S phase and under genotoxic stress. Disruption of SLX4's interaction with TRF2 or SLX1 and SLX1's nuclease activity independently causes telomere fragility, suggesting a requirement of the SLX4 complex for nucleolytic resolution of branched intermediates during telomere replication. Indeed, the SLX1-SLX4 complex processes a variety of telomeric joint molecules in vitro. The nucleolytic activity of SLX1-SLX4 is negatively regulated by telomeric DNA-binding proteins TRF1 and TRF2 and is suppressed by the RecQ helicase BLM in vitro. In vivo, in the presence of functional BLM, telomeric circle formation and telomere sister chromatid exchange, both arising out of nucleolytic processing of telomeric homologous recombination intermediates, are suppressed. We propose that the SLX4-toolkit is a telomere accessory complex that, in conjunction with other telomere maintenance proteins, ensures unhindered, but regulated telomere maintenance.


Asunto(s)
Recombinasas/metabolismo , Telómero/metabolismo , Ciclo Celular , ADN/metabolismo , Endodesoxirribonucleasas , Endonucleasas/metabolismo , Células HeLa , Recombinación Homóloga , Humanos , RecQ Helicasas/metabolismo , Intercambio de Cromátides Hermanas , Proteínas de Unión a Telómeros/metabolismo
8.
J Biol Chem ; 290(9): 5502-11, 2015 Feb 27.
Artículo en Inglés | MEDLINE | ID: mdl-25572391

RESUMEN

Uracil in the genome can result from misincorporation of dUTP instead of dTTP during DNA synthesis, and is primarily removed by uracil DNA glycosylase (UNG) during base excision repair. Telomeres contain long arrays of TTAGGG repeats and may be susceptible to uracil misincorporation. Using model telomeric DNA substrates, we showed that the position and number of uracil substitutions of thymine in telomeric DNA decreased recognition by the telomere single-strand binding protein, POT1. In primary mouse hematopoietic cells, uracil was detectable at telomeres, and UNG deficiency further increased uracil loads and led to abnormal telomere lengthening. In UNG-deficient cells, the frequencies of sister chromatid exchange and fragility in telomeres also significantly increased in the absence of telomerase. Thus, accumulation of uracil and/or UNG deficiency interferes with telomere maintenance, thereby underscoring the necessity of UNG-initiated base excision repair for the preservation of telomere integrity.


Asunto(s)
Células de la Médula Ósea/metabolismo , Reparación del ADN , Telómero/metabolismo , Uracilo/metabolismo , Animales , Secuencia de Bases , Células Cultivadas , Daño del ADN , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Hibridación Fluorescente in Situ , Ratones Noqueados , Unión Proteica , Complejo Shelterina , Telómero/genética , Homeostasis del Telómero/genética , Proteínas de Unión a Telómeros , Proteína 1 de Unión a Repeticiones Teloméricas/genética , Proteína 1 de Unión a Repeticiones Teloméricas/metabolismo , Proteína 2 de Unión a Repeticiones Teloméricas/genética , Proteína 2 de Unión a Repeticiones Teloméricas/metabolismo , Timina/metabolismo , Uracil-ADN Glicosidasa/deficiencia , Uracil-ADN Glicosidasa/genética
9.
Nucleic Acids Res ; 42(9): 5671-88, 2014 May.
Artículo en Inglés | MEDLINE | ID: mdl-24623817

RESUMEN

A variety of human tumors employ alternative and recombination-mediated lengthening for telomere maintenance (ALT). Human RecQ helicases, such as BLM and WRN, can efficiently unwind alternate/secondary structures during telomere replication and/or recombination. Here, we report a novel role for RECQL1, the most abundant human RecQ helicase but functionally least studied, in telomere maintenance. RECQL1 associates with telomeres in ALT cells and actively resolves telomeric D-loops and Holliday junction substrates. RECQL1 physically and functionally interacts with telomere repeat-binding factor 2 that in turn regulates its helicase activity on telomeric substrates. The telomeric single-stranded binding protein, protection of telomeres 1 efficiently stimulates RECQL1 on telomeric substrates containing thymine glycol, a replicative blocking lesion. Loss of RECQL1 results in dysfunctional telomeres, telomere loss and telomere shortening, elevation of telomere sister-chromatid exchanges and increased aphidicolin-induced telomere fragility, indicating a role for RECQL1 in telomere maintenance. Further, our results indicate that RECQL1 may participate in the same pathway as WRN, probably in telomere replication.


Asunto(s)
RecQ Helicasas/fisiología , Homeostasis del Telómero , Animales , Replicación del ADN , Ensayo de Cambio de Movilidad Electroforética , Exodesoxirribonucleasas/metabolismo , Células HeLa , Humanos , Unión Proteica , Transporte de Proteínas , RecQ Helicasas/metabolismo , Telomerasa/metabolismo , Proteína 2 de Unión a Repeticiones Teloméricas/metabolismo , Helicasa del Síndrome de Werner
10.
PLoS Genet ; 9(7): e1003639, 2013.
Artículo en Inglés | MEDLINE | ID: mdl-23874233

RESUMEN

Telomeres are chromosome end structures and are essential for maintenance of genome stability. Highly repetitive telomere sequences appear to be susceptible to oxidative stress-induced damage. Oxidation may therefore have a severe impact on telomere integrity and function. A wide spectrum of oxidative pyrimidine-derivatives has been reported, including thymine glycol (Tg), that are primarily removed by a DNA glycosylase, Endonuclease III-like protein 1 (Nth1). Here, we investigate the effect of Nth1 deficiency on telomere integrity in mice. Nth1 null (Nth1(-/-) ) mouse tissues and primary MEFs harbor higher levels of Endonuclease III-sensitive DNA lesions at telomeric repeats, in comparison to a non-telomeric locus. Furthermore, oxidative DNA damage induced by acute exposure to an oxidant is repaired slowly at telomeres in Nth1(-/-) MEFs. Although telomere length is not affected in the hematopoietic tissues of Nth1(-/-) adult mice, telomeres suffer from attrition and increased recombination and DNA damage foci formation in Nth1(-/-) bone marrow cells that are stimulated ex vivo in the presence of 20% oxygen. Nth1 deficiency also enhances telomere fragility in mice. Lastly, in a telomerase null background, Nth1(-/-) bone marrow cells undergo severe telomere loss at some chromosome ends and cell apoptosis upon replicative stress. These results suggest that Nth1 plays an important role in telomere maintenance and base repair against oxidative stress-induced base modifications. The fact that telomerase deficiency can exacerbate telomere shortening in Nth1 deficient mouse cells supports that base excision repair cooperates with telomerase to maintain telomere integrity.


Asunto(s)
Daño del ADN , Desoxirribonucleasa (Dímero de Pirimidina)/genética , Inestabilidad Genómica , Telómero/genética , Animales , Células de la Médula Ósea/metabolismo , Células de la Médula Ósea/patología , Cromosomas/genética , Cromosomas/metabolismo , Cromosomas/ultraestructura , Ratones , Ratones Noqueados , Estrés Oxidativo , Oxígeno/metabolismo , Recombinación Genética , Secuencias Repetitivas de Ácidos Nucleicos/genética , Telomerasa/genética , Telomerasa/metabolismo , Telómero/metabolismo , Telómero/patología
11.
EMBO J ; 29(2): 398-409, 2010 Jan 20.
Artículo en Inglés | MEDLINE | ID: mdl-19942858

RESUMEN

Telomeres consist of short guanine-rich repeats. Guanine can be oxidized to 8-oxo-7,8-dihydroguanine (8-oxoG) and 2,6-diamino-4-hydroxy-5-formamidopyrimidine (FapyG). 8-oxoguanine DNA glycosylase (Ogg1) repairs these oxidative guanine lesions through the base excision repair (BER) pathway. Here we show that in Saccharomyces cerevisiae ablation of Ogg1p leads to an increase in oxidized guanine level in telomeric DNA. The ogg1 deletion (ogg1Delta) strain shows telomere lengthening that is dependent on telomerase and/or Rad52p-mediated homologous recombination. 8-oxoG in telomeric repeats attenuates the binding of the telomere binding protein, Rap1p, to telomeric DNA in vitro. Moreover, the amount of telomere-bound Rap1p and Rif2p is reduced in ogg1Delta strain. These results suggest that oxidized guanines may perturb telomere length equilibrium by attenuating telomere protein complex to function in telomeres, which in turn impedes their regulation of pathways engaged in telomere length maintenance. We propose that Ogg1p is critical in maintaining telomere length homoeostasis through telomere guanine damage repair, and that interfering with telomere length homoeostasis may be one of the mechanism(s) by which oxidative DNA damage inflicts the genome.


Asunto(s)
ADN Glicosilasas/genética , Eliminación de Gen , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/genética , Telómero/metabolismo , ADN/metabolismo , ADN Glicosilasas/metabolismo , Guanina/metabolismo , Oxidación-Reducción , Unión Proteica , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Complejo Shelterina , Telómero/química , Proteínas de Unión a Telómeros/metabolismo , Factores de Transcripción/metabolismo
13.
Proc Natl Acad Sci U S A ; 108(39): 16434-9, 2011 Sep 27.
Artículo en Inglés | MEDLINE | ID: mdl-21903926

RESUMEN

Telomere repeat-binding factor 2 (TRF2) is critical for telomere integrity in dividing stem and somatic cells, but its role in postmitotic neurons is unknown. Apart from protecting telomeres, nuclear TRF2 interacts with the master neuronal gene-silencer repressor element 1-silencing transcription factor (REST), and disruption of this interaction induces neuronal differentiation. Here we report a developmental switch from the expression of TRF2 in proliferating neural progenitor cells to expression of a unique short nontelomeric isoform of TRF2 (TRF2-S) as neurons establish a fully differentiated state. Unlike nuclear TRF2, which enhances REST-mediated gene repression, TRF2-S is located in the cytoplasm where it sequesters REST, thereby maintaining the expression of neuronal genes, including those encoding glutamate receptors, cell adhesion, and neurofilament proteins. In neurons, TRF2-S-mediated antagonism of REST nuclear activity is greatly attenuated by either overexpression of TRF2 or administration of the excitatory amino acid kainic acid. Overexpression of TRF2-S rescues kainic acid-induced REST nuclear accumulation and its gene-silencing effects. Thus, TRF2-S acts as part of a unique developmentally regulated molecular switch that plays critical roles in the maintenance and plasticity of neurons.


Asunto(s)
Silenciador del Gen , Neuronas/citología , Proteínas Represoras/metabolismo , Proteínas de Unión a Telómeros/fisiología , Telómero , Factores de Transcripción/metabolismo , Animales , Encéfalo/crecimiento & desarrollo , Encéfalo/metabolismo , Humanos , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Proteínas de Unión a Telómeros/genética
14.
J Biol Chem ; 287(1): 196-209, 2012 Jan 02.
Artículo en Inglés | MEDLINE | ID: mdl-22039056

RESUMEN

Telomeres are structures at the ends of chromosomes and are composed of long tracks of short tandem repeat DNA sequences bound by a unique set of proteins (shelterin). Telomeric DNA is believed to form G-quadruplex and D-loop structures, which presents a challenge to the DNA replication and repair machinery. Although the RecQ helicases WRN and BLM are implicated in the resolution of telomeric secondary structures, very little is known about RECQL4, the RecQ helicase mutated in Rothmund-Thomson syndrome (RTS). Here, we report that RTS patient cells have elevated levels of fragile telomeric ends and that RECQL4-depleted human cells accumulate fragile sites, sister chromosome exchanges, and double strand breaks at telomeric sites. Further, RECQL4 localizes to telomeres and associates with shelterin proteins TRF1 and TRF2. Using recombinant proteins we showed that RECQL4 resolves telomeric D-loop structures with the help of shelterin proteins TRF1, TRF2, and POT1. We also found a novel functional synergistic interaction of this protein with WRN during D-loop unwinding. These data implicate RECQL4 in telomere maintenance.


Asunto(s)
Proteínas Mutantes/metabolismo , Mutación , RecQ Helicasas/metabolismo , Síndrome Rothmund-Thomson/genética , Telómero/metabolismo , Afidicolina/farmacología , Secuencia de Bases , ADN/biosíntesis , ADN/química , ADN/metabolismo , Replicación del ADN/efectos de los fármacos , Exodesoxirribonucleasas/metabolismo , Técnicas de Silenciamiento del Gen , Células HeLa , Humanos , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Datos de Secuencia Molecular , Proteínas Mutantes/genética , Conformación de Ácido Nucleico/efectos de los fármacos , Transporte de Proteínas/efectos de los fármacos , ARN Interferente Pequeño/genética , RecQ Helicasas/deficiencia , RecQ Helicasas/genética , Síndrome Rothmund-Thomson/metabolismo , Síndrome Rothmund-Thomson/patología , Telómero/efectos de los fármacos , Telómero/genética , Proteína 2 de Unión a Repeticiones Teloméricas/metabolismo , Proteína 1 de Unión al Supresor Tumoral P53 , Helicasa del Síndrome de Werner
15.
Nucleic Acids Res ; 39(18): 7992-8004, 2011 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-21737425

RESUMEN

Oxidative DNA damage plays a role in disease development and the aging process. A prominent participant in orchestrating the repair of oxidative DNA damage, particularly single-strand breaks, is the scaffold protein XRCC1. A series of chronological and biological aging parameters in XRCC1 heterozygous (HZ) mice were examined. HZ and wild-type (WT) C57BL/6 mice exhibit a similar median lifespan of ~26 months and a nearly identical maximal life expectancy of ~37 months. However, a number of HZ animals (7 of 92) showed a propensity for abdominal organ rupture, which may stem from developmental abnormalities given the prominent role of XRCC1 in endoderm and mesoderm formation. For other end-points evaluated-weight, fat composition, blood chemistries, condition of major organs, tissues and relevant cell types, behavior, brain volume and function, and chromosome and telomere integrity-HZ mice exhibited by-and-large a normal phenotype. Treatment of animals with the alkylating agent azoxymethane resulted in both liver toxicity and an increased incidence of precancerous lesions in the colon of HZ mice. Our study indicates that XRCC1 haploinsufficiency in mammals has little effect on chronological longevity and many key biological markers of aging in the absence of environmental challenges, but may adversely affect normal animal development or increase disease susceptibility to a relevant genotoxic exposure.


Asunto(s)
Envejecimiento/genética , Proteínas de Unión al ADN/genética , Haploinsuficiencia , Alquilantes/toxicidad , Animales , Conducta Animal , Peso Corporal , Células de la Médula Ósea/efectos de los fármacos , Encéfalo/anatomía & histología , Encéfalo/metabolismo , Supervivencia Celular/efectos de los fármacos , Enfermedad Hepática Inducida por Sustancias y Drogas/patología , Susceptibilidad a Enfermedades , Femenino , Inestabilidad Genómica , Masculino , Ratones , Ratones Endogámicos C57BL , Mutágenos/toxicidad , Proteína 1 de Reparación por Escisión del Grupo de Complementación Cruzada de las Lesiones por Rayos X
16.
PLoS Genet ; 6(5): e1000951, 2010 May 13.
Artículo en Inglés | MEDLINE | ID: mdl-20485567

RESUMEN

8-oxo-7,8-dihydroguanine (8-oxoG) and 2,6-diamino-4-hydroxy-5-formamidopyrimidine (FapyG) are among the most common oxidative DNA lesions and are substrates for 8-oxoguanine DNA glycosylase (OGG1)-initiated DNA base excision repair (BER). Mammalian telomeres consist of triple guanine repeats and are subject to oxidative guanine damage. Here, we investigated the impact of oxidative guanine damage and its repair by OGG1 on telomere integrity in mice. The mouse cells were analyzed for telomere integrity by telomere quantitative fluorescence in situ hybridization (telomere-FISH), by chromosome orientation-FISH (CO-FISH), and by indirect immunofluorescence in combination with telomere-FISH and for oxidative base lesions by Fpg-incision/Southern blot assay. In comparison to the wild type, telomere lengthening was observed in Ogg1 null (Ogg1(-/-)) mouse tissues and primary embryonic fibroblasts (MEFs) cultivated in hypoxia condition (3% oxygen), whereas telomere shortening was detected in Ogg1(-/-) mouse hematopoietic cells and primary MEFs cultivated in normoxia condition (20% oxygen) or in the presence of an oxidant. In addition, telomere length abnormalities were accompanied by altered telomere sister chromatid exchanges, increased telomere single- and double-strand breaks, and preferential telomere lagging- or G-strand losses in Ogg1(-/-) mouse cells. Oxidative guanine lesions were increased in telomeres in Ogg1(-/-) mice with aging and primary MEFs cultivated in 20% oxygen. Furthermore, oxidative guanine lesions persisted at high level in Ogg1(-/-) MEFs after acute exposure to hydrogen peroxide, while they rapidly returned to basal level in wild-type MEFs. These findings indicate that oxidative guanine damage can arise in telomeres where it affects length homeostasis, recombination, DNA replication, and DNA breakage repair. Our studies demonstrate that BER pathway is required in repairing oxidative guanine damage in telomeres and maintaining telomere integrity in mammals.


Asunto(s)
Guanina/química , Guanosina/análogos & derivados , Telómero , Animales , Células Cultivadas , Técnica del Anticuerpo Fluorescente Indirecta , Guanosina/química , Hibridación Fluorescente in Situ , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Oxidación-Reducción , Estrés Oxidativo , Recombinación Genética , Intercambio de Cromátides Hermanas
17.
Cancers (Basel) ; 15(7)2023 Apr 06.
Artículo en Inglés | MEDLINE | ID: mdl-37046839

RESUMEN

Telomeric repeat containing RNA (TERRA) is transcribed from subtelomeric regions to telomeres. TERRA RNA can invade telomeric dsDNA and form telomeric R-loop structures. A growing body of evidence suggests that TERRA-mediated R-loops are critical players in telomere length homeostasis. Here, we will review current knowledge on the regulation of R-loop levels at telomeres. In particular, we will discuss how the central player TERRA and its binding proteins modulate R-loop levels through various mechanisms. We will further provide an overview of the consequences of TERRA-mediated persistent or unscheduled R-loops at telomeres in human ALT cancers and other organisms, with a focus on telomere length regulation after replication interference-induced damage and DNA homologous recombination-mediated repair.

18.
Geroscience ; 45(4): 2213-2228, 2023 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-36826621

RESUMEN

Short telomeres are a defining feature of telomere biology disorders (TBDs), including dyskeratosis congenita (DC), for which there is no effective general cure. Patients with TBDs often experience bone marrow failure. NAD, an essential metabolic coenzyme, is decreased in models of DC. Herein, using telomerase reverse transcriptase null (Tert-/-) mice with critically short telomeres, we investigated the effect of NAD supplementation with the NAD precursor, nicotinamide riboside (NR), on features of health span disrupted by telomere impairment. Our results revealed that NR ameliorated body weight loss in Tert-/- mice and improved telomere integrity and telomere dysfunction-induced systemic inflammation. NR supplementation also mitigated myeloid skewing of Tert-/- hematopoietic stem cells. Furthermore, NR alleviated villous atrophy and inflammation in the small intestine of Tert-/- transplant recipient mice. Altogether, our findings support NAD intervention as a potential therapeutic strategy to enhance aspects of health span compromised by telomere attrition.


Asunto(s)
Disqueratosis Congénita , Trasplante de Células Madre Hematopoyéticas , Humanos , Animales , Ratones , NAD , Telómero/metabolismo , Disqueratosis Congénita/genética , Disqueratosis Congénita/metabolismo , Inflamación
19.
J Extracell Biol ; 2(8)2023 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-37744304

RESUMEN

Extracellular vesicles and particles (EVPs) are secreted by organs across the body into different circulatory systems, including the bloodstream, and reflect pathophysiologic conditions of the organ. However, the heterogeneity of EVPs in the blood makes it challenging to determine their organ of origin. We hypothesized that small (s)EVPs (<100 nm in diameter) in the bloodstream carry distinctive protein signatures associated with each originating organ, and we investigated this possibility by studying the proteomes of sEVPs produced by six major organs (brain, liver, lung, heart, kidney, fat). We found that each organ contained distinctive sEVP proteins: 68 proteins were preferentially found in brain sEVPs, 194 in liver, 39 in lung, 15 in heart, 29 in kidney, and 33 in fat. Furthermore, we isolated sEVPs from blood and validated the presence of sEVP proteins associated with the brain (DPP6, SYT1, DNM1L), liver (FABPL, ARG1, ASGR1/2), lung (SFPTA1), heart (CPT1B), kidney (SLC31), and fat (GDN). We further discovered altered levels of these proteins in serum sEVPs prepared from old mice compared to young mice. In sum, we have cataloged sEVP proteins that can serve as potential biomarkers for organ identification in serum and show differential expression with age.

20.
J Clin Invest ; 133(12)2023 06 15.
Artículo en Inglés | MEDLINE | ID: mdl-37097759

RESUMEN

Senescent vascular smooth muscle cells (VSMCs) accumulate in the vasculature with age and tissue damage and secrete factors that promote atherosclerotic plaque vulnerability and disease. Here, we report increased levels and activity of dipeptidyl peptidase 4 (DPP4), a serine protease, in senescent VSMCs. Analysis of the conditioned media from senescent VSMCs revealed a unique senescence-associated secretory phenotype (SASP) signature comprising many complement and coagulation factors; silencing or inhibiting DPP4 reduced these factors and increased cell death. Serum samples from persons with high risk for cardiovascular disease contained high levels of DPP4-regulated complement and coagulation factors. Importantly, DPP4 inhibition reduced senescent cell burden and coagulation and improved plaque stability, while single-cell resolution of senescent VSMCs reflected the senomorphic and senolytic effects of DPP4 inhibition in murine atherosclerosis. We propose that DPP4-regulated factors could be exploited therapeutically to reduce senescent cell function, reverse senohemostasis, and improve vascular disease.


Asunto(s)
Aterosclerosis , Placa Aterosclerótica , Ratones , Animales , Placa Aterosclerótica/genética , Dipeptidil Peptidasa 4/genética , Dipeptidil Peptidasa 4/metabolismo , Senescencia Celular/genética , Músculo Liso Vascular/metabolismo , Aterosclerosis/tratamiento farmacológico , Aterosclerosis/genética , Aterosclerosis/metabolismo
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