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1.
Nucleic Acids Res ; 48(4): 2189-2197, 2020 02 28.
Artigo em Inglês | MEDLINE | ID: mdl-31950178

RESUMO

Understanding the mechanism by which biological macromolecules fold into their functional native conformations represents a problem of fundamental interest. DNA oligonucleotides derived from human telomeric repeat d[TAGGG(TTAGGG)3] and d[TAGGG(TTAGGG)3TT] fold into G-quadruplexes through diverse steps. Varying the pH and temperature by the use of nuclear magnetic resonance and other methods enabled detection of pre-folded structures that exist in solution before completely formed G-quadruplexes upon addition of cations. Pre-folded structures are in general hard to detect, however their knowledge is crucial to set up folding pathways into final structure since they are believed to be a starting point. Unexpectedly well-defined pre-folded structures composed of base triples for both oligonucleotides were detected at certain pH and temperature. These kinds of structures were up to now only hypothesized as intermediates in the folding process. All revealed pre-folded structures irrespective of the pH and temperature exhibited one common structural feature that could govern folding process.


Assuntos
DNA/química , Quadruplex G , Conformação de Ácido Nucleico , Telômero/genética , Dicroísmo Circular , DNA/genética , Humanos , Concentração de Íons de Hidrogênio , Ligantes , Modelos Moleculares , Oligonucleotídeos/química , Oligonucleotídeos/genética , Telômero/química , Temperatura Ambiente
2.
Genes Dev ; 34(1-2): 7-23, 2020 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-31896689

RESUMO

53BP1 is an enigmatic DNA damage response factor that gained prominence because it determines the efficacy of PARP1 inhibitory drugs (PARPi) in BRCA1-deficient cancers. Recent studies have elevated 53BP1 from its modest status of (yet another) DNA damage factor to master regulator of double-strand break (DSB) repair pathway choice. Our review of the literature suggests an alternative view. We propose that 53BP1 has evolved to avoid mutagenic repair outcomes and does so by controlling the processing of DNA ends and the dynamics of DSBs. The consequences of 53BP1 deficiency, such as diminished PARPi efficacy in BRCA1-deficient cells and altered repair of damaged telomeres, can be explained from this viewpoint. We further propose that some of the fidelity functions of 53BP1 coevolved with class switch recombination (CSR) in the immune system. We speculate that, rather than being deterministic in DSB repair pathway choice, 53BP1 functions as a DSB escort that guards against illegitimate and potentially tumorigenic recombination.


Assuntos
Quebras de DNA de Cadeia Dupla , Reparo do DNA/genética , Proteína 1 de Ligação à Proteína Supressora de Tumor p53/metabolismo , Evolução Molecular , Humanos , Switching de Imunoglobulina/genética , Telômero/genética , Proteína 1 de Ligação à Proteína Supressora de Tumor p53/deficiência , Proteína 1 de Ligação à Proteína Supressora de Tumor p53/genética
3.
J Cancer Res Clin Oncol ; 146(2): 381-389, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31960186

RESUMO

PURPOSE: To evaluate the prevalence of two recurrent somatic mutations (-124 C>T and -146 C>T) within the promoter of the gene encoding telomerase reverse transcriptase (TERT) as well as their relationship with TERT level, telomeres length, and outcome in patients with head and neck squamous cell carcinomas (HNSCCs). METHODS: We evaluate the prevalence of TERT promoter mutations, TERT levels, and telomere length in paired cancer tissue and adjacent mucosa (AM) in a series of HNSCCs. RESULTS: Cancer tissue and AM specimens from 105 patients were analyzed. Telomere length and TERT mRNA levels were estimated using real-time polymerase chain reaction. TERT promoter mutations were assessed using Sanger sequencing. Out of 105 cases, 101 were considered suitable for the analysis. TERT promoter harbored mutations in 12 tumors (11.9%), with -124 C>T and -146 C>T accounting for 83.3% and 16.7% of the alterations, respectively. No mutations were detected in AM samples. The prevalence of TERT promoter mutations was significantly higher in oral cavity SCCs (10 out of 27 tumors; 37%), and telomere length in AM was shorter in patients with tumors carrying TERT promoter mutations than in patients with unmutated TERT promoter cancers (p = 0.023). TERT levels in tumor did not significantly differ according to the mutational status of TERT promoter. No significant association was found between TERT promoter status and overall survival. CONCLUSION: TERT promoter mutations are most likely a late event in tumor development, occurring in a context of critically short telomeres, mostly in patients with oral cavity SCC. TERT levels, but not TERT promoter mutational status impact clinical outcome.


Assuntos
Neoplasias de Cabeça e Pescoço/genética , Mutação , Carcinoma de Células Escamosas de Cabeça e Pescoço/genética , Telomerase/genética , Telômero/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Estudos de Coortes , Feminino , Neoplasias de Cabeça e Pescoço/metabolismo , Neoplasias de Cabeça e Pescoço/patologia , Humanos , Masculino , Pessoa de Meia-Idade , Regiões Promotoras Genéticas , Estudos Prospectivos , Carcinoma de Células Escamosas de Cabeça e Pescoço/metabolismo , Carcinoma de Células Escamosas de Cabeça e Pescoço/patologia , Análise de Sobrevida , Telomerase/metabolismo , Telômero/genética , Telômero/patologia
4.
Chemosphere ; 238: 124863, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31551201

RESUMO

Peripheral blood leukocyte telomere length in omethoate-exposed workers is related to environmental exposure and single nucleotide polymorphisms (SNPs) in genes including p21, GSTM1, miR-145, etc. However, the roles of SNPs in tankyrase (TNKS) gene in telomere length are still unknown. The aim of this study was to explore the association between SNPs in TNKS gene and telomere length in omethoate-exposed workers. Telomere length in peripheral blood leukocyte DNA from 180 omethoate-exposed workers and 115 healthy controls was measured using Real-time quantitative polymerase chain reaction (PCR). Genotyping of the selected functional and susceptible SNPs was performed by the flight mass spectrometry based on PCR and single-base extension. The analysis of covariance was performed to find effects of SNPs on telomere length. Generalized linear models were used to analyze the environment, gene, and interaction on telomere length. The results showed that telomere length in the CG + CC genotypes in rs1055328 in TNKS gene was significantly longer than that in the wild homozygous GG genotype both in exposure group (P = 0.017) and in control group (P = 0.038) after adjusting the covariates. The variables kept in the generalized linear models included omethoate-exposure (ß = 0.580, P = 0.001) and rs1055328 (CG + CC) in TNKS gene (ß = 0.339, P = 0.002). The study suggests that the prolongation of telomere length is associated with omethoate-exposure and the CG + CC genotypes in rs1055328 in TNKS gene.


Assuntos
Dimetoato/análogos & derivados , Exposição Ocupacional/efeitos adversos , Tanquirases/genética , Homeostase do Telômero/efeitos dos fármacos , Telômero/fisiologia , Adulto , Inibidor de Quinase Dependente de Ciclina p21/genética , DNA/genética , Dano ao DNA/efeitos dos fármacos , Dimetoato/toxicidade , Feminino , Genótipo , Glutationa Transferase/genética , Humanos , Leucócitos/citologia , Masculino , MicroRNAs/genética , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único/genética , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Telômero/genética
5.
Cytogenet Genome Res ; 159(4): 208-214, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31846969

RESUMO

The karyotypes and other chromosomal markers of 4 catfish species, namely Lasiancistrus schomburgkii, Lasiancistrus sp., Araichthysloro, and Megalancistrus sp., members of a taxonomically complex and speciose tribe of catfishes Ancistrini, Hypostominae, were examined using conventional (Giemsa staining, Ag-NOR, and C-banding) and molecular cytogenetic protocols (FISH) and DNA barcoding. In L. schomburgkii, Lasiancistrus sp., and A.loro a diploid number 2n = 54 was observed, with karyotypes composed of 28m + 16sm + 10st, 36m + 12sm + 6st chromosomes, while Megalancistrus sp. had 2n = 52, with the karyotype composed of 28m + 16sm + 8st chromosomes. The Ag-NOR phenotypes were simple in all 4 species, which was confirmed by FISH with an 18S rDNA probe. However, the positive 5S rDNA sites varied among species: 2 chromosome pairs in L. schomburgkii, Lasiancistrus sp., and A. loro, and only 1 pair in Megalancistrus sp. The blocks of constitutive heterochromatin were poorly visible in the pericentromeric and telomeric regions of most chromosomes of the examined species by C-banding. The genetic distance analysis, based on mtDNA COI gene sequences (DNA barcoding), confirmed the species as 4 taxonomic units. Ours and other published data indicate that karyotype differentiation among Ancistrini is complex and divergent and indicates the occurrence of common chromosomal rearrangements, such as pericentric inversions conserving the diploid number, and other rearrangements that are more frequent in some genera, such as centric fusions in Ancistrus.


Assuntos
Peixes-Gato/genética , Animais , Inversão Cromossômica/genética , Cromossomos/genética , Citogenética/métodos , DNA Ribossômico/genética , Diploide , Feminino , Heterocromatina/genética , Cariótipo , Masculino , Filogenia , Telômero/genética
6.
Hum Genet ; 138(11-12): 1323-1330, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31677132

RESUMO

Phenotypic heterogeneity is often observed in patients with telomeropathies caused by pathogenic variants in telomere biology genes. However, the roles of recessive variants in these different phenotypes are not fully characterized. Our goal is to describe the biological roles of a novel homozygous RTEL1 variant identified in a consanguineous Lebanese family with unusual presentation of telomeropathies. A proband was screened for germline variants in telomere biology genes by whole exome sequencing. Leukocytes' telomere length was measured in the proband and eight relatives. We identified a novel homozygous p.E665K RTEL1 variant in the proband, his mother, and seven siblings that associated with telomere shortening and a broad spectrum of clinical manifestations, ranging from mild unspecific findings to severe phenotypes. Consanguinity in at least three family generations led to increased frequency of the homozygous p.E665K variant in the youngest generation and progressive telomere shortening. The increased frequency of the homozygous RTEL1 variant due to consanguinity in this Lebanese family allowed us to infer novel behaviors of recessive RTEL1 variants, as the expressivity and penetrance of this gene are very heterogenous between inter- and intra-generations. Progressive telomere shortening was associated with disease anticipation, first reported in recessive autosomal telomeropathies. Both genetic testing and telomere length measurement were critical for the clinical diagnosis of this family with telomere diseases marked by phenotypic heterogeneity.


Assuntos
Consanguinidade , DNA Helicases/genética , Doenças Genéticas Inatas/epidemiologia , Homozigoto , Mutação , Telômero/genética , Adolescente , Adulto , Feminino , Doenças Genéticas Inatas/genética , Humanos , Líbano/epidemiologia , Masculino , Pessoa de Meia-Idade , Linhagem , Fenótipo , Adulto Jovem
7.
PLoS Genet ; 15(11): e1008426, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31765372

RESUMO

DNA polymerase epsilon (Pol ε) is critical for genome duplication, but little is known about how post-translational modification regulates its function. Here we report that the Pol ε catalytic subunit Pol2 in yeast is sumoylated at a single lysine within a catalytic domain insertion uniquely possessed by Pol2 family members. We found that Pol2 sumoylation occurs specifically in S phase and is increased under conditions of replication fork blockade. Analyses of the genetic requirements of this modification indicate that Pol2 sumoylation is associated with replication fork progression and dependent on the Smc5/6 SUMO ligase known to promote DNA synthesis. Consistently, the pol2 sumoylation mutant phenotype suggests impaired replication progression and increased levels of gross chromosomal rearrangements. Our findings thus indicate a direct role for SUMO in Pol2-mediated DNA synthesis and a molecular basis for Smc5/6-mediated regulation of genome stability.


Assuntos
Proteínas de Ciclo Celular/genética , DNA Polimerase II/genética , DNA/biossíntese , Proteínas de Saccharomyces cerevisiae/genética , Sumoilação/genética , Domínio Catalítico/genética , DNA/genética , Replicação do DNA/genética , Instabilidade Genômica/genética , Lisina/genética , Complexos Multiproteicos/genética , Mutação/genética , Fase S/genética , Saccharomyces cerevisiae/genética , Proteínas Modificadoras Pequenas Relacionadas à Ubiquitina/genética , Telômero/genética
8.
PLoS Genet ; 15(10): e1008412, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31609962

RESUMO

During meiosis, each chromosome must selectively pair and synapse with its own unique homolog to enable crossover formation and subsequent segregation. How homolog pairing is maintained in early meiosis to ensure synapsis occurs exclusively between homologs is unknown. We aimed to further understand this process by examining the meiotic defects of a unique Drosophila mutant, Mcm5A7. We found that Mcm5A7 mutants are proficient in homolog pairing at meiotic onset yet fail to maintain pairing as meiotic synapsis ensues, causing seemingly normal synapsis between non-homologous loci. This pairing defect corresponds with a reduction of SMC1-dependent centromere clustering at meiotic onset. Overexpressing SMC1 in this mutant significantly restores centromere clustering, homolog pairing, and crossover formation. These data indicate that the initial meiotic pairing of homologs is not sufficient to yield synapsis exclusively between homologs and provide a model in which meiotic homolog pairing must be stabilized by centromeric SMC1 to ensure proper synapsis.


Assuntos
Proteínas de Ciclo Celular/genética , Centrômero/genética , Proteínas Cromossômicas não Histona/genética , Recombinação Homóloga/genética , Meiose/genética , Animais , Pareamento Cromossômico/genética , Segregação de Cromossomos/genética , Drosophila/genética , Complexo Sinaptonêmico , Telômero/genética
9.
Cytogenet Genome Res ; 159(1): 48-53, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31610539

RESUMO

Visualizing the spatiotemporal organization of the genome will improve our understanding of how chromatin structure and function are intertwined. Here, we describe a further development of the CRISPR/Cas9-based RNA-guided endonuclease-in situ labeling (RGEN-ISL) method. RGEN-ISL allowed the differentiation between vertebrate-type (TTAGGG)n and Arabidopsis-type (TTTAGGG)n telomere repeats. Using maize as an example, we established a combination of RGEN-ISL, immunostaining, and EdU labeling to visualize in situ specific repeats, histone marks, and DNA replication sites, respectively. The effects of the non-denaturing RGEN-ISL and standard denaturing FISH on the chromatin structure were compared using super-resolution microscopy. 3D structured illumination microscopy revealed that denaturation and acetic acid fixation impaired and flattened the chromatin. The broad range of adaptability of RGEN-ISL to different combinations of methods has the potential to advance the field of chromosome biology.


Assuntos
Amaryllidaceae/genética , Arabidopsis/genética , Sistemas CRISPR-Cas/genética , Replicação do DNA/genética , Zea mays/genética , Cromatina/metabolismo , Cromossomos/genética , DNA de Plantas/genética , Endonucleases/genética , Hibridização in Situ Fluorescente/métodos , RNA Guia/genética , Telômero/genética
10.
Int J Mol Sci ; 20(19)2019 Oct 08.
Artigo em Inglês | MEDLINE | ID: mdl-31597307

RESUMO

Senescence is defined as a stress-induced durable cell cycle arrest. We herein revisit the origin of two of these stresses, namely mitochondrial metabolic compromise, associated with reactive oxygen species (ROS) production, and replicative senescence, activated by extreme telomere shortening. We discuss how replication stress-induced DNA damage of telomeric DNA (telDNA) and mitochondrial DNA (mtDNA) can be considered a common origin of senescence in vitro, with consequences on ageing in vivo. Unexpectedly, mtDNA and telDNA share common features indicative of a high degree of replicative stress, such as G-quadruplexes, D-loops, RNA:DNA heteroduplexes, epigenetic marks, or supercoiling. To avoid these stresses, both compartments use similar enzymatic strategies involving, for instance, endonucleases, topoisomerases, helicases, or primases. Surprisingly, many of these replication helpers are active at both telDNA and mtDNA (e.g., RNAse H1, FEN1, DNA2, RecQ helicases, Top2α, Top2ß, TOP3A, DNMT1/3a/3b, SIRT1). In addition, specialized telomeric proteins, such as TERT (telomerase reverse transcriptase) and TERC (telomerase RNA component), or TIN2 (shelterin complex), shuttle from telomeres to mitochondria, and, by doing so, modulate mitochondrial metabolism and the production of ROS, in a feedback manner. Hence, mitochondria and telomeres use common weapons and cooperate to resist/prevent replication stresses, otherwise producing common consequences, namely senescence and ageing.


Assuntos
Replicação do DNA , DNA Mitocondrial/genética , Estresse Fisiológico/genética , Telômero/genética , Envelhecimento/genética , Animais , Senescência Celular/genética , Dano ao DNA , Epigênese Genética , Humanos , Mitocôndrias/genética , Mitocôndrias/metabolismo , Estresse Oxidativo , Homeostase do Telômero , Encurtamento do Telômero
11.
Nat Commun ; 10(1): 4723, 2019 10 17.
Artigo em Inglês | MEDLINE | ID: mdl-31624261

RESUMO

Short telomeres trigger age-related pathologies and shorter lifespans in mice and humans. In the past, we generated mouse embryonic (ES) cells with longer telomeres than normal (hyper-long telomeres) in the absence of genetic manipulations, which contributed to all mouse tissues. To address whether hyper-long telomeres have deleterious effects, we generated mice in which 100% of their cells are derived from hyper-long telomere ES cells. We observe that these mice have longer telomeres and less DNA damage with aging. Hyper-long telomere mice are lean and show low cholesterol and LDL levels, as well as improved glucose and insulin tolerance. Hyper-long telomere mice also have less incidence of cancer and an increased longevity. These findings demonstrate that longer telomeres than normal in a given species are not deleterious but instead, show beneficial effects.


Assuntos
Envelhecimento/genética , Células-Tronco Embrionárias/metabolismo , Longevidade/genética , Homeostase do Telômero/genética , Telômero/genética , Envelhecimento/metabolismo , Animais , Dano ao DNA , Humanos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Neoplasias/genética
12.
Int J Mol Sci ; 20(20)2019 Oct 09.
Artigo em Inglês | MEDLINE | ID: mdl-31601021

RESUMO

Cichlid fishes are the subject of scientific interest because of their rapid adaptive radiation, resulting in extensive ecological and taxonomic diversity. In this study, we examined 11 morphologically distinct cichlid species endemic to Barombi Mbo, the largest crater lake in western Cameroon, namely Konia eisentrauti, Konia dikume, Myaka myaka, Pungu maclareni, Sarotherodon steinbachi, Sarotherodon lohbergeri, Sarotherodon linnellii, Sarotherodon caroli, Stomatepia mariae, Stomatepia pindu, and Stomatepia mongo. These species supposedly evolved via sympatric ecological speciation from a common ancestor, which colonized the lake no earlier than one million years ago. Here we present the first comparative cytogenetic analysis of cichlid species from Barombi Mbo Lake using both conventional (Giemsa staining, C-banding, and CMA3/DAPI staining) and molecular (fluorescence in situ hybridization with telomeric, 5S, and 28S rDNA probes) methods. We observed stability on both macro and micro-chromosomal levels. The diploid chromosome number was 2n = 44, and the karyotype was invariably composed of three pairs of meta/submetacentric and 19 pairs of subtelo/acrocentric chromosomes in all analysed species, with the same numbers of rDNA clusters and distribution of heterochromatin. The results suggest the evolutionary stability of chromosomal set; therefore, the large-scale chromosomal rearrangements seem to be unlikely associated with the sympatric speciation in Barombi Mbo.


Assuntos
Adaptação Biológica/genética , Adaptação Biológica/efeitos da radiação , Instabilidade Cromossômica/efeitos da radiação , Ciclídeos/genética , Animais , Evolução Biológica , Camarões , Bandeamento Cromossômico , Mapeamento Cromossômico , Hibridização in Situ Fluorescente , Cariótipo , Cariotipagem , Lagos , Telômero/genética
13.
PLoS Genet ; 15(9): e1008368, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31518356

RESUMO

Telomere shortening is associated with aging and age-associated diseases. Additionally, telomere dysfunction resulting from telomerase gene mutation can lead to premature aging, such as apparent skin atrophy and hair loss. However, the molecular signaling linking telomere dysfunction to skin atrophy remains elusive. Here we show that dysfunctional telomere disrupts BMP/pSmad/P63 signaling, impairing epidermal stem cell specification and differentiation of skin and hair follicles. We find that telomere shortening mediated by Terc loss up-regulates Follistatin (Fst), inhibiting pSmad signaling and down-regulating P63 and epidermal keratins in an ESC differentiation model as well as in adult development of telomere-shortened mice. Mechanistically, short telomeres disrupt PRC2/H3K27me3-mediated repression of Fst. Our findings reveal that skin atrophy due to telomere dysfunction is caused by a previously unappreciated link with Fst and BMP signaling that could be explored in the development of therapies.


Assuntos
Células-Tronco/metabolismo , Encurtamento do Telômero/fisiologia , Animais , Atrofia/genética , Proteínas Morfogenéticas Ósseas/metabolismo , Diferenciação Celular/genética , Linhagem da Célula/genética , Proliferação de Células/genética , Células Epidérmicas/metabolismo , Epiderme/metabolismo , Regulação da Expressão Gênica/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Nus , Transdução de Sinais/genética , Proteínas Smad/metabolismo , Telômero/genética , Encurtamento do Telômero/genética , Transativadores/metabolismo
14.
Nat Commun ; 10(1): 4224, 2019 09 17.
Artigo em Inglês | MEDLINE | ID: mdl-31530811

RESUMO

Mitotic catastrophe is a broad descriptor encompassing unclear mechanisms of cell death. Here we investigate replication stress-driven mitotic catastrophe in human cells and identify that replication stress principally induces mitotic death signalled through two independent pathways. In p53-compromised cells we find that lethal replication stress confers WAPL-dependent centromere cohesion defects that maintain spindle assembly checkpoint-dependent mitotic arrest in the same cell cycle. Mitotic arrest then drives cohesion fatigue and triggers mitotic death through a primary pathway of BAX/BAK-dependent apoptosis. Simultaneously, a secondary mitotic death pathway is engaged through non-canonical telomere deprotection, regulated by TRF2, Aurora B and ATM. Additionally, we find that suppressing mitotic death in replication stressed cells results in distinct cellular outcomes depending upon how cell death is averted. These data demonstrate how replication stress-induced mitotic catastrophe signals cell death with implications for cancer treatment and cancer genome evolution.


Assuntos
Apoptose , Proteínas de Transporte/metabolismo , Replicação do DNA , Mitose , Neoplasias/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Telômero/metabolismo , Morte Celular , Linhagem Celular Tumoral , Humanos , Neoplasias/genética , Neoplasias/fisiopatologia , Telômero/genética , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Proteína Killer-Antagonista Homóloga a bcl-2/genética , Proteína Killer-Antagonista Homóloga a bcl-2/metabolismo , Proteína X Associada a bcl-2/genética , Proteína X Associada a bcl-2/metabolismo
15.
Nucleic Acids Res ; 47(18): 9950-9966, 2019 10 10.
Artigo em Inglês | MEDLINE | ID: mdl-31504744

RESUMO

HMGB1 is a ubiquitous non-histone protein, which biological effects depend on its expression and subcellular location. Inside the nucleus, HMGB1 is engaged in many DNA events such as DNA repair, transcription and telomere maintenance. HMGB1 has been reported to bind preferentially to bent DNA as well as to noncanonical DNA structures like 4-way junctions and, more recently, to G-quadruplexes. These are four-stranded conformations of nucleic acids involved in important cellular processes, including telomere maintenance. In this frame, G-quadruplex recognition by specific proteins represents a key event to modulate physiological or pathological pathways. Herein, to get insights into the telomeric G-quadruplex DNA recognition by HMGB1, we performed detailed biophysical studies complemented with biological analyses. The obtained results provided information about the molecular determinants for the interaction and showed that the structural variability of human telomeric G-quadruplex DNA may have significant implications in HMGB1 recognition. The biological data identified HMGB1 as a telomere-associated protein in both telomerase-positive and -negative tumor cells and showed that HMGB1 gene silencing in such cells induces telomere DNA damage foci. Altogether, these findings provide a deeper understanding of telomeric G-quadruplex recognition by HMGB1 and suggest that this protein could actually represent a new target for cancer therapy.


Assuntos
Quadruplex G , Proteína HMGB1/genética , Conformação de Ácido Nucleico , Telômero/genética , DNA/química , DNA/genética , Escherichia coli/genética , Proteína HMGB1/química , Humanos , Telomerase/química , Telomerase/genética , Telômero/química
16.
Int J Mol Sci ; 20(17)2019 Aug 30.
Artigo em Inglês | MEDLINE | ID: mdl-31480228

RESUMO

Karyotypic data from Australian native freshwater fishes are scarce, having been described from relatively few species. Golden perch (Macquaria ambigua) and Murray cod (Maccullochella peelii) are two large-bodied freshwater fish species native to Australia with significant indigenous, cultural, recreational and commercial value. The arid landscape over much of these fishes' range, coupled with the boom and bust hydrology of their habitat, means that these species have potential to provide useful evolutionary insights, such as karyotypes and sex chromosome evolution in vertebrates. Here we applied standard and molecular cytogenetic techniques to characterise karyotypes for golden perch and Murray cod. Both species have a diploid chromosome number 2n = 48 and a male heterogametic sex chromosome system (XX/XY). While the karyotype of golden perch is composed exclusively of acrocentric chromosomes, the karyotype of Murray cod consists of two submetacentric and 46 subtelocentric/acrocentric chromosomes. We have identified variable accumulation of repetitive sequences (AAT)10 and (CGG)10 along with diverse methylation patterns, especially on the sex chromosomes in both species. Our study provides a baseline for future cytogenetic analyses of other Australian freshwater fishes, especially species from the family Percichthyidae, to better understand their genome and sex chromosome evolution.


Assuntos
Água Doce , Cariótipo , Percas/genética , Perciformes/genética , Cromossomos Sexuais/genética , Animais , Bandeamento Cromossômico , Metilação de DNA/genética , Feminino , Geografia , Masculino , Metáfase , Repetições de Microssatélites/genética , Filogenia , Especificidade da Espécie , Telômero/genética
17.
Parasitol Res ; 118(10): 2789-2800, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31485863

RESUMO

An original cytogenetic study combining classical karyotype analysis and modern fluorescence in situ hybridization using telomeric (TTAGGG)n and ribosomal sequences (18S rDNA) was performed in Khawia abbottinae (Cestoda, Caryophyllidea), a parasite of Chinese false gudgeon (Abbottina rivularis) from China. Analyses based on conventional Giemsa staining, DAPI, YOYO-1 dye, and silver (Ag) staining were also carried out. The karyotype is composed of eight pairs of metacentric and telocentric chromosomes (2n = 16, n=5m + 3t). Constitutive heterochromatin was mainly positioned at pericentromeric regions, and telomeric sequences (TTAGGG)n were restricted to the end of all chromosomes. In mitotic preparations stained with Giemsa, both homologues of chromosome pair 4 showed a distinct secondary constriction. FISH with rDNA probe confirmed that this secondary constriction contains a nucleolar organizer region (NOR). The process of spermatocyte meiosis and the dynamics of nucleolus degradation in dividing cell were scrutinized. The present study and its results enhance the limited knowledge on basic karyotype characteristics and 18S rDNA clusters location in caryophyllidean tapeworms.


Assuntos
Cestoides/genética , Cromossomos/genética , DNA Ribossômico/genética , Telômero/genética , Animais , Cestoides/classificação , Cestoides/isolamento & purificação , China , Cyprinidae/genética , DNA de Helmintos/genética , Proteínas de Helminto/genética , Heterocromatina/genética , Hibridização in Situ Fluorescente , Cariótipo , Cariotipagem , Ribossomos/genética
18.
Nat Commun ; 10(1): 4067, 2019 09 06.
Artigo em Inglês | MEDLINE | ID: mdl-31492885

RESUMO

ARID1A inactivation causes mitotic defects. Paradoxically, cancers with high ARID1A mutation rates typically lack copy number alterations (CNAs). Here, we show that ARID1A inactivation causes defects in telomere cohesion, which selectively eliminates gross chromosome aberrations during mitosis. ARID1A promotes the expression of cohesin subunit STAG1 that is specifically required for telomere cohesion. ARID1A inactivation causes telomere damage that can be rescued by STAG1 expression. Colony formation capability of single cells in G2/M, but not G1 phase, is significantly reduced by ARID1A inactivation. This correlates with an increase in apoptosis and a reduction in tumor growth. Compared with ARID1A wild-type tumors, ARID1A-mutated tumors display significantly less CNAs across multiple cancer types. Together, these results show that ARID1A inactivation is selective against gross chromosome aberrations through causing defects in telomere cohesion, which reconciles the long-standing paradox between the role of ARID1A in maintaining mitotic integrity and the lack of genomic instability in ARID1A-mutated cancers.


Assuntos
Instabilidade Genômica , Mutação , Proteínas Nucleares/genética , Neoplasias Ovarianas/genética , Telômero/genética , Fatores de Transcrição/genética , Animais , Apoptose/genética , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular , Linhagem Celular Tumoral , Proteínas Cromossômicas não Histona/genética , Proteínas Cromossômicas não Histona/metabolismo , Variações do Número de Cópias de DNA , Feminino , Humanos , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos SCID , Camundongos Transgênicos , Proteínas Nucleares/metabolismo , Neoplasias Ovarianas/metabolismo , Neoplasias Ovarianas/patologia , Telômero/metabolismo , Fatores de Transcrição/metabolismo , Transplante Heterólogo/métodos , Carga Tumoral/genética
19.
Genetics ; 213(2): 411-429, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31391264

RESUMO

Telomeres progressively shorten at every round of DNA replication in the absence of telomerase. When they become critically short, telomeres trigger replicative senescence by activating a DNA damage response that is governed by the Mec1/ATR and Tel1/ATM protein kinases. While Mec1/ATR is known to block cell division when extended single-stranded DNA (ssDNA) accumulates at eroded telomeres, the molecular mechanism by which Tel1/ATM promotes senescence is still unclear. By characterizing a Tel1-hy184 mutant variant that compensates for the lack of Mec1 functions, we provide evidence that Tel1 promotes senescence by signaling to a Rad9-dependent checkpoint. Tel1-hy184 anticipates senescence onset in telomerase-negative cells, while the lack of Tel1 or the expression of a kinase-defective (kd) Tel1 variant delays it. Both Tel1-hy184 and Tel1-kd do not alter ssDNA generation at telomeric DNA ends. Furthermore, Rad9 and (only partially) Mec1 are responsible for the precocious senescence promoted by Tel1-hy184. This precocious senescence is mainly caused by the F1751I, D1985N, and E2133K amino acid substitutions, which are located in the FRAP-ATM-TRAPP domain of Tel1 and also increase Tel1 binding to DNA ends. Altogether, these results indicate that Tel1 induces replicative senescence by directly signaling dysfunctional telomeres to the checkpoint machinery.


Assuntos
Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteínas Serina-Treonina Quinases/genética , Proteínas de Saccharomyces cerevisiae/genética , Encurtamento do Telômero/genética , Telômero/genética , Substituição de Aminoácidos/genética , Proteínas Mutadas de Ataxia Telangiectasia/genética , Pontos de Checagem do Ciclo Celular/genética , Divisão Celular/genética , Senescência Celular/genética , Dano ao DNA/genética , Replicação do DNA/genética , DNA de Cadeia Simples/genética , Proteínas de Ligação a DNA/genética , Proteínas Mutantes/genética , Saccharomyces cerevisiae/genética , Telomerase/genética
20.
Nucleic Acids Res ; 47(18): 9842-9856, 2019 10 10.
Artigo em Inglês | MEDLINE | ID: mdl-31392988

RESUMO

To elucidate the molecular nature of evolutionary changes of telomeres in the plant order Asparagales, we aimed to characterize telomerase RNA subunits (TRs) in these plants. The unusually long telomere repeat unit in Allium plants (12 nt) allowed us to identify TRs in transcriptomic data of representative species of the Allium genus. Orthologous TRs were then identified in Asparagales plants harbouring telomere DNA composed of TTAGGG (human type) or TTTAGGG (Arabidopsis-type) repeats. Further, we identified TRs across the land plant phylogeny, including common model plants, crop plants, and plants with unusual telomeres. Several lines of functional testing demonstrate the templating telomerase function of the identified TRs and disprove a functionality of the only previously reported plant telomerase RNA in Arabidopsis thaliana. Importantly, our results change the existing paradigm in plant telomere biology which has been based on the existence of a relatively conserved telomerase reverse transcriptase subunit (TERT) associating with highly divergent TRs even between closely related plant taxa. The finding of a monophyletic origin of genuine TRs across land plants opens the possibility to identify TRs directly in transcriptomic or genomic data and/or predict telomere sequences synthesized according to the respective TR template region.


Assuntos
Evolução Molecular , Filogenia , RNA/genética , Telomerase/genética , Telômero/genética , Allium/genética , Arabidopsis/genética , Asparagales/genética , Embriófitas/genética , Genoma de Planta/genética , Humanos
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