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1.
Nat Commun ; 12(1): 3318, 2021 06 03.
Artigo em Inglês | MEDLINE | ID: mdl-34083536

RESUMO

Dormancy, a reversible quiescent cellular state characterized by greatly reduced metabolic activity, protects from genetic damage, prolongs survival and is crucial for tissue homeostasis and cellular response to injury or transplantation. Dormant cells have been characterized in many tissues, but their identification, isolation and characterization irrespective of tissue of origin remains elusive. Here, we develop a live cell ratiometric fluorescent Optical Stem Cell Activity Reporter (OSCAR) based on the observation that phosphorylation of RNA Polymerase II (RNApII), a hallmark of active mRNA transcription elongation, is largely absent in dormant stem cells from multiple lineages. Using the small intestinal crypt as a model, OSCAR reveals in real time the dynamics of dormancy induction and cellular differentiation in vitro, and allows the identification and isolation of several populations of transcriptionally diverse OSCARhigh and OSCARlow intestinal epithelial cell states in vivo. In particular, this reporter is able to identify a dormant OSCARhigh cell population in the small intestine. OSCAR therefore provides a tool for a better understanding of dormant stem cell biology.


Assuntos
RNA Polimerase II/metabolismo , Fase de Repouso do Ciclo Celular/fisiologia , Animais , Separação Celular , Quinase 9 Dependente de Ciclina/metabolismo , Citometria de Fluxo , Corantes Fluorescentes/metabolismo , Humanos , Mucosa Intestinal/citologia , Mucosa Intestinal/metabolismo , Intestino Delgado/citologia , Intestino Delgado/metabolismo , Proteínas Luminescentes/metabolismo , Camundongos , Camundongos Transgênicos , RNA Mensageiro/metabolismo , Transcrição Gênica
2.
Yeast ; 38(1): 30-38, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33350501

RESUMO

A subset of Saccharomyces cerevisiae cells in a stationary phase culture achieve a unique quiescent state characterized by increased cell density, stress tolerance, and longevity. Trehalose accumulation is necessary but not sufficient for conferring this state, and it is not recapitulated by abrupt starvation. The fraction of cells that achieve this state varies widely in haploids and diploids and can approach 100%, indicating that both mother and daughter cells can enter quiescence. The transition begins when about half the glucose has been taken up from the medium. The high affinity glucose transporters are turned on, glycogen storage begins, the Rim15 kinase enters the nucleus and the accumulation of cells in G1 is initiated. After the diauxic shift (DS), when glucose is exhausted from the medium, growth promoting genes are repressed by the recruitment of the histone deacetylase Rpd3 by quiescence-specific repressors. The final division that takes place post-DS is highly asymmetrical and G1 arrest is complete after 48 h. The timing of these events can vary considerably, but they are tightly correlated with total biomass of the culture, suggesting that the transition to quiescence is tightly linked to changes in external glucose levels. After 7 days in culture, there are massive morphological changes at the protein and organelle level. There are global changes in histone modification. An extensive array of condensin-dependent, long-range chromatin interactions lead to genome-wide chromatin compaction that is conserved in yeast and human cells. These interactions are required for the global transcriptional repression that occurs in quiescent yeast.


Assuntos
Fase de Repouso do Ciclo Celular , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/fisiologia , Saccharomycetales/genética , Saccharomycetales/fisiologia , Adenosina Trifosfatases/genética , Adenosina Trifosfatases/metabolismo , Divisão Celular/genética , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Regulação Fúngica da Expressão Gênica , Genoma Fúngico , Glucose/metabolismo , Código das Histonas , Complexos Multiproteicos/genética , Complexos Multiproteicos/metabolismo , Fase de Repouso do Ciclo Celular/genética , Fase de Repouso do Ciclo Celular/fisiologia , Saccharomyces cerevisiae/crescimento & desenvolvimento , Proteínas de Saccharomyces cerevisiae/genética , Transcrição Gênica
3.
Yeast ; 38(1): 12-29, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33350503

RESUMO

Cellular quiescence, the temporary and reversible exit from proliferative growth, is the predominant state of all cells. However, our understanding of the biological processes and molecular mechanisms that underlie cell quiescence remains incomplete. As with the mitotic cell cycle, budding and fission yeast are preeminent model systems for studying cellular quiescence owing to their rich experimental toolboxes and the evolutionary conservation across eukaryotes of pathways and processes that control quiescence. Here, we review current knowledge of cell quiescence in budding yeast and how it pertains to cellular quiescence in other organisms, including multicellular animals. Quiescence entails large-scale remodeling of virtually every cellular process, organelle, gene expression, and metabolic state that is executed dynamically as cells undergo the initiation, maintenance, and exit from quiescence. We review these major transitions, our current understanding of their molecular bases, and highlight unresolved questions. We summarize the primary methods employed for quiescence studies in yeast and discuss their relative merits. Understanding cell quiescence has important consequences for human disease as quiescent single-celled microbes are notoriously difficult to kill and quiescent human cells play important roles in diseases such as cancer. We argue that research on cellular quiescence will be accelerated through the adoption of common criteria, and methods, for defining cell quiescence. An integrated approach to studying cell quiescence, and a focus on the behavior of individual cells, will yield new insights into the pathways and processes that underlie cell quiescence leading to a more complete understanding of the life cycle of cells. TAKE AWAY: Quiescent cells are viable cells that have reversibly exited the cell cycle Quiescence is induced in response to a variety of nutrient starvation signals Quiescence is executed dynamically through three phases: initiation, maintenance, and exit Quiescence entails large-scale remodeling of gene expression, organelles, and metabolism Single-cell approaches are required to address heterogeneity among quiescent cells.


Assuntos
Divisão Celular/fisiologia , Fase de Repouso do Ciclo Celular/genética , Saccharomycetales/genética , Saccharomycetales/fisiologia , Divisão Celular/genética , Fase de Repouso do Ciclo Celular/fisiologia , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/fisiologia , Schizosaccharomyces , Transdução de Sinais/fisiologia
4.
J Pharm Pharmacol ; 72(10): 1383-1393, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32567066

RESUMO

OBJECTIVES: Uncontrolled cell proliferation was caused by multiple deficient pathways that inhibition of one pathway may result to activate an alternative pathway. Therefore, combination of drugs which targeted multiple pathways could be beneficial to overcome drug resistance. Ciprofloxacin (CPF) cytotoxicity was widely investigated on cancer cell lines, and results revealed hepatoma-derived Hep G2 cells are relatively resistant. So, this study aimed to increase CPF cytotoxicity by rational design of a supplement which targeted Ca2+ homoeostasis as major hub in unchecked proliferation. METHODS: Cells were treated by CPF and/or pilocarpine (PILO), and cell cycle distribution, caspases activity and regulatory proteins were evaluated. KEY FINDINGS: MTT and flow cytometry analysis confirmed administration of CPF + PILO causes more cytotoxicity. CPF-exposed cells accumulated in S phase due to DNA damages while PILO + CPF imposed G0 stage arrest through cyclin D1 and P-Akt downregulation. Caspase 8 was activated in cells treated by CPF but accompaniment of PILO with CPF led to activation of caspase 9, 8 and 3 and ROS overproduction. CONCLUSIONS: Ciprofloxacin imposed mitochondrial-independent apoptosis while PILO + CPF caused mitochondrial-dependent and independent apoptosis simultaneously. Consequently, coadministration of PILO and CPF causes intense cytotoxic effects through targeting the mitochondria, DNA gyrase enzyme and other unknown mechanisms.


Assuntos
Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Ciprofloxacina/administração & dosagem , Citotoxinas/administração & dosagem , Pilocarpina/administração & dosagem , Fase de Repouso do Ciclo Celular/efeitos dos fármacos , Antibacterianos/administração & dosagem , Pontos de Checagem do Ciclo Celular/fisiologia , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Combinação de Medicamentos , Células Hep G2 , Humanos , Agonistas Muscarínicos/administração & dosagem , Fase de Repouso do Ciclo Celular/fisiologia
5.
Artigo em Inglês | MEDLINE | ID: mdl-31385777

RESUMO

BACKGROUND: Nowadays, the potential therapeutic role of various bioflavonoids including Curcumin, Luteolin and Resveratrol has currently been well-documented in a vast range of fatal complications including synaptic failure and cancers. These bioflavonoids are widely being implemented for the treatment of various cancers as they possess anti-cancerous, anti-oxidant and anti-inflammatory properties. Moreover, they are also used as a better alternative to conventional therapies since; these are non-toxic to cells and having no or least side effects. Notably, the pertinent therapeutic role of Rutin in cervical cancer is still unsettled however, its anti-cancerous role has already been reported in other cancers including prostate and colon cancer. Rutin (Vitamin P or Rutoside) is a polyphenolics flavonoid exhibiting multi-beneficial roles against several carcinomas. OBJECTIVE: Despite the evidence for its several biological activities, the anticancer effects of Rutin on human cervical cancer (C33A) cells remain to be explored. In this study, the anticancer potential of Rutin was investigated by employing the key biomarkers such as nuclear condensation reactive oxygen species (ROS), apoptosis, and changes in mitochondrial membrane potential (MMP). RESULTS: Our findings showed that Rutin treatment reduced the cell viability, induced significant increase in ROS production and nuclear condensation in dose-dependent manner. Moreover, Rutin provoked apoptosis by inducing decrease in MMP and activation of caspase-3. Cell cycle analysis further confirmed the efficacy of Rutin by showing cell cycle arrest at G0/G1 phase. CONCLUSION: Thus, our study is envisaged to open up interests for elucidating Rutin as an anticancerous agent against cervical cancer.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Fase de Repouso do Ciclo Celular/efeitos dos fármacos , Rutina/farmacologia , Neoplasias do Colo do Útero/fisiopatologia , Alphapapillomavirus , Antineoplásicos Fitogênicos/uso terapêutico , Apoptose/fisiologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/fisiologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Relação Dose-Resposta a Droga , Feminino , Pontos de Checagem da Fase G1 do Ciclo Celular/fisiologia , Células HEK293 , Humanos , Fase de Repouso do Ciclo Celular/fisiologia , Rutina/uso terapêutico , Neoplasias do Colo do Útero/tratamento farmacológico
6.
Exp Cell Res ; 387(1): 111776, 2020 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-31838060

RESUMO

Microtubule-binding proteins provide an alternative and vital pathway to the functional diversity of microtubules. Considerable work is still required to understand the complexities of microtubule-associated cellular processes and to identify novel microtubule-binding proteins. In this study, we identify Bcl2-associated athanogene cochaperone 6 (BAG6) as a novel microtubule-binding protein and reveal that it is crucial for primary ciliogenesis. By immunofluorescence we show that BAG6 largely colocalizes with intracellular microtubules and by co-immunoprecipitation we demonstated that it can interact with α-tubulin. Additionally, both the UBL and BAG domains of BAG6 are indispensable for its interaction with α-tubulin. Moreover, the assembly of primary cilia in RPE-1 cells is significantly inhibited upon the depletion of BAG6. Notably, BAG6 inhibition leads to an abnormal G0/G1 transition during the cell cycle. In addition, BAG6 colocalizes and interactes with the centrosomal protein γ-tubulin, suggesting that BAG6 might regulate primary ciliogenesis through its action in centrosomal function. Collectively, our findings suggest that BAG6 is a novel microtubule-bindng protein crucial for primary ciliogenesis.


Assuntos
Proteínas Associadas aos Microtúbulos/metabolismo , Microtúbulos/metabolismo , Chaperonas Moleculares/metabolismo , Tubulina (Proteína)/metabolismo , Proteínas de Transporte/metabolismo , Linhagem Celular , Linhagem Celular Tumoral , Fase G1/fisiologia , Células HEK293 , Células HeLa , Humanos , Ligação Proteica/fisiologia , Fase de Repouso do Ciclo Celular/fisiologia
7.
Biomed Pharmacother ; 121: 109598, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31733572

RESUMO

Lung cancer remains the leading cause of cancer mortality because of highly malignant and metastatic potential. The current status of lung cancer treatment is limited, and more treatment options are needed. Interesting, antipsychotic drugs have been reported to show anti-cancer effects. In this present study, we investigated the anticancer potential of penfluridol (PF), an anti-schizophrenic drug, in lung cancer and its underlying mechanism in vitro and in vivo. In vitro, it could inhibit the viability of various lung cancer cells with G0/G1 phase arrest via increasing the expression level of p21/p27 and decreasing the expression levels of cyclin-CDK complex. Meanwhile, cell-cycle arrest causes DNA repair in the nucleus, which was associated with the upregulation of H2A.X and p-H2A.X. Moreover, PF could also decrease mitochondrial membrane potential and increase reactive oxygen species levels in the lung cancer cells. These results implied that PF might induce the mitochondria-mediated intrinsic apoptosis. In addition, PF inhibits the migration and invasion of lung cancer cells via downregulation of FAK-MMP signaling. In vivo, oral administration of PF at concentration of 10 mg/kg inhibited tumor growth in A549 xenograft model. Notably, PF is an approved drug and the price is exceedingly cheap, so this study demonstrates the potential of PF to treat lung cancer.


Assuntos
Antipsicóticos/uso terapêutico , Apoptose/efeitos dos fármacos , Fase G1/efeitos dos fármacos , Neoplasias Pulmonares/tratamento farmacológico , Penfluridol/uso terapêutico , Fase de Repouso do Ciclo Celular/efeitos dos fármacos , Células A549 , Animais , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Antipsicóticos/farmacologia , Apoptose/fisiologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/fisiologia , Relação Dose-Resposta a Droga , Feminino , Fase G1/fisiologia , Inibidores do Crescimento/farmacologia , Inibidores do Crescimento/uso terapêutico , Humanos , Neoplasias Pulmonares/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Invasividade Neoplásica/patologia , Penfluridol/farmacologia , Fase de Repouso do Ciclo Celular/fisiologia , Ensaios Antitumorais Modelo de Xenoenxerto/métodos
8.
Dev Biol ; 456(1): 17-24, 2019 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-31390535

RESUMO

Cell proliferation and cell death are opposing but fundamental aspects of development that must be tightly controlled to ensure proper tissue organization and organismal health. Developmental apoptosis of abdominal neuroblasts in the Drosophila ventral nerve cord is controlled by multiple upstream spatial and temporal signals, which have also been implicated in control of cell proliferation. It has therefore remained unclear whether developmental apoptosis is linked to active cell proliferation. Previous investigations into this topic have focused on the effect of cell cycle arrests on exogenous induction of apoptosis, and thus have not addressed whether potential effects of the cell cycle lie with the sensing of damage signals or the execution of apoptosis itself. In this report, we show that developmental apoptosis is not inhibited by cell cycle arrest, and that endogenous cell death occurs independently of cell cycle phase. We also find that ectopic neuroblasts rescued from cell death retain the competency to respond to quiescence cues at the end of embryogenesis. In addition, we observe multiple quiescence types in neuroblasts, and we show that cell death mutant embryos display a specific loss of presumptive G2 quiescent abdominal neuroblasts at the end of embryogenesis. This study demonstrates that upstream control of neuroblast proliferation and apoptosis represent independent mechanisms of regulating stem cell fate, and that execution of apoptosis occurs in a cell cycle-independent manner. Our findings also indicate that a subset of G2Q-fated abdominal neuroblasts are eliminated from the embryo through a non-apoptotic mechanism.


Assuntos
Apoptose/fisiologia , Ciclo Celular/fisiologia , Neurogênese/fisiologia , Animais , Pontos de Checagem do Ciclo Celular/fisiologia , Diferenciação Celular/fisiologia , Divisão Celular/fisiologia , Proliferação de Células/fisiologia , Sistema Nervoso Central/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/genética , Células-Tronco Neurais/citologia , Neurônios/metabolismo , Fenótipo , Fase de Repouso do Ciclo Celular/fisiologia , Transdução de Sinais/fisiologia
9.
Epigenetics Chromatin ; 12(1): 45, 2019 07 17.
Artigo em Inglês | MEDLINE | ID: mdl-31315658

RESUMO

BACKGROUND: Cellular quiescence is a reversible differentiation state during which cells modify their gene expression program to inhibit metabolic functions and adapt to a new cellular environment. The epigenetic changes accompanying these alterations are not well understood. We used fission yeast cells as a model to study the regulation of quiescence. When these cells are starved for nitrogen, the cell cycle is arrested in G1, and the cells enter quiescence (G0). A gene regulatory program is initiated, including downregulation of thousands of genes-for example, those related to cell proliferation-and upregulation of specific genes-for example, autophagy genes-needed to adapt to the physiological challenge. These changes in gene expression are accompanied by a marked alteration of nuclear organization and chromatin structure. RESULTS: Here, we investigated the role of Leo1, a subunit of the conserved RNA polymerase-associated factor 1 (Paf1) complex, in the quiescence process using fission yeast as the model organism. Heterochromatic regions became very dynamic in fission yeast in G0 during nitrogen starvation. The reduction of heterochromatin in early G0 was correlated with reduced target of rapamycin complex 2 (TORC2) signaling. We demonstrated that cells lacking Leo1 show reduced survival in G0. In these cells, heterochromatic regions, including subtelomeres, were stabilized, and the expression of many genes, including membrane transport genes, was abrogated. TOR inhibition mimics the effect of nitrogen starvation, leading to the expression of subtelomeric genes, and this effect was suppressed by genetic deletion of leo1. CONCLUSIONS: We identified a protein, Leo1, necessary for survival during quiescence. Leo1 is part of a conserved protein complex, Paf1C, linked to RNA polymerase II. We showed that Leo1, acting downstream of TOR, is crucial for the dynamic reorganization of chromosomes and the regulation of gene expression during cellular quiescence. Genes encoding membrane transporters are not expressed in quiescent leo1 mutant cells, and cells die after 2 weeks of nitrogen starvation. Taken together, our results suggest that Leo1 is essential for the dynamic regulation of heterochromatin and gene expression during cellular quiescence.


Assuntos
Heterocromatina/metabolismo , Proteínas de Ligação a RNA/metabolismo , Fase de Repouso do Ciclo Celular/genética , Ciclo Celular/genética , Epigênese Genética , Regulação Fúngica da Expressão Gênica , Heterocromatina/genética , Histonas/metabolismo , Proteínas Nucleares/metabolismo , RNA Polimerase II/genética , Proteínas de Ligação a RNA/genética , Fase de Repouso do Ciclo Celular/fisiologia , Schizosaccharomyces/genética , Schizosaccharomyces/metabolismo , Proteínas de Schizosaccharomyces pombe/genética , Proteínas de Schizosaccharomyces pombe/metabolismo
10.
Acta Pharm ; 69(1): 75-86, 2019 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-31259717

RESUMO

Recent studies suggest that annexin A1 (ANXA1) promotes apoptosis in cancerous cells. This study aims to investigate the effects of ANXA1 on apoptosis and cell cycle arrest in K562, Jurkat and U937 cells and peripheral blood mononu-clear cells (PBMC). Cells were treated with ANXA1 and cyclophosphamide prior to flow cytometry analysis for apoptosis and cell cycle arrest induction. At 2.5µM, ANXA1 induced significant apoptosis in K562 (p ≤ 0.001) and U937 (p ≤ 0.05) cells, with EC50 values of 3.6 and 3.8 µM, respectively. In Jurkat cells, induction was not significant (EC50, 17.0 µM). No significant apoptosis induction was observed in PBMC. ANXA1 caused cycle arrest in the G0/G1 phase in K562 and U937 cells with p ≤ 0.001 for both, and (p ≤ 0.01) for Jurkat cells. ANXA1 induced apoptosis and cycle arrest in the G0/G1 phase in K562 and U937 cells, causing only cell cycle arrest in Jurkat cells.


Assuntos
Anexina A1/metabolismo , Apoptose/fisiologia , Pontos de Checagem do Ciclo Celular/fisiologia , Leucócitos Mononucleares/metabolismo , Leucócitos Mononucleares/fisiologia , Adolescente , Adulto , Linhagem Celular , Linhagem Celular Tumoral , Feminino , Fase G1/fisiologia , Humanos , Células Jurkat , Células K562 , Masculino , Pessoa de Meia-Idade , Fase de Repouso do Ciclo Celular/fisiologia , Células U937 , Adulto Jovem
11.
Cell Death Dis ; 10(7): 502, 2019 06 26.
Artigo em Inglês | MEDLINE | ID: mdl-31243265

RESUMO

Previously, several protein-coding tumor suppressors localized at 1p36 have been reported. In the present work, we focus on functional long non-coding RNAs (lncRNAs) embedded in this locus. Small interfering RNA was used to identify lncRNA candidates with growth-suppressive activities in breast cancer. The mechanism involved was also explored. LINC01355 were downregulated in breast cancer cells relative to non-malignant breast epithelial cells. Overexpression of LINC01355 significantly inhibited proliferation, colony formation, and tumorigenesis of breast cancer cells. LINC01355 arrested breast cancer cells at the G0/G1 phase by repressing CCND1. Moreover, LINC01355 interacted with and stabilized FOXO3 protein, leading to transcriptional repression of CCND1. Importantly, LINC01355-mediated suppression of breast cancer growth was reversed by knockdown of FOXO3 or overexpression of CCND1. Clinically, LINC01355 was downregulated in breast cancer specimens and correlated with more aggressive features. There was a negative correlation between LINC01355 and CCND1 expression in breast cancer samples. LINC01355 acts as a tumor suppressor in breast cancer, which is ascribed to enhancement of FOXO3-mediated transcriptional repression of CCND1. Re-expression of LINC01355 may provide a potential therapeutic strategy to block breast cancer growth and progression.


Assuntos
Neoplasias da Mama/metabolismo , Neoplasias da Mama/terapia , Proliferação de Células/fisiologia , Ciclina D1/metabolismo , Proteína Forkhead Box O3/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/fisiologia , Animais , Western Blotting , Neoplasias da Mama/genética , Proliferação de Células/genética , Imunoprecipitação da Cromatina , Ciclina D1/genética , Feminino , Citometria de Fluxo , Proteína Forkhead Box O3/genética , Fase G1/genética , Fase G1/fisiologia , Regulação Neoplásica da Expressão Gênica/genética , Regulação Neoplásica da Expressão Gênica/fisiologia , Humanos , Técnicas In Vitro , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Plasmídeos/genética , Reação em Cadeia da Polimerase em Tempo Real , Fase de Repouso do Ciclo Celular/genética , Fase de Repouso do Ciclo Celular/fisiologia , Ensaios Antitumorais Modelo de Xenoenxerto
12.
Dev Cell ; 49(2): 293-300.e3, 2019 04 22.
Artigo em Inglês | MEDLINE | ID: mdl-30905769

RESUMO

Quiescent neural stem cells (NSCs) in the adult brain are regenerative cells that could be activated therapeutically to repair damage. It is becoming apparent that quiescent NSCs exhibit heterogeneity in their propensity for activation and in the progeny that they generate. We discovered recently that NSCs undergo quiescence in either G0 or G2 in the Drosophila brain, challenging the notion that all quiescent stem cells are G0 arrested. We found that G2-quiescent NSCs become activated prior to G0 NSCs. Here, we show that the cyclin-dependent kinase inhibitor Dacapo (Dap; ortholog of p57KIP2) determines whether NSCs enter G0 or G2 quiescence during embryogenesis. We demonstrate that the dorsal patterning factor, Muscle segment homeobox (Msh; ortholog of MSX1/2/3) binds directly to the Dap locus and induces Dap expression in dorsal NSCs, resulting in G0 arrest, while more ventral NSCs undergo G2 quiescence. Our results reveal region-specific regulation of stem cell quiescence.


Assuntos
Inibidor de Quinase Dependente de Ciclina p57/metabolismo , Proteínas de Drosophila/metabolismo , Células-Tronco Neurais/metabolismo , Proteínas Nucleares/metabolismo , Células-Tronco Adultas/citologia , Células-Tronco Adultas/metabolismo , Animais , Ciclo Celular/fisiologia , Divisão Celular/fisiologia , Drosophila melanogaster/citologia , Drosophila melanogaster/metabolismo , Fase G2/fisiologia , Proteínas de Homeodomínio/metabolismo , Células-Tronco Neurais/citologia , Neurogênese/fisiologia , Fase de Repouso do Ciclo Celular/fisiologia
13.
Insect Biochem Mol Biol ; 102: 75-83, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-30287269

RESUMO

Insects enter diapause to synchronize their life cycle with biotic and abiotic conditions favorable for their development, reproduction, and survival. Adult females of the band-legged ground cricket Dianemobius nigrofasciatus (Orthoptera, Glyllidae) respond to environmental factors in autumn and lay diapause-destined eggs. The eggs arrest their development and enter diapause at a very early embryonic stage, specifically the cellular blastoderm. To elucidate the physiological mechanisms underlying this very early stage programmed developmental arrest, we investigated the cell division cycle as well as the expression of cell cycle regulators, small silencing RNAs, and segment patterning genes. The diapause embryo arrests its cell cycle predominantly at the G0/G1 phase. The proportion of cells in the S phase of the cell cycle abruptly decreased at the time of developmental arrest, but further changes of the G0/G1 and G2/M were later observed. Thus, cell cycle arrest in the diapause embryo is not an immediate event, but it takes longer to reach the steady state. We further elucidated molecular events possibly involved in diapause preparation and entry. Downregulation of Proliferating cellular antigen (PCNA; a cell cycle regulator), caudal and pumilio (cad and pum; early segmentation genes) as well as P-element induced wimpy testis (piwi) (a small silencing RNA) prior to the onset of developmental arrest was notable. The downregulation of PCNA, cad and pum continued even after entry into developmental arrest. In contrast to upregulation in non-diapause eggs, Cyclin D (another cell cycle regulator) and hunchback, Krüppel, and runt (gap and pair-rule genes) were downregulated in diapause eggs. These molecular events may contribute to embryonic diapause of D. nigrofasciatus.


Assuntos
Blastoderma/embriologia , Pontos de Checagem da Fase G1 do Ciclo Celular/fisiologia , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Gryllidae/embriologia , RNA Interferente Pequeno/biossíntese , Fase de Repouso do Ciclo Celular/fisiologia , Animais , Gryllidae/genética , RNA Interferente Pequeno/genética
14.
Sci Adv ; 4(8): eaat5685, 2018 08.
Artigo em Inglês | MEDLINE | ID: mdl-30116786

RESUMO

Quiescent (G0 phase) cells must maintain mitotic competence (MC) to restart the cell cycle. This is essential for reproduction in unicellular organisms and also for development and cell replacement in higher organisms. Recently, suppression of MC has gained attention as a possible therapeutic strategy for cancer. Using a Schizosaccharomyces pombe deletion-mutant library, we identified 85 genes required to maintain MC during the G0 phase induced by nitrogen deprivation. G0 cells must recycle proteins and RNA, governed by anabolism, catabolism, transport, and availability of small molecules such as antioxidants. Protein phosphatases are also essential to maintain MC. In particular, Nem1-Spo7 protects the nucleus from autophagy by regulating Ned1, a lipin. These genes, designated GZE (G-Zero Essential) genes, reveal the landscape of genetic regulation of MC.


Assuntos
Autofagia , Núcleo Celular/genética , Regulação Fúngica da Expressão Gênica , Mitose , Fase de Repouso do Ciclo Celular/fisiologia , Proteínas de Schizosaccharomyces pombe/genética , Schizosaccharomyces/genética , Células Cultivadas , Metaboloma , Schizosaccharomyces/crescimento & desenvolvimento , Schizosaccharomyces/metabolismo , Proteínas de Schizosaccharomyces pombe/metabolismo
15.
Chromosome Res ; 26(3): 179-189, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-29679205

RESUMO

In female mammals, each cell silences one X chromosome by converting it into transcriptionally inert heterochromatin. The inactivation is concomitant with epigenetic changes including methylation of specific histone residues and incorporation of macroH2A. Such epigenetic changes may exert influence on the positioning of the inactive X chromosome (Xi) within the nucleus beyond the level of chromatin structure. However, the dynamic positioning of the inactive X chromosome during cell cycle remains unclear. Here, we show that H3K27me3 is a cell-cycle-independent marker for the inactivated X chromosomes in WI38 cells. By utilizing this marker, three types of Xi locations in the nuclei are classified, which are envelope position (associated with envelope), mid-position (between the envelope and nucleolus), and nucleolus position (associated with the nucleolus). Moreover, serial-section analysis revealed that the inactive X chromosomes in the mid-position appear to be sparser and less condensed than those associated with the nuclear envelope or nucleolus. During the transition from G0 to G1 phase, the inactive X chromosomes tend to move from the envelope position to the nucleolus position in WI38 cells. Our results imply a role of chromosome positioning in maintaining the organization of the inactive X chromosomes in different cell phases.


Assuntos
Cromossomos de Mamíferos/metabolismo , Fase G1/fisiologia , Fase de Repouso do Ciclo Celular/fisiologia , Inativação do Cromossomo X/fisiologia , Cromossomo X/metabolismo , Animais , Linhagem Celular , Feminino , Camundongos
16.
Leukemia ; 32(4): 911-919, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-29209041

RESUMO

The E3 ubiquitin ligase (E3) WWP1 is an oncogenic factor implicated in the maintenance of different types of epithelial cancers. The role of WW domain-containing E3 ubiquitin protein ligase 1 (WWP1) in haematological neoplasms remains unknown. Acute myeloid leukaemia (AML) is characterized by the expansion of malignant myeloid cells blocked at different stages of differentiation. Here we report that the expression of WWP1 is significantly augmented in a large cohort of primary AML patients and in AML cell lines, compared with haematopoietic cells from healthy donors. We show that WWP1 inactivation severely impairs the growth of primary AML blasts and cell lines in vitro. In vivo, we observed a reduced leukaemogenic potential of WWP1-depleted AML cells upon transplantation into immunocompromised mice. Mechanistically, WWP1 inactivation induces the accumulation of its protein substrate p27Kip1, which ultimately contributes to G0/G1 cell cycle arrest of AML blasts. In addition, WWP1 depletion triggers the autophagy signalling and reduces survival of leukaemic cells. Collectively, our findings provide molecular insights into the anti-cancer potential of WWP1 inhibition, suggesting that this E3 is a promising biomarker and druggable target in AML.


Assuntos
Leucemia Mieloide Aguda/metabolismo , Leucemia Mieloide Aguda/patologia , Ubiquitina-Proteína Ligases/metabolismo , Animais , Pontos de Checagem do Ciclo Celular/fisiologia , Diferenciação Celular/fisiologia , Linhagem Celular Tumoral , Proliferação de Células/fisiologia , Sobrevivência Celular/fisiologia , Inibidor de Quinase Dependente de Ciclina p27/metabolismo , Fase G1/fisiologia , Regulação Neoplásica da Expressão Gênica/fisiologia , Humanos , Camundongos , Fase de Repouso do Ciclo Celular/fisiologia , Transdução de Sinais/fisiologia , Células U937 , Ubiquitinação/fisiologia
17.
J Biol Chem ; 293(4): 1353-1362, 2018 01 26.
Artigo em Inglês | MEDLINE | ID: mdl-29217771

RESUMO

It has been long assumed that post-mitotic neurons only utilize the error-prone non-homologous end-joining pathway to repair double-strand breaks (DSBs) associated with oxidative damage to DNA, given the inability of non-replicating neuronal DNA to utilize a sister chromatid template in the less error-prone homologous recombination (HR) repair pathway. However, we and others have found recently that active transcription triggers a replication-independent recombinational repair mechanism in G0/G1 phase of the cell cycle. Here we observed that the HR repair protein RAD52 is recruited to sites of DNA DSBs in terminally differentiated, post-mitotic neurons. This recruitment is dependent on the presence of a nascent mRNA generated during active transcription, providing evidence that an RNA-templated HR repair mechanism exists in non-dividing, terminally differentiated neurons. This recruitment of RAD52 in neurons is decreased by transcription inhibition. Importantly, we found that high concentrations of amyloid ß, a toxic protein associated with Alzheimer's disease, inhibits the expression and DNA damage response of RAD52, potentially leading to a defect in the error-free, RNA-templated HR repair mechanism. This study shows a novel RNA-dependent repair mechanism of DSBs in post-mitotic neurons and demonstrates that defects in this pathway may contribute to neuronal genomic instability and consequent neurodegenerative phenotypes such as those seen in Alzheimer's disease.


Assuntos
Quebras de DNA de Cadeia Dupla , Mitose/fisiologia , Neurônios/metabolismo , RNA/metabolismo , Proteína Rad52 de Recombinação e Reparo de DNA/metabolismo , Recombinação Genética/fisiologia , Animais , Fase G1/fisiologia , Neurônios/citologia , RNA/genética , Proteína Rad52 de Recombinação e Reparo de DNA/genética , Ratos , Fase de Repouso do Ciclo Celular/fisiologia
18.
Cell Physiol Biochem ; 43(5): 2022-2036, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-29059680

RESUMO

BACKGROUND/AIMS: In order to further characterize the biological traits of Dp71, HBE over expressing two most abundantly expressed Dp71 spliced isoforms, Dp71d and Dp71f, were established and their biological traits were explored. METHODS: The proliferation, migration and invasion capabilities of HBE-Dp71d and HBE-Dp71f cells were evaluated by MTT, colony formation, transwell and scratch assay. Cell cycle and apoptosis induced by H2O2 were measured by flow cytometer. Co-IP was performed to prove the interaction between lamin B1, FAK and Dp71. Western blot was performed to detect lamin B1, FAK, ERK and Cyclin D expression in HBE-Dp71d and HBE-Dp71f cells. RESULTS: HBE-Dp71d and HBE-Dp71f cells proliferated faster than their mock and blank controls; shortened their G0/G1 phase; enhanced their invasion and migration capabilities; reduced their apoptosis induced by H2O2. Co-IP proved Dp71 directly interacting with focal adhesion kinase (FAK) and lamin B1 in HBE cells. Increased lamin B1, FAK mRNA and protein expression, over activation of integrin/focal adhesion kinase/extracellular signal-regulated kinase (ERK)/cyclin D pathway were observed in HBE-Dp71d and HBE-Dp71f cells. CONCLUSIONS: Via increasing FAK in the cytoplasmic FAK-Dp71 , lamin B1 of nucleus laminB1-Dp71 complex, HBE-Dp71d and HBE-Dp71f cells alter their proliferation, migration, invasion, cell cycle and apoptosis rate induced by H2O2.


Assuntos
Distrofina/metabolismo , Animais , Apoptose/efeitos dos fármacos , Apoptose/genética , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Movimento Celular/genética , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Ciclina D/metabolismo , Distrofina/genética , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Quinase 1 de Adesão Focal/metabolismo , Fase G1/genética , Fase G1/fisiologia , Células HeLa , Humanos , Peróxido de Hidrogênio/farmacologia , Células PC12 , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Ratos , Fase de Repouso do Ciclo Celular/genética , Fase de Repouso do Ciclo Celular/fisiologia
19.
Ann Bot ; 120(4): 495-509, 2017 10 17.
Artigo em Inglês | MEDLINE | ID: mdl-28981580

RESUMO

Background: Quiescence is a fundamental feature of plant life, which enables plasticity, renewal and fidelity of the somatic cell line. Cellular quiescence is defined by arrest in a particular phase of the cell cycle, typically G1 or G2; however, the regulation of quiescence and proliferation can also be considered across wider scales in space and time. As such, quiescence is a defining feature of plant development and phenology, from meristematic stem cell progenitors to terminally differentiated cells, as well as dormant or suppressed seeds and buds. While the physiology of each of these states differs considerably, each is referred to as 'cell cycle arrest' or 'G1 arrest'. Scope: Here the physiology and molecular regulation of (1) meristematic quiescence, (2) dormancy and (3) terminal differentiation (cell cycle exit) are considered in order to determine whether and how the molecular decisions guiding these nuclear states are distinct. A brief overview of the canonical cell cycle regulators is provided, and the genetic and genomic, as well as physiological, evidence is considered regarding two primary questions: (1) Are the canonical cell cycle regulators superior or subordinate in the regulation of quiescence? (2) Are these three modes of quiescence governed by distinct molecular controls? Conclusion: Meristematic quiescence, dormancy and terminal differentiation are each predominantly characterized by G1 arrest but regulated distinctly, at a level largely superior to the canonical cell cycle. Meristematic quiescence is intrinsically linked to non-cell-autonomous regulation of meristem cell identity, and particularly through the influence of ubiquitin-dependent proteolysis, in partnership with reactive oxygen species, abscisic acid and auxin. The regulation of terminal differentiation shares analogous features with meristematic quiescence, albeit with specific activators and a greater role for cytokinin signalling. Dormancy meanwhile appears to be regulated at the level of chromatin accessibility, by Polycomb group-type histone modifications of particular dormancy genes.


Assuntos
Pontos de Checagem do Ciclo Celular/fisiologia , Desenvolvimento Vegetal/fisiologia , Dormência de Plantas/fisiologia , Fase G1/fisiologia , Meristema/crescimento & desenvolvimento , Meristema/fisiologia , Raízes de Plantas/crescimento & desenvolvimento , Raízes de Plantas/fisiologia , Brotos de Planta/crescimento & desenvolvimento , Brotos de Planta/fisiologia , Plantas , Fase de Repouso do Ciclo Celular/fisiologia , Fase S/fisiologia
20.
Biomed Pharmacother ; 84: 177-184, 2016 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-27657825

RESUMO

OBJECTIVE: Curcumin as an effective anticancer bioactive extract has been proved to induce apoptosis in many cancer cells. Notch signaling regulates prostate cancer apoptosis, but it is still unknown whether curcumin induces apoptosis in DU-145 cells by regulating Notch pathway. The aim of this study was to investigate the effect of curcumin on regulating Notch signaling and provide basic data for using curcumin in prostate cancer therapy. METHODS: Notch pathway signal related proteins Notch 1, Jagged-1 and NICD were detected using Western blotting and RT-PCR. The proliferation and apoptosis were determined by MTT method and Elisa kits after curcumin treatment, respectively. Dual-Luciferase Reporter Assay was carried out to confirm that curcumin could target Notching signaling. In order to study whether Notch 1 expression could be downregulated by curcumin, Notch 1 siRNA and Notch 1 plasmid were used in Notch 1 down-regulation and over-expression. The effects of curcumin on cell cycle distribution and apoptosis related proteins expression were analyzed by flow cytometry and western blotting, separately. RESULTS: We found that Notch 1 signaling was down regulated in Notch 1 siRNA or Notch 1 plasmid transfected 145 cells after curcumin treatment. Curcumin induced G0/G1 arrest in DU-145 cells, and G0/G1 phase related regulatory factors Cyclin D1 and CDK2 expressions were inhibited. Meanwhile, p21 and p27 were up regulated. The apoptosis related protein p53 expression was increased, and apoptosis suppressor Bcl-2 was inhibited in DU-145 after curcumin treatment. Additionally, Caspase-3 and Caspase-9 were activated by curcumin. CONCLUSION: Curcumin induced apoptosis and G0/G1 arrest in DU-145 cells by down regulating Notch signaling.


Assuntos
Apoptose/efeitos dos fármacos , Curcumina/farmacologia , Fase G1/efeitos dos fármacos , Neoplasias da Próstata , Receptor Notch1/antagonistas & inibidores , Fase de Repouso do Ciclo Celular/efeitos dos fármacos , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Apoptose/fisiologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/fisiologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Curcumina/uso terapêutico , Relação Dose-Resposta a Droga , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/fisiologia , Fase G1/fisiologia , Humanos , Masculino , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/metabolismo , Receptor Notch1/metabolismo , Fase de Repouso do Ciclo Celular/fisiologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia
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