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2.
Mol Cell Biol ; 40(22)2020 10 26.
Artigo em Inglês | MEDLINE | ID: mdl-32900821

RESUMO

The rRNA gene, which consists of tandem repetitive arrays (ribosomal DNA [rDNA] repeat), is one of the most unstable regions in the genome. The rDNA repeat in the budding yeast Saccharomyces cerevisiae is known to become unstable as the cell ages. However, it is unclear how the rDNA repeat changes in aging mammalian cells. Using quantitative single-cell analyses, we identified age-dependent alterations in rDNA copy number and levels of methylation in mice. The degree of methylation and copy number of rDNA from bone marrow cells of 2-year-old mice were increased by comparison to levels in 4-week-old mice in two mouse strains, BALB/cA and C57BL/6. Moreover, the level of pre-rRNA transcripts was reduced in older BALB/cA mice. We also identified many sequence variations in the rDNA. Among them, three mutations were unique to old mice, and two of them were found in the conserved region in budding yeast. We established yeast strains with the old-mouse-specific mutations and found that they shortened the life span of the cells. Our findings suggest that rDNA is also fragile in mammalian cells and that alterations within this region have a profound effect on cellular function.


Assuntos
Envelhecimento/genética , Variações do Número de Cópias de DNA , Metilação de DNA , DNA Ribossômico/metabolismo , Envelhecimento/metabolismo , Animais , Células da Medula Óssea/metabolismo , Senescência Celular/genética , Sequência Conservada , Análise Mutacional de DNA , Sequência de DNA Instável , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Saccharomycetales/genética , Análise de Célula Única , Especificidade da Espécie , Transcrição Genética
3.
Life Sci ; 259: 118199, 2020 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-32781064

RESUMO

Cellular senescence, a process whereby cells enter a state of permanent growth arrest, appears to regulate cardiac pathological remodeling and dysfunction in response to various stresses including myocardial infarction (MI). However, the precise role as well as the underlying regulatory mechanism of cardiac cellular senescence in the ischemic heart disease remain to be further determined. Herein we report an inhibitory role of Nrf2, a key transcription factor of cellular defense, in regulating cardiac senescence in infarcted hearts as well as a therapeutic potential of targeting Nrf2-mediated suppression of cardiac senescence in the treatment of MI-induced cardiac dysfunction. MI was induced by left coronary artery ligation for 28 days in mice. Heart tissues from the infarct border zone were used for the analyses. The MI-induced cardiac dysfunction was associated with increased myocardial cell senescence, oxidative stress and apoptosis in adult wild type (WT) mice. In addition, a downregulated Nrf2 activity was associated with upregulated Keap1 levels and increased phosphorylation of JAK and FYN in the infarcted border zone heart tissues. Nrf2 Knockout (Nrf2-/-) enhanced the MI-induced myocardial, cardiac dysfunction and senescence. Qiliqiangxin (QLQX), a herbal medicine which could reverse the MI-induced suppression of Nrf2 activity, significantly inhibited the MI-induced cardiac senescence, apoptosis, and cardiac dysfunction in WT mice but not in Nrf2-/- mice. These results indicate that MI downregulates Nrf2 activity thus promoting oxidative stress to accelerate cellular senescence in the infarcted heart towards cardiac dysfunction and Nrf2 may be a drug target for suppressing the cellular senescence-associated pathologies in infarcted hearts.


Assuntos
Cardiomiopatias/genética , Cardiomiopatias/patologia , Senescência Celular/genética , Infarto do Miocárdio/genética , Infarto do Miocárdio/patologia , Miocárdio/patologia , Fator 2 Relacionado a NF-E2/genética , Animais , Cardiomiopatias/diagnóstico por imagem , Ecocardiografia , Inativação Gênica , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Infarto do Miocárdio/diagnóstico por imagem , Miócitos Cardíacos/metabolismo , RNA Interferente Pequeno/farmacologia , Remodelação Ventricular/fisiologia
4.
Nat Commun ; 11(1): 3321, 2020 07 03.
Artigo em Inglês | MEDLINE | ID: mdl-32620872

RESUMO

Human telomeres are bound by the telomere repeat binding proteins TRF1 and TRF2. Telomere shortening in human cells leads to a DNA damage response that signals replicative senescence. While insufficient loading of TRF2 at shortened telomeres contributes to the DNA damage response in senescence, the contribution of TRF1 to senescence induction has not been determined. Here we show that counter to TRF2 deficiency-mediated induction of DNA damage, TRF1 deficiency serves a protective role to limit induction of DNA damage induced by subtelomere recombination. Shortened telomeres recruit insufficient TRF1 and as a consequence inadequate tankyrase 1 to resolve sister telomere cohesion. Our findings suggest that the persistent cohesion protects short telomeres from inappropriate recombination. Ultimately, in the final division, telomeres are no longer able to maintain cohesion and subtelomere copying ensues. Thus, the gradual loss of TRF1 and concomitant persistent cohesion that occurs with telomere shortening ensures a measured approach to replicative senescence.


Assuntos
Encurtamento do Telômero/genética , Telômero/genética , Proteína 1 de Ligação a Repetições Teloméricas/genética , Proteína 2 de Ligação a Repetições Teloméricas/genética , Sequência de Bases , Linhagem Celular , Linhagem Celular Tumoral , Senescência Celular/genética , Dano ao DNA , Células HEK293 , Heterocromatina/genética , Heterocromatina/metabolismo , Humanos , Hibridização in Situ Fluorescente , Mutação , Interferência de RNA , Tanquirases/metabolismo , Telômero/metabolismo , Proteína 1 de Ligação a Repetições Teloméricas/deficiência , Proteína 1 de Ligação a Repetições Teloméricas/metabolismo , Proteína 2 de Ligação a Repetições Teloméricas/deficiência , Proteína 2 de Ligação a Repetições Teloméricas/metabolismo
5.
Am J Physiol Endocrinol Metab ; 319(2): E447-E454, 2020 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-32691630

RESUMO

The aim of the present study was to determine if the training status decreases inflammation, slows down senescence, and preserves telomere health in skeletal muscle in older compared with younger subjects, with a specific focus on satellite cells. Analyses were conducted on skeletal muscle and cultured satellite cells from vastus lateralis biopsies (n = 34) of male volunteers divided into four groups: young sedentary (YS), young trained cyclists (YT), old sedentary (OS), and old trained cyclists (OT). The senescence state and inflammatory profile were evaluated by telomere dysfunction-induced foci (TIF) quantification, senescence-associated ß-galactosidase (SA-ß-Gal) staining, and quantitative (q)RT-PCR. Independently of the endurance training status, TIF levels (+35%, P < 0.001) and the percentage of SA-ß-Gal-positive cells (+30%, P < 0.05) were higher in cultured satellite cells of older compared with younger subjects. p16 (4- to 5-fold) and p21 (2-fold) mRNA levels in skeletal muscle were higher with age but unchanged by the training status. Aging induced higher CD68 mRNA levels in human skeletal muscle (+102%, P = 0.009). Independently of age, both trained groups had lower IL-8 mRNA levels (-70%, P = 0.011) and tended to have lower TNF-α mRNA levels (-40%, P = 0.10) compared with the sedentary subjects. All together, we found that the endurance training status did not slow down senescence in skeletal muscle and satellite cells in older compared with younger subjects despite reduced inflammation in skeletal muscle. These findings highlight that the link between senescence and inflammation can be disrupted in skeletal muscle.


Assuntos
Envelhecimento/fisiologia , Treino Aeróbico , Inflamação/prevenção & controle , Músculo Esquelético/fisiologia , Resistência Física/fisiologia , Homeostase do Telômero/fisiologia , Idoso , Senescência Celular/genética , Senescência Celular/fisiologia , Inibidor p16 de Quinase Dependente de Ciclina/genética , Inibidor de Quinase Dependente de Ciclina p21/genética , Humanos , Masculino , Músculo Esquelético/química , Músculo Esquelético/citologia , RNA Mensageiro/análise , Células Satélites de Músculo Esquelético/fisiologia , Células Satélites de Músculo Esquelético/ultraestrutura , Telômero/fisiologia , Telômero/ultraestrutura , Adulto Jovem , beta-Galactosidase/análise
6.
Nat Cell Biol ; 22(8): 960-972, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32719551

RESUMO

It remains unknown if biophysical or material properties of biomolecular condensates regulate cancer. Here we show that AKAP95, a nuclear protein that regulates transcription and RNA splicing, plays an important role in tumorigenesis by supporting cancer cell growth and suppressing oncogene-induced senescence. AKAP95 forms phase-separated and liquid-like condensates in vitro and in nucleus. Mutations of key residues to different amino acids perturb AKAP95 condensation in opposite directions. Importantly, the activity of AKAP95 in splice regulation is abolished by disruption of condensation, significantly impaired by hardening of condensates, and regained by substituting its condensation-mediating region with other condensation-mediating regions from irrelevant proteins. Moreover, the abilities of AKAP95 in regulating gene expression and supporting tumorigenesis require AKAP95 to form condensates with proper liquidity and dynamicity. These results link phase separation to tumorigenesis and uncover an important role of appropriate biophysical properties of protein condensates in gene regulation and cancer.


Assuntos
Proteínas de Ancoragem à Quinase A/fisiologia , Carcinogênese/genética , Transformação Celular Neoplásica/genética , Proteínas Nucleares/fisiologia , Processamento de RNA , Proteínas de Ancoragem à Quinase A/química , Animais , Carcinogênese/metabolismo , Transformação Celular Neoplásica/metabolismo , Células Cultivadas , Senescência Celular/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Camundongos , Proteínas Nucleares/química , Transição de Fase , Processamento de RNA/fisiologia , Relação Estrutura-Atividade
7.
Nature ; 583(7817): 590-595, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32669714

RESUMO

Ageing is characterized by a progressive loss of physiological integrity, leading to impaired function and increased vulnerability to death1. Despite rapid advances over recent years, many of the molecular and cellular processes that underlie the progressive loss of healthy physiology are poorly understood2. To gain a better insight into these processes, here we generate a single-cell transcriptomic atlas across the lifespan of Mus musculus that includes data from 23 tissues and organs. We found cell-specific changes occurring across multiple cell types and organs, as well as age-related changes in the cellular composition of different organs. Using single-cell transcriptomic data, we assessed cell-type-specific manifestations of different hallmarks of ageing-such as senescence3, genomic instability4 and changes in the immune system2. This transcriptomic atlas-which we denote Tabula Muris Senis, or 'Mouse Ageing Cell Atlas'-provides molecular information about how the most important hallmarks of ageing are reflected in a broad range of tissues and cell types.


Assuntos
Envelhecimento/genética , Senescência Celular/genética , Especificidade de Órgãos/genética , Análise de Célula Única , Transcriptoma/genética , Animais , Análise Mutacional de DNA , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Instabilidade Genômica , Imunidade/genética , Imunidade/imunologia , Fígado/citologia , Fígado/metabolismo , Masculino , Camundongos , Modelos Animais , Linfócitos T/citologia , Linfócitos T/metabolismo
8.
Nucleic Acids Res ; 48(14): 7883-7898, 2020 08 20.
Artigo em Inglês | MEDLINE | ID: mdl-32609810

RESUMO

Circular DNA can arise from all parts of eukaryotic chromosomes. In yeast, circular ribosomal DNA (rDNA) accumulates dramatically as cells age, however little is known about the accumulation of other chromosome-derived circles or the contribution of such circles to genetic variation in aged cells. We profiled circular DNA in Saccharomyces cerevisiae populations sampled when young and after extensive aging. Young cells possessed highly diverse circular DNA populations but 94% of the circular DNA were lost after ∼15 divisions, whereas rDNA circles underwent massive accumulation to >95% of circular DNA. Circles present in both young and old cells were characterized by replication origins including circles from unique regions of the genome and repetitive regions: rDNA and telomeric Y' regions. We further observed that circles can have flexible inheritance patterns: [HXT6/7circle] normally segregates to mother cells but in low glucose is present in up to 50% of cells, the majority of which must have inherited this circle from their mother. Interestingly, [HXT6/7circle] cells are eventually replaced by cells carrying stable chromosomal HXT6 HXT6/7 HXT7 amplifications, suggesting circular DNAs are intermediates in chromosomal amplifications. In conclusion, the heterogeneity of circular DNA offers flexibility in adaptation, but this heterogeneity is remarkably diminished with age.


Assuntos
Senescência Celular/genética , Replicação do DNA , DNA Circular/química , Saccharomyces cerevisiae/genética , DNA Circular/análise , Variação Genética , Padrões de Herança , Proteínas de Transporte de Monossacarídeos/genética , Sequências Repetitivas de Ácido Nucleico , Origem de Replicação , Proteínas de Saccharomyces cerevisiae/genética
9.
Nucleic Acids Res ; 48(11): 6001-6018, 2020 06 19.
Artigo em Inglês | MEDLINE | ID: mdl-32427330

RESUMO

Zinc finger protein with KRAB and SCAN domains 3 (ZKSCAN3) has long been known as a master transcriptional repressor of autophagy. Here, we identify a novel role for ZKSCAN3 in alleviating senescence that is independent of its autophagy-related activity. Downregulation of ZKSCAN3 is observed in aged human mesenchymal stem cells (hMSCs) and depletion of ZKSCAN3 accelerates senescence of these cells. Mechanistically, ZKSCAN3 maintains heterochromatin stability via interaction with heterochromatin-associated proteins and nuclear lamina proteins. Further study shows that ZKSCAN3 deficiency results in the detachment of genomic lamina-associated domains (LADs) from the nuclear lamina, loss of heterochromatin, a more accessible chromatin status and consequently, aberrant transcription of repetitive sequences. Overexpression of ZKSCAN3 not only rescues premature senescence phenotypes in ZKSCAN3-deficient hMSCs but also rejuvenates physiologically and pathologically senescent hMSCs. Together, these data reveal for the first time that ZKSCAN3 functions as an epigenetic modulator to maintain heterochromatin organization and thereby attenuate cellular senescence. Our findings establish a new functional link among ZKSCAN3, epigenetic regulation, and stem cell aging.


Assuntos
Senescência Celular , Epigênese Genética , Heterocromatina/metabolismo , Fatores de Transcrição/metabolismo , Animais , Senescência Celular/genética , Regulação para Baixo , Heterocromatina/genética , Humanos , Masculino , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Mesenquimais/patologia , Camundongos , Fatores de Transcrição/deficiência
10.
Artigo em Inglês | MEDLINE | ID: mdl-32233952

RESUMO

The role of cellular senescence induced by radiation in bone loss has attracted much attention. As one of the common complications of anticancer radiotherapy, irradiation-induced bone deterioration is common and clinically significant, but the pathological mechanism has not been elucidated. This study was performed to explore the cellular senescence and senescence-associated secretory phenotype (SASP) induction of bone marrow-derived mesenchymal stem cells (BMSCs) by irradiation and its role in osteogenic differentiation dysfunction. It was observed that irradiated BMSCs lost typical fibroblast-like morphology, exhibited suppressed viability and differentiation potential accompanied with senescence phenotypes, including an increase in senescence-associated ß-galactosidase (SA-ß-gal) staining-positive cells, and upregulated senescence-related genes p53/p21, whereas no changes happened to p16. Additionally, DNA damage γ-H2AX foci, G0/G1 phase of cell cycle arrest, and cellular and mitochondrial reactive oxygen species (ROS) increased in an irradiation dose-dependent manner. Meanwhile, the JAK1/STAT3 pathway was activated and accompanied by an increase in SASP secretion, such as IL-6, IL-8, and matrix metalloproteinase-9 (MMP9), whereas 0.8 µM JAK1 inhibitor (JAKi) treatment effectively inhibited the JAK pathway and SASP production. Furthermore, conditioned medium (CM) from irradiation-induced senescent (IRIS) BMSCs exhibited a markedly reduced ability in osteogenic differentiation and marker gene expression of osteoblasts, whereas CM with JAKi intervention may effectively improve these deterioration effects. In conclusion, irradiation could provoke BMSC senescence and SASP secretion and further aggravate osteogenic differentiation dysfunction via paracrine signaling, whereas SASP targeting may be a possible intervention strategy for alleviating irradiation-induced bone loss.


Assuntos
Diferenciação Celular/genética , Senescência Celular/genética , Células-Tronco Mesenquimais/citologia , Osteogênese/genética , Reabsorção Óssea/genética , Reabsorção Óssea/terapia , Pontos de Checagem do Ciclo Celular/genética , Proliferação de Células/genética , Senescência Celular/efeitos da radiação , Dano ao DNA/efeitos da radiação , Regulação da Expressão Gênica no Desenvolvimento/efeitos da radiação , Histonas/genética , Humanos , Janus Quinase 1/genética , Células-Tronco Mesenquimais/efeitos da radiação , Mitocôndrias/genética , Mitocôndrias/efeitos da radiação , Comunicação Parácrina/genética , Radiação , Espécies Reativas de Oxigênio/metabolismo , Fator de Transcrição STAT3/genética , Transdução de Sinais/efeitos da radiação
11.
Exp Mol Pathol ; 114: 104433, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32243892

RESUMO

Combination antiretroviral therapy (cART) has greatly improved the prognosis of patients with human immunodeficiency virus type-1 (HIV-1) infection. However, cardiovascular disease (CVD) remains a serious issue even in the post-cART era. Viral protein R (Vpr), an accessory gene product of HIV-1, exerts pleiotropic activities such as the induction of DNA damage signals, apoptosis by mitochondrial membrane depolarization, G2/M-phase cell cycle abnormalities, and retrotransposition. Importantly, some of these cellular responses are induced by the trans-acting activity of Vpr. Recently, we established an enzyme-linked immunosorbent assay to detect Vpr and reported that about 22% of blood samples from 100 HIV-1-positive patients were positive for Vpr. Here, we investigated the biological effects of recombinant Vpr (rVpr) in vivo. We observed that repeated injections of rVpr increased the copy number of long interspersed element-1 (L1) in the heart genome in mice. rVpr also increased the number of cells positive for senescence-associated ß-galactosidase (SA-ß-gal) and fibrosis in the heart. Notably, co-administration of a reverse transcriptase inhibitor reduced the number of rVpr-induced SA-ß-gal-positive cells and fibrosis concomitantly with the attenuation of L1 retrotransposition. Interestingly, a Vpr mutant defective for mitochondrial dysfunction also induced heart senescence and increased L1 copy number. Together with a recent report that L1 retrotransposition functions as a molecular basis of senescence, our current data suggest that rVpr-induced L1 retrotransposition is linked with senescence in heart tissue. We would propose that Vpr in the bloodstream may be one of risk factors for CVD, and that its monitoring will lead to well understanding of the heterogeneity and multifactorial mechanisms of CVD in HIV-1 patients.


Assuntos
Senescência Celular/genética , Infecções por HIV/genética , Retroelementos/genética , Produtos do Gene vpr do Vírus da Imunodeficiência Humana/genética , Animais , Infecções por HIV/virologia , HIV-1/patogenicidade , Coração/fisiopatologia , Coração/virologia , Humanos , Camundongos
12.
EMBO Rep ; 21(5): e50322, 2020 05 06.
Artigo em Inglês | MEDLINE | ID: mdl-32346980

RESUMO

The natural progressive dysfunction of most living organisms-ageing-has captured the attention of several studies, with the intention to develop rejuvenation strategies. Evidence is emerging of a positive correlation between natural ageing and chromosomal instability (CIN). In this issue of EMBO Reports, Barroso-Vilares et al [1] now show a link between ageing and the erroneous assembly of the apparatus required for a proper cellular division. They compare this mechanism in young and naturally aged human cells and describe a strategy to delay age-related CIN.


Assuntos
Segregação de Cromossomos , Neoplasias , Envelhecimento/genética , Senescência Celular/genética , Instabilidade Cromossômica , Humanos
15.
Tissue Cell ; 63: 101323, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32223958

RESUMO

Previous reports showed that fibronectin (FN) was effective in stimulating the recovery of damaged dermis. However, native FN has multifunctional domains transmitting beneficial as well as unbeneficial signals to dermal tissue cells through the mediation of integrin heterodimers. The use of a functional domain [FN type III9-10 fragments (FNIII9-10)] providing beneficial effects on the physiology of dermal tissue cells would enhance an in vitro culture system for dermal fibroblasts (DFs). We therefore investigated the FNIII9-10-derived extracellular signaling effect on the physiology of DFs during in vitro culture. Recombinant FNIII9-10 proteins were constructed and their functionality was determined by observing the adhesion of adult human DFs (aHDFs) to recombinant FNIII9-10 and of low adhesion integrin α5ß1- and αvß3-blocked aHDFs to recombinant FNIII9-10. Cellular proliferation, morphology, and senescence were measured and compared in the aHDFs cultured on native FN and recombinant FNIII9-10 for short or long periods. The results show that recombinant FNIII9-10-derived extracellular signaling stimulated increased proliferation of aHDF (both in short- and long-term cultures) and inhibited the generation of morphological abnormalities (in short- and long-term cultures) and cellular senescence (long-term culture) when compared with native FN-derived extracellular signaling. Our results suggest that, instead of native FN, recombinant FNIII9-10 better enhanced the in vitro culture of aHDFs while diminishing the adverse effects associated with the use of human-derived materials.


Assuntos
Senescência Celular/genética , Derme/efeitos dos fármacos , Fibroblastos/efeitos dos fármacos , Fibronectinas/genética , Adesão Celular/efeitos dos fármacos , Adesão Celular/genética , Proliferação de Células/genética , Células Cultivadas , Derme/metabolismo , Fibroblastos/metabolismo , Domínio de Fibronectina Tipo III/genética , Fibronectinas/farmacologia , Humanos , Integrina alfa5beta1/genética , Transdução de Sinais/genética
16.
Gene ; 743: 144582, 2020 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-32173543

RESUMO

As the main energy source for generating ATP during plant growth and development, sugars are synthesized in leaves, while sugar allocation depends on both intracellular transport between different organelles and source-to-sink transport. However, sugar transport related research is limited in pear. Here, a sugar transporter PbSWEET4 was identified that control sugar content and senescence in leaf. Phylogenetic analysis and multiple sequence alignment results indicated that PbSWEET4 was homologous to AtSWEET15, which contained two conserved domains and could promote senescence. The qRT-PCR and transcriptome database result showed that the expression of PbSWEET4 was positively correlated with leaf development, especially highly expressed in older leaves. Furthermore, the evaluation of promoter-GUS activity also indicated that PbSWEET4 exhibited the highest expression level in older leaves. The subcellular localization revealed that the PbSWEET4 localized in the plasma membrane. Finally, overexpression of the PbSWEET4 in strawberry plants could reduce leaf sugar content and chlorophyll content, while accelerate leaf senescence, which might be due to enhanced export of sugars from leaves. These results enrich the knowledge about the function of sugar exporter in regulating the fruit species development, and provide a novel genetic resource for future improvement in carbohydrate partitioning for pear and other fruit trees.


Assuntos
Regulação da Expressão Gênica de Plantas , Proteínas de Membrana Transportadoras/genética , Proteínas de Plantas/genética , Pyrus/fisiologia , Membrana Celular/metabolismo , Senescência Celular/genética , Clorofila/análise , Fragaria/genética , Proteínas de Membrana Transportadoras/metabolismo , Folhas de Planta/química , Folhas de Planta/genética , Folhas de Planta/crescimento & desenvolvimento , Folhas de Planta/metabolismo , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas , Pyrus/química , Açúcares/análise , Açúcares/metabolismo
17.
DNA Cell Biol ; 39(5): 747-755, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32182129

RESUMO

Polo-like kinase 1 (PLK1) is a ubiquitous serine/threonine protein kinase. It is reported to be involved in the occurrence and progression of various human cancers. In the present study, we explored the role and molecular mechanism of PLK1 in the proliferation of osteosarcoma (OS) cells. We found that PLK1 expression was higher in MG63/Dox cells than in MG63 cells, while inhibiting or interfering with the level of PLK1 suppressed cell proliferation of MG63/Dox cells. TargetScan analysis predicted that miR-4779 would interact with the 3'-UTR of PLK1 mRNAs and also inhibit cell autophagy of MG63/Dox cells. The data demonstrated that miR-4779 negatively regulates the expression of PLK1, and both miR-4779 and PLK1 regulate cell proliferation and cell apoptosis of MG63/Dox cells, processes that are involved in the drug resistance of OS cells.


Assuntos
Apoptose/genética , Proteínas de Ciclo Celular/deficiência , Proteínas de Ciclo Celular/genética , Técnicas de Silenciamento de Genes , MicroRNAs/genética , Proteínas Serina-Treonina Quinases/deficiência , Proteínas Serina-Treonina Quinases/genética , Proteínas Proto-Oncogênicas/deficiência , Proteínas Proto-Oncogênicas/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Sobrevivência Celular/genética , Senescência Celular/genética , Regulação Neoplásica da Expressão Gênica/genética , Humanos
18.
Mol Cell ; 78(3): 522-538.e9, 2020 05 07.
Artigo em Inglês | MEDLINE | ID: mdl-32220303

RESUMO

To understand the role of the extensive senescence-associated 3D genome reorganization, we generated genome-wide chromatin interaction maps, epigenome, replication-timing, whole-genome bisulfite sequencing, and gene expression profiles from cells entering replicative senescence (RS) or upon oncogene-induced senescence (OIS). We identify senescence-associated heterochromatin domains (SAHDs). Differential intra- versus inter-SAHD interactions lead to the formation of senescence-associated heterochromatin foci (SAHFs) in OIS but not in RS. This OIS-specific configuration brings active genes located in genomic regions adjacent to SAHDs in close spatial proximity and favors their expression. We also identify DNMT1 as a factor that induces SAHFs by promoting HMGA2 expression. Upon DNMT1 depletion, OIS cells transition to a 3D genome conformation akin to that of cells in replicative senescence. These data show how multi-omics and imaging can identify critical features of RS and OIS and discover determinants of acute senescence and SAHF formation.


Assuntos
Senescência Celular/genética , DNA (Citosina-5-)-Metiltransferase 1/genética , Genoma Humano , Oncogenes , Células Cultivadas , Montagem e Desmontagem da Cromatina/genética , DNA (Citosina-5-)-Metiltransferase 1/metabolismo , Metilação de DNA , Fibroblastos , Heterocromatina/genética , Humanos , Hibridização in Situ Fluorescente
19.
Clin Sci (Lond) ; 134(7): 727-746, 2020 04 17.
Artigo em Inglês | MEDLINE | ID: mdl-32202295

RESUMO

We previously identified genomic instability as a causative factor for vascular aging. In the present study, we determined which vascular aging outcomes are due to local endothelial DNA damage, which was accomplished by genetic removal of ERCC1 (excision repair cross-complementation group 1) DNA repair in mice (EC-knockout (EC-KO) mice). EC-KO showed a progressive decrease in microvascular dilation of the skin, increased microvascular leakage in the kidney, decreased lung perfusion, and increased aortic stiffness compared with wild-type (WT). EC-KO showed expression of DNA damage and potential senescence marker p21 exclusively in the endothelium, as demonstrated in aorta. Also the kidney showed p21-positive cells. Vasodilator responses measured in organ baths were decreased in aorta, iliac and coronary artery EC-KO compared with WT, of which coronary artery was the earliest to be affected. Nitric oxide-mediated endothelium-dependent vasodilation was abolished in aorta and coronary artery, whereas endothelium-derived hyperpolarization and responses to exogenous nitric oxide (NO) were intact. EC-KO showed increased superoxide production compared with WT, as measured in lung tissue, rich in endothelial cells (ECs). Arterial systolic blood pressure (BP) was increased at 3 months, but normal at 5 months, at which age cardiac output (CO) was decreased. Since no further signs of cardiac dysfunction were detected, this decrease might be an adaptation to prevent an increase in BP. In summary, a selective DNA repair defect in the endothelium produces features of age-related endothelial dysfunction, largely attributed to loss of endothelium-derived NO. Increased superoxide generation might contribute to the observed changes affecting end organ perfusion, as demonstrated in kidney and lung.


Assuntos
Envelhecimento/genética , Senescência Celular/genética , Dano ao DNA , Reparo do DNA , Proteínas de Ligação a DNA/deficiência , Endonucleases/deficiência , Células Endoteliais/metabolismo , Endotélio Vascular/metabolismo , Fatores Etários , Envelhecimento/metabolismo , Envelhecimento/patologia , Animais , Permeabilidade Capilar , Inibidor de Quinase Dependente de Ciclina p21/genética , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Proteínas de Ligação a DNA/genética , Endonucleases/genética , Células Endoteliais/patologia , Endotélio Vascular/patologia , Endotélio Vascular/fisiopatologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Superóxidos/metabolismo , Rigidez Vascular , Vasodilatação
20.
Oxid Med Cell Longev ; 2020: 9369524, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32190179

RESUMO

Aging has been characterized with the accumulation of oxidized proteins, as a consequence of progressive decline in proteostasis capacity. Among others, proteasomal system is an efficient protein turnover complex to avoid aggregation of oxidized proteins. Heat shock protein 70 (HSP70) is another critical player that is involved in some key processes including the correct folding of misfolded proteins and targeting aggregated proteins to the proteasome for rapid degradation. The aim of this study was to determine the role of proteasomal system and heat shock proteins to maintain proteome balance during replicative senescence in mild hyperthermia conditions. Our results demonstrated that HSP40/70 machinery is induced by mild hyperthermia conditions independent from senescence conditions. Since HSP70 is largely responsible for the rapidly inducible cell protection following hyperthermia, the activation of "heat shock response" resulted in the elevation of HSP40/70 expressions as well as the proteasome activity. Interestingly, when HSP70 expression was inhibited, increased proteasomal activation was shown to be responsive to mild hyperthermia. Since HSP70 is involved in various stress-related pathways such as oxidative and endoplasmic reticulum stress, depletion of HSP70 expression may induce proteasomal degradation to maintain proteome balance of the cell. Thus, our data suggests that in mild heat stress conditions, molecular chaperone HSP70 plays an important role to avoid protein oxidation and aggregation; however, activities of proteasomal system are induced when HSP70 expression is depleted.


Assuntos
Senescência Celular , Fibroblastos/citologia , Fibroblastos/metabolismo , Proteínas de Choque Térmico HSP70/antagonistas & inibidores , Hipertermia Induzida , Complexo de Endopeptidases do Proteassoma/metabolismo , Compostos Benzidrílicos/farmacologia , Senescência Celular/genética , Inativação Gênica , Proteínas de Choque Térmico HSP40/genética , Proteínas de Choque Térmico HSP40/metabolismo , Proteínas de Choque Térmico HSP70/genética , Proteínas de Choque Térmico HSP70/metabolismo , Resposta ao Choque Térmico/genética , Humanos , Masculino , Proteostase , Pirrolidinonas/farmacologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
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