RESUMO
Telomeres are essential for genome stability. Oxidative stress caused by excess reactive oxygen species (ROS) accelerates telomere shortening. Although telomeres are hypersensitive to ROS-mediated 8-oxoguanine (8-oxoG) formation, the biological effect of this common lesion at telomeres is poorly understood because ROS have pleiotropic effects. Here we developed a chemoptogenetic tool that selectively produces 8-oxoG only at telomeres. Acute telomeric 8-oxoG formation increased telomere fragility in cells lacking OGG1, the enzyme that removes 8-oxoG, but did not compromise cell survival. However, chronic telomeric 8-oxoG induction over time shortens telomeres and impairs cell growth. Accumulation of telomeric 8-oxoG in chronically exposed OGG1-deficient cells triggers replication stress, as evidenced by mitotic DNA synthesis at telomeres, and significantly increases telomere losses. These losses generate chromosome fusions, leading to chromatin bridges and micronucleus formation upon cell division. By confining base damage to the telomeres, we show that telomeric 8-oxoG accumulation directly drives telomere crisis.
Assuntos
Aberrações Cromossômicas/efeitos da radiação , DNA Glicosilases/genética , Reparo do DNA/efeitos da radiação , Instabilidade Genômica/efeitos da radiação , Guanina/análogos & derivados , Telômero/efeitos da radiação , Divisão Celular/efeitos da radiação , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos da radiação , Dano ao DNA , DNA Glicosilases/deficiência , Replicação do DNA/efeitos da radiação , Expressão Gênica , Guanina/agonistas , Guanina/biossíntese , Células HeLa , Humanos , Luz/efeitos adversos , Micronúcleos com Defeito Cromossômico/efeitos da radiação , Optogenética , Osteoblastos/citologia , Osteoblastos/metabolismo , Osteoblastos/efeitos da radiação , Estresse Oxidativo/efeitos da radiação , Oxigênio Singlete/agonistas , Oxigênio Singlete/metabolismo , Telômero/metabolismo , Homeostase do Telômero/efeitos da radiaçãoRESUMO
Biological signals need to be robust and filter small fluctuations yet maintain sensitivity to signals across a wide range of magnitudes. Here, we studied how fluctuations in DNA damage signaling relate to maintenance of long-term cell-cycle arrest. Using live-cell imaging, we quantified division profiles of individual human cells in the course of 1 week after irradiation. We found a subset of cells that initially establish cell-cycle arrest and then sporadically escape and divide. Using fluorescent reporters and mathematical modeling, we determined that fluctuations in the oscillatory pattern of the tumor suppressor p53 trigger a sharp switch between p21 and CDK2, leading to escape from arrest. Transient perturbation of p53 stability mimicked the noise in individual cells and was sufficient to trigger escape from arrest. Our results show that the self-reinforcing circuitry that mediates cell-cycle transitions can translate small fluctuations in p53 signaling into large phenotypic changes.
Assuntos
Quinase 2 Dependente de Ciclina/metabolismo , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Células Epiteliais/metabolismo , Modelos Estatísticos , Transdução de Sinais , Proteína Supressora de Tumor p53/metabolismo , Pontos de Checagem do Ciclo Celular/genética , Pontos de Checagem do Ciclo Celular/efeitos da radiação , Divisão Celular/efeitos da radiação , Linhagem Celular Transformada , Proliferação de Células/efeitos da radiação , Quinase 2 Dependente de Ciclina/genética , Inibidor de Quinase Dependente de Ciclina p21/genética , Dano ao DNA , Células Epiteliais/citologia , Células Epiteliais/efeitos da radiação , Raios gama , Regulação da Expressão Gênica , Genes Reporter , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Humanos , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Estabilidade Proteica , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Epitélio Pigmentado da Retina/citologia , Epitélio Pigmentado da Retina/metabolismo , Epitélio Pigmentado da Retina/efeitos da radiação , Imagem com Lapso de Tempo , Proteína Supressora de Tumor p53/antagonistas & inibidores , Proteína Supressora de Tumor p53/genética , Proteína Vermelha FluorescenteRESUMO
Light-regulated modules offer unprecedented new ways to control cellular behaviour with precise spatial and temporal resolution. Among a variety of bacterial light-switchable gene expression systems, single-component systems consisting of single transcription factors would be more useful due to the advantages of speed, simplicity, and versatility. In the present study, we developed a single-component light-activated bacterial gene expression system (eLightOn) based on a novel LOV domain from Rhodobacter sphaeroides (RsLOV). The eLightOn system showed significant improvements over the existing single-component bacterial light-activated expression systems, with benefits including a high ON/OFF ratio of >500-fold, a high activation level, fast activation kinetics, and/or good adaptability. Additionally, the induction characteristics, including regulatory windows, activation kinetics and light sensitivities, were highly tunable by altering the expression level of LexRO. We demonstrated the usefulness of the eLightOn system in regulating cell division and swimming by controlling the expression of the FtsZ and CheZ genes, respectively, as well as constructing synthetic Boolean logic gates using light and arabinose as the two inputs. Taken together, our data indicate that the eLightOn system is a robust and highly tunable tool for quantitative and spatiotemporal control of bacterial gene expression.
Assuntos
Regulação Bacteriana da Expressão Gênica/efeitos da radiação , Luz , Rhodobacter sphaeroides/citologia , Rhodobacter sphaeroides/efeitos da radiação , Proteínas de Bactérias/metabolismo , Divisão Celular/efeitos da radiação , Cinética , Lógica , Fatores de Transcrição/metabolismoRESUMO
The cell fusion is a widespread process, which takes place in many systems in vivo and in vitro. Fusion of cells is frequently related to tetraploidy, which can be found within natural physiological conditions, e.g., placentation, and in pathophysiological conditions, such as cancer and early pregnancy failure in humans. Here we investigate the mechanism of tetraploidization with help of femtosecond laser-induced mouse blastomere fusion by the means of Hoechst staining, GFP, BODIPY dyes and fluorescent species generated intracellularly by a femtosecond laser. We establish diffusive mixing of cytosol, whereas the large components of a cytoplasm (organelles, cytoskeleton) are poorly diffusible and are not completely mixed after cell fusion and a subsequent division. We show that mechanisms which are responsible for the formation of a common metaphase plate triggered tetraploidization in fused mouse embryos and could be a significant factor in polyploidy formation in vivo. Thus, our results suggest that microtubules play a critical role in tetraploidization.
Assuntos
Blastômeros/fisiologia , Blastômeros/efeitos da radiação , Lasers , Tetraploidia , Animais , Blastômeros/citologia , Divisão Celular/efeitos da radiação , Fusão Celular/métodos , Embrião de Mamíferos/citologia , Embrião de Mamíferos/efeitos da radiação , Feminino , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Masculino , Metáfase/fisiologia , Metáfase/efeitos da radiação , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , GravidezRESUMO
All organisms regulate cell cycle progression by coordinating cell division with DNA replication status. In eukaryotes, DNA damage or problems with replication fork progression induce the DNA damage response (DDR), causing cyclin-dependent kinases to remain active, preventing further cell cycle progression until replication and repair are complete. In bacteria, cell division is coordinated with chromosome segregation, preventing cell division ring formation over the nucleoid in a process termed nucleoid occlusion. In addition to nucleoid occlusion, bacteria induce the SOS response after replication forks encounter DNA damage or impediments that slow or block their progression. During SOS induction, Escherichia coli expresses a cytoplasmic protein, SulA, that inhibits cell division by directly binding FtsZ. After the SOS response is turned off, SulA is degraded by Lon protease, allowing for cell division to resume. Recently, it has become clear that SulA is restricted to bacteria closely related to E. coli and that most bacteria enforce the DNA damage checkpoint by expressing a small integral membrane protein. Resumption of cell division is then mediated by membrane-bound proteases that cleave the cell division inhibitor. Further, many bacterial cells have mechanisms to inhibit cell division that are regulated independently from the canonical LexA-mediated SOS response. In this review, we discuss several pathways used by bacteria to prevent cell division from occurring when genome instability is detected or before the chromosome has been fully replicated and segregated.
Assuntos
Divisão Celular/efeitos dos fármacos , Divisão Celular/efeitos da radiação , Replicação do DNA/efeitos dos fármacos , Replicação do DNA/efeitos da radiação , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Bleomicina/farmacologia , Ciclo Celular/efeitos dos fármacos , Ciclo Celular/genética , Ciclo Celular/efeitos da radiação , Divisão Celular/genética , Dano ao DNA/efeitos dos fármacos , Dano ao DNA/efeitos da radiação , Replicação do DNA/genética , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Escherichia coli/metabolismo , Escherichia coli/efeitos da radiação , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Protease La/genética , Protease La/metabolismo , Radiação Ionizante , Resposta SOS em Genética/efeitos dos fármacos , Resposta SOS em Genética/genética , Resposta SOS em Genética/efeitos da radiaçãoRESUMO
Both SIRT1 and UVA radiation are involved in cellular damage processes such as apoptosis, senescence and ageing. MicroRNAs (miRNAs) have been reported to be closely related to UV radiation, as well as to SIRT1. In this study, we investigated the connections among SIRT1, UVA and miRNA in human skin primary fibroblasts. Our results showed that UVA altered the protein level of SIRT1 in a time point-dependent manner. Using miRNA microarray, bioinformatics analysis, we found that knocking down SIRT1 could cause up-regulation of miR-27a-5p and the latter could down-regulate SMAD2, and these results were verified by qRT-PCR or Western blot. Furthermore, UVA radiation (5 J/cm2 ), knocking down SIRT1 or overexpression of miR-27a-5p led to increased expression of MMP1, and decreased expressions of COL1 and BCL2. We also found additive impacts on MMP1, COL1 and BCL2 under the combination of UVA radiation + Sirtinol (SIRT1 inhibitor), or UVA radiation + miR-27a-5p mimic. SIRT1 activator resveratrol could reverse damage changes caused by UVA radiation. Besides, absent of SIRT1 or overexpression of miR-27a-5p increased cell apoptosis and induced cell arrest in G2/M phase. Taken together, these results demonstrated that UVA could influence a novel SIRT1-miR-27a-5p-SMAD2-MMP1/COL1/BCL2 axis in skin primary fibroblasts, and may provide potential therapeutic targets for UVA-induced skin damage.
Assuntos
Fibroblastos/metabolismo , Fibroblastos/efeitos da radiação , Proteínas/metabolismo , Transdução de Sinais/efeitos da radiação , Pele/metabolismo , Pele/efeitos da radiação , Raios Ultravioleta/efeitos adversos , Adolescente , Adulto , Apoptose/efeitos da radiação , Pontos de Checagem do Ciclo Celular/efeitos da radiação , Divisão Celular/efeitos da radiação , Células Cultivadas , Regulação para Baixo/efeitos da radiação , Fase G2/efeitos da radiação , Humanos , Regulação para Cima/efeitos da radiação , Adulto JovemRESUMO
Gastrointestinal toxicity limits the clinical application of abdominal and pelvic radiotherapy and currently has no effective treatment. Intestinal leucine-rich-repeat-containing GPCR 5 (Lgr5)-positive stem cell depletion and loss of proliferative ability due to radiation may be the primary factors causing intestinal injury following radiation. Here, we report the critical role of ß-arrestin1 (ßarr1) in radiation-induced intestinal injury. Intestinal ßarr1 was highly expressed in radiation enteritis and in a radiation model. ßarr1 knockout (KO) or knockdown mice exhibited increased proliferation in intestinal Lgr5+ stem cell, crypt reproduction, and survival following radiation. Unexpectedly, the beneficial effects of ßarr1 deficiency on intestinal stem cells in response to radiation were compromised when the endoplasmic reticulum stress-related protein kinase RNA-like ER kinase (PERK)/eukaryotic initiation factor-2α (eIF2α) pathway was inhibited, and this result was further supported in vitro. Furthermore, we found that ßarr1 knockdown with small interfering RNA significantly enhanced intestinal Lgr5+ stem cell proliferation after radiation via directly targeting PERK. ßarr1 offers a promising target for mitigating radiation-induced intestinal injury.-Liu, Z., Jiang, J., He, Q., Liu, Z., Yang, Z., Xu, J., Huang, Z., Wu, B. ß-Arrestin1-mediated decrease in endoplasmic reticulum stress impairs intestinal stem cell proliferation following radiation.
Assuntos
Estresse do Retículo Endoplasmático/fisiologia , Enterite/patologia , Intestinos/efeitos da radiação , Lesões Experimentais por Radiação/patologia , Lesões por Radiação/patologia , Células-Tronco/efeitos da radiação , beta-Arrestina 1/fisiologia , eIF-2 Quinase/fisiologia , Idoso , Animais , Divisão Celular/efeitos da radiação , Ensaio de Unidades Formadoras de Colônias , Enterite/etiologia , Enterite/fisiopatologia , Fator de Iniciação 2 em Eucariotos/fisiologia , Feminino , Técnicas de Silenciamento de Genes , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pessoa de Meia-Idade , Interferência de RNA , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/farmacologia , Quimera por Radiação , Lesões por Radiação/fisiopatologia , Lesões Experimentais por Radiação/fisiopatologia , Radioterapia/efeitos adversos , Receptores Acoplados a Proteínas G/análise , Regeneração , Transdução de Sinais/fisiologia , Células-Tronco/patologia , beta-Arrestina 1/deficiência , beta-Arrestina 1/genéticaRESUMO
Temporal programs synchronised with the daily cycle are of adaptive importance for organisms exposed to periodic fluctuations. This study deepens into several aspects of the exogenous and endogenous nature of microbial grazers. We investigated the diel rhythms of cell division and feeding activity of four marine protists under different light regimes. In particular, we tested if the feeding cycle of protistan grazers could be mediated by a light-aided enhancement of prey digestion, and also explored the consequences of cell division on diel feeding rhythms. Cell division occurred at night for the heterotrophic dinoflagellates Gyrodinium dominans and Oxyrrhis marina. In contrast, the mixotrophic dinoflagellate Karlodinium armiger and the ciliate Strombidium sp. mostly divided during the day. Additionally, a significant diurnal feeding rhythm was observed in all species. When exposed to continuous darkness, nearly all species maintained the cell division rhythm, but lost the feeding cycle within several hours/days (with the exception of O. marina that kept the rhythm for 9.5 days). Additional feeding experiments under continuous light also showed the same pattern. We conclude that the feeding rhythms of protistan grazers are generally regulated not by cell division nor by the enhancement of digestion by light. Our study, moreover, indicates that the cell division cycle is under endogenous control, whereas an external trigger is required to maintain the feeding rhythm, at least for most of the species studied here.
Assuntos
Cilióforos/fisiologia , Dinoflagellida/fisiologia , Divisão Celular/efeitos da radiação , Cilióforos/efeitos da radiação , Dinoflagellida/efeitos da radiação , Processos Heterotróficos , LuzRESUMO
Ciprofloxacin hydrochloride and Norfloxacin are second-generation fluoroquinolone antibiotic against bacterial DNA gyrase, which reduces DNA strain throughout replication. As DNA gyrase is essential through DNA replication, subsequent DNA synthesis and cell division are inhibited. Direct photolysis of fluoroquinolones was studied by using UV irradiation in the presence or absence of other substances that generate free radicals. This study aimed to assess the effect of Ultraviolet B (UVB) irradiation in removing ciprofloxacin and norfloxacin by using a simulating model of wastewater contained urea at pH 4. A known concentration of ciprofloxacin and norfloxacin were prepared in an appropriate aqueous solution in presence or absence 0.2M urea and adjusted at pH 4. The dis-solved drugs were irradiated with UVB-lamp in a dark place for 60 minutes. The percent of removal and the rate of elimination (k) of each drug were calculated. The direct photolysis effect of UVB irradiation was observed with ciprofloxacin which amounted to 24.4% removal compared with12.4% removal of norfloxacin after 60 minutes of irradiation. The effect of UVB irradiation was enhanced by urea to reach 38.9% and 15% for ciprofloxacin and norfloxacin. The calculated k of ciprofloxacin has amounted to three folds of that of norfloxacin. Direct photolysis of ciprofloxacin and norfloxacin can be achieved simply by using a simulation model of 0.2 M urea and UVB irradiation at pH 4. UVB is highly effective in removing ciprofloxacin compared with norfloxacin by 2-3 folds.
Assuntos
Divisão Celular/efeitos dos fármacos , Ciprofloxacina/farmacologia , Replicação do DNA/efeitos dos fármacos , DNA Bacteriano/efeitos dos fármacos , Norfloxacino/farmacologia , Raios Ultravioleta , Ureia/química , Divisão Celular/efeitos da radiação , Ciprofloxacina/efeitos da radiação , Meios de Cultura , Replicação do DNA/efeitos da radiação , DNA Bacteriano/efeitos da radiação , Norfloxacino/efeitos da radiação , Análise de RegressãoRESUMO
Cellular fate decisions are influenced by their topographical location in the adult body. For instance, tissue repair and neoplastic growth are greater in anterior than in posterior regions of adult animals. However, the molecular underpinnings of these regional differences are unknown. We identified a regional switch in the adult planarian body upon systemic disruption of homologous recombination with RNA-interference of Rad51 Rad51 knockdown increases DNA double-strand breaks (DSBs) throughout the body, but stem cells react differently depending on their location along the anteroposterior axis. In the presence of extensive DSBs, cells in the anterior part of the body resist death, whereas cells in the posterior region undergo apoptosis. Furthermore, we found that proliferation of cells with DNA damage is induced in the presence of brain tissue and that the retinoblastoma pathway enables overproliferation of cells with DSBs while attending to the demands of tissue growth and repair. Our results implicate both autonomous and non-autonomous mechanisms as key mediators of regional cell behavior and cellular transformation in the adult body.
Assuntos
Linhagem da Célula , Instabilidade Genômica , Planárias/citologia , Planárias/genética , Células-Tronco/citologia , Animais , Padronização Corporal/efeitos da radiação , Morte Celular/efeitos da radiação , Diferenciação Celular/efeitos da radiação , Divisão Celular/efeitos da radiação , Linhagem da Célula/efeitos da radiação , Proliferação de Células/efeitos da radiação , DNA/metabolismo , Quebras de DNA de Cadeia Dupla/efeitos da radiação , Dano ao DNA , Reparo do DNA por Junção de Extremidades/efeitos da radiação , Regulação para Baixo/efeitos da radiação , Raios gama , Instabilidade Genômica/efeitos da radiação , Recombinação Homóloga/efeitos da radiação , Especificidade de Órgãos/efeitos da radiação , Planárias/efeitos da radiação , Interferência de RNA/efeitos da radiação , Rad51 Recombinase/metabolismo , Radiação Ionizante , Proteína do Retinoblastoma/metabolismo , Transdução de Sinais/efeitos da radiação , Células-Tronco/metabolismo , Células-Tronco/efeitos da radiação , Cicatrização/efeitos da radiaçãoRESUMO
Ultraviolet-B (UV-B) radiation affects leaf growth in a wide range of species. In this work, we demonstrate that UV-B levels present in solar radiation inhibit maize (Zea mays) leaf growth without causing any other visible stress symptoms, including the accumulation of DNA damage. We conducted kinematic analyses of cell division and expansion to understand the impact of UV-B radiation on these cellular processes. Our results demonstrate that the decrease in leaf growth in UV-B-irradiated leaves is a consequence of a reduction in cell production and a shortened growth zone (GZ). To determine the molecular pathways involved in UV-B inhibition of leaf growth, we performed RNA sequencing on isolated GZ tissues of control and UV-B-exposed plants. Our results show a link between the observed leaf growth inhibition and the expression of specific cell cycle and developmental genes, including growth-regulating factors (GRFs) and transcripts for proteins participating in different hormone pathways. Interestingly, the decrease in the GZ size correlates with a decrease in the concentration of GA19, the immediate precursor of the active gibberellin, GA1, by UV-B in this zone, which is regulated, at least in part, by the expression of GRF1 and possibly other transcription factors of the GRF family.
Assuntos
Giberelinas/metabolismo , Reguladores de Crescimento de Plantas/metabolismo , Folhas de Planta/crescimento & desenvolvimento , Folhas de Planta/efeitos da radiação , Raios Ultravioleta , Zea mays/crescimento & desenvolvimento , Zea mays/efeitos da radiação , Fenômenos Biomecânicos , Divisão Celular/efeitos da radiação , Proliferação de Células/efeitos da radiação , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas/efeitos da radiação , MicroRNAs/genética , MicroRNAs/metabolismo , Epiderme Vegetal/citologia , Epiderme Vegetal/efeitos da radiação , Transcriptoma/genética , Zea mays/genéticaRESUMO
BACKGROUND: Cantharidin is an inhibitor of protein phosphatase 2â¯A (PP2A), and has been frequently used in clinical practice. In our previous study, we proved that cantharidin could arrest cell cycle in G2/M phase. Since cells at G2/M phase are sensitive to radiotherapy, in the present study, we investigated the radiotherapy-sesitization effect of cantharidin and the potential mechanisms involved. METHODS: Cell growth was determined by MTT assay. Cell cycle was evaluated by flow cytometry. DNA damage was visualized by phospho-Histone H2A.X staining. Expression of mRNA was tested by microarray assay and real-time PCR. Clinical information and RNA-Seq expression data were derived from The Cancer Genome Atlas (TCGA) pancreatic cancer cohort. Survival analysis was obtained by Kaplan-Meier estimates. RESULTS: Cantharidin strengthened the growth inhibition effect of irradiation. Cantharidin drove pancreatic cancer cells out of quiescent G0/G1 phase and arrested cell cycle in G2/M phase. As a result, cantharidin strengthened DNA damage which was induced by irradiation. Moreover, cantharidin repressed expressions of several genes participating in DNA damage repair, including UBE2T, RPA1, GTF2HH5, LIG1, POLD3, RMI2, XRCC1, PRKDC, FANC1, FAAP100, RAD50, RAD51D, RAD51B and DMC1, through JNK, ERK, PKC, p38 and/or NF-κB pathway dependent manners. Among these genes, worse overall survival for pancreatic cancer patients were associated with high mRNA expressions of POLD3, RMI2, PRKDC, FANC1, RAD50 and RAD51B, all of which could be down-regulated by cantharidin. CONCLUSION: Cantharidin can sensitize pancreatic cancer cells to radiotherapy. Multiple mechanisms, including cell cycle regulation, enhanced DNA damage, and inhibited DNA damage repair, may be involved.
Assuntos
Cantaridina/farmacologia , Radiossensibilizantes/farmacologia , Radioterapia , Ciclo Celular , Divisão Celular/efeitos dos fármacos , Divisão Celular/efeitos da radiação , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos da radiação , Dano ao DNA , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Neoplasias Pancreáticas , Proteína Fosfatase 2/antagonistas & inibidoresRESUMO
There is compelling biological data to suggest that cancer arises from a series of mutations in single target cells, resulting in defects in cell renewal and differentiation processes which lead to malignancy. Because much mutagenic damage is expressed following cell division, more-rapidly renewing tissues could be at higher risk because of the larger number of cell replications. Cairns suggested that renewing tissues may reduce cancer risk by partitioning the dividing cell populations into lineages comprising infrequently-dividing long-lived stem cells and frequently-dividing short-lived daughter transit cells. We develop generalizations of three recent cancer-induction models that account for the joint maintenance and renewal of stem and transit cells, also competing processes of partially transformed cell proliferation and differentiation/apoptosis. We are particularly interested in using these models to separately assess the probabilities of mutation and development of cancer associated with "spontaneous" processes and with those linked to a specific environmental mutagen, specifically ionizing radiation or cigarette smoking. All three models demonstrate substantial variation in cancer risks, by at least 20 orders of magnitude, depending on the assumed number of critical mutations required for cancer, and the stem-cell and transition-cell mutation rates. However, in most cases the conditional probabilities of cancer being mutagen-induced range between 7-96%. The relative risks associated with mutagen exposure compared to background rates are also stable, ranging from 1.0-16.0. Very few cancers, generally <0.5%, arise from mutations occurring solely in stem cells rather than in a combination of stem and transit cells. However, for cancers with 2 or 3 critical mutations, a substantial proportion of cancers, in some cases 100%, have at least one mutation derived from a mutated stem cell. Little difference is made to relative risks if competing processes of proliferation and differentiation in the partially transformed stem and transit cell population are allowed for, nor is any difference made if one assumes that transit cells require an extra mutation to confer malignancy from the number required by stem cells. The probability of a cancer being mutagen-induced correlates across cancer sites with the estimated cumulative number of stem cell divisions in the associated tissue (p<0.05), although in some cases there is sensitivity of findings to removal of high-leverage outliers and in some cases only modest variation in probability, but these issues do not affect the validity of the findings. There are no significant correlations (p>0.3) between lifetime cancer-site specific radiation risk and the probability of that cancer being mutagen-induced. These results do not depend on the assumed critical number of mutations leading to cancer, or on the assumed mutagen-associated mutation rate, within the generally-accepted ranges tested. However, there are borderline significant negative correlations (p = 0.08) between the smoking-associated mortality rate difference (current vs former smokers) and the probability of cancer being mutagen-induced. This is only the case where values of the critical number of mutations leading to cancer, k, is 3 or 4 and not for smaller values (1 or 2), but does not strongly depend on the assumed mutagen-associated mutation rate.
Assuntos
Modelos Genéticos , Neoplasias Induzidas por Radiação/epidemiologia , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/efeitos da radiação , Nicotiana/toxicidade , Fumaça , Fumar , Animais , Divisão Celular/efeitos dos fármacos , Divisão Celular/efeitos da radiação , Transformação Celular Neoplásica/induzido quimicamente , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/efeitos da radiação , Simulação por Computador , Epigênese Genética/efeitos dos fármacos , Epigênese Genética/genética , Epigênese Genética/efeitos da radiação , Humanos , Mutação , Neoplasias Induzidas por Radiação/genéticaRESUMO
BACKGROUND: Cyanobacteria are important agents in global carbon and nitrogen cycling and hold great promise for biotechnological applications. Model organisms such as Synechocystis sp. and Synechococcus sp. have advanced our understanding of photosynthetic capacity and circadian behavior, mostly using population-level measurements in which the behavior of individuals cannot be monitored. Synechocystis sp. cells are small and divide slowly, requiring long-term experiments to track single cells. Thus, the cumulative effects of drift over long periods can cause difficulties in monitoring and quantifying cell growth and division dynamics. RESULTS: To overcome this challenge, we enhanced a microfluidic cell-culture device and developed an image analysis pipeline for robust lineage reconstruction. This allowed simultaneous tracking of many cells over multiple generations, and revealed that cells expand exponentially throughout their cell cycle. Generation times were highly correlated for sister cells, but not between mother and daughter cells. Relationships between birth size, division size, and generation time indicated that cell-size control was inconsistent with the "sizer" rule, where division timing is based on cell size, or the "timer" rule, where division occurs after a fixed time interval. Instead, single cell growth statistics were most consistent with the "adder" rule, in which division occurs after a constant increment in cell volume. Cells exposed to light-dark cycles exhibited growth and division only during the light period; dark phases pause but do not disrupt cell-cycle control. CONCLUSIONS: Our analyses revealed that the "adder" model can explain both the growth-related statistics of single Synechocystis cells and the correlation between sister cell generation times. We also observed rapid phenotypic response to light-dark transitions at the single cell level, highlighting the critical role of light in cyanobacterial cell-cycle control. Our findings suggest that by monitoring the growth kinetics of individual cells we can build testable models of circadian control of the cell cycle in cyanobacteria.
Assuntos
Divisão Celular , Rastreamento de Células/métodos , Microfluídica/métodos , Synechocystis/citologia , Divisão Celular/efeitos da radiação , Proliferação de Células , Processamento de Imagem Assistida por Computador , Luz , Modelos Biológicos , Probabilidade , Synechocystis/crescimento & desenvolvimento , Synechocystis/efeitos da radiação , Fatores de TempoRESUMO
In this study, we examined the dynamics of phenotypic and transcriptional profiles in Saccharomyces cerevisiae following semi-lethal X-ray irradiation. Post-irradiation, reproductive death was revealed as the predominant form of death in S. cerevisiae and almost all the irradiated cells were physically present and intact. In addition, cell cycle arrest reached its peak and cell division was at its valley at 2 h. Cell cycle arrest, cell division potential, DNA damage, and mitochondrial transmembrane potential (MTP) showed significant recovery at 4 h (P > 0.05 vs. control). The improvements of DNA damage and MTP decrease were evaluated as at least 77% and 84% for the original irradiated cells at 4 h, respectively. In the transcriptional profile, the amount of differentially expressed genes (DEGs) and the fold change in the repair-related DEGs were highest at 1 h post-irradiation and then decreased. The DEGs at 1 h (but not 2 h or 3 h) were significantly enriched in gene ontology (GO) categories of detoxification (up) and antioxidant activity (up). Although the transcriptional profile supported the repair time frame observed in the phenotypic profile, the complete repair may take a longer duration as the transcriptional levels of several important repair-related DEGs did not show a decrease and the DNA repair-related pathways (up) were the major enriched pathway in Kyoto Encyclopaedia of Genes and Genomes pathway analysis throughout the whole course of the study. These results provide an important reference for the selection of key time points in further studies.
Assuntos
Saccharomyces cerevisiae/crescimento & desenvolvimento , Saccharomyces cerevisiae/efeitos da radiação , Estresse Fisiológico , Raios X , Divisão Celular/efeitos da radiação , Dano ao DNA/efeitos da radiação , Perfilação da Expressão Gênica , Membranas Mitocondriais/efeitos da radiação , Fatores de Tempo , Transcrição GênicaRESUMO
The unicellular green microalga Haematococcus pluvialis has the highest content of the natural antioxidant, astaxanthin. Previously, it was determined that astaxanthin accumulation in H. pluvialis could be induced by blue-wavelength irradiation; however, the molecular mechanism remains unknown. The present study aimed to compare the transcriptome of H. pluvialis, with respect to astaxanthin biosynthesis, under the monochromatic red (660 nm) or blue (450 nm) light-emitting diode (LED) irradiation. Among a total of 165,372 transcripts, we identified 67,703 unigenes, of which 2245 and 171 were identified as differentially expressed genes (DEGs) in response to blue and red irradiation, respectively. Interestingly, expressional changes of blue light receptor cryptochromes were detected in response to blue and/or red LED irradiation in H. pluvialis, which may directly and indirectly regulate astaxanthin biosynthesis. In accordance with this observation, expression of the BKT and CHY genes, which are part of the downstream section of the astaxanthin biosynthetic pathway, was significantly upregulated by blue LED irradiation compared with their expression under control white irradiation. Contrastingly, they were downregulated by red LED irradiation. Our transcriptome study provided molecular insights that highlighted the different of responses of H. pluvialis to red and blue irradiation, especially for astaxanthin biosynthesis.
Assuntos
Clorófitas/genética , Clorófitas/metabolismo , Clorófitas/efeitos da radiação , Regulação da Expressão Gênica de Plantas/efeitos da radiação , Luz , Transcriptoma , Vias Biossintéticas/genética , Vias Biossintéticas/efeitos da radiação , Divisão Celular/efeitos da radiação , Clorófitas/crescimento & desenvolvimento , Análise por Conglomerados , Cor , Perfilação da Expressão Gênica , Ontologia Genética , Genes de Plantas/genética , Genes de Plantas/efeitos da radiação , Microbiologia Industrial , Iluminação , Espécies Reativas de Oxigênio/metabolismo , Análise de Sequência de RNA , Regulação para Cima , Xantofilas/biossíntese , Xantofilas/genéticaRESUMO
Oral squamous cell carcinoma (OSCC) is a serious public health problem caused primarily by smoking and alcohol consumption or human papillomavirus. The cancer stem cell (CSC) theory posits that CSCs show unique characteristics, including self-renewal and therapeutic resistance. Examining biomarkers and other features of CSCs is critical to better understanding their biology. To this end, the results show that cellular SOX2 immunostaining correlates with other CSC biomarkers in OSCC cell lines and marks the rare CSC population. To assess whether CSC division patterns are symmetrical, resulting in two CSC, or asymmetrical, leading to one CSC and one cancer cell, cell size and fluorescence intensity of mitotic cells stained with SOX2 were analyzed. Asymmetrical SOX2 distribution in ≈25% of the mitoses analyzed was detected. Chromosomal instability, some of which is caused by chromosome segregation defects (CSDs), is a feature of cancer cells that leads to altered gene copy numbers. We compare chromosomal instability (as measured by CSDs) between CSCs (SOX2+) and non-CSCs (SOX2-) from the same OSCC cell lines. CSDs were more common in non-CSCs (SOX2-) than CSCs (SOX2+) and in symmetrical CSC (SOX2+) mitotic pairs than asymmetrical CSC (SOX2+/SOX2-) mitotic pairs. CSCs showed fewer and different types of CSDs after ionizing radiation treatment than non-CSCs. Overall, these data are the first to demonstrate both symmetrical and asymmetrical cell divisions with CSDs in OSCC CSC. Further, the results suggest that CSCs may undergo altered behavior, including therapeutic resistance as a result of chromosomal instability due to chromosome segregation defects. © 2016 Wiley Periodicals, Inc.
Assuntos
Biomarcadores Tumorais/genética , Carcinoma de Células Escamosas/patologia , Divisão Celular/genética , Segregação de Cromossomos/genética , Neoplasias Bucais/patologia , Células-Tronco Neoplásicas/patologia , Fatores de Transcrição SOXB1/genética , Biomarcadores Tumorais/metabolismo , Carcinoma de Células Escamosas/genética , Diferenciação Celular/genética , Diferenciação Celular/efeitos da radiação , Divisão Celular/efeitos da radiação , Segregação de Cromossomos/efeitos da radiação , Imunofluorescência , Humanos , Raios Infravermelhos , Neoplasias Bucais/genética , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/efeitos da radiação , Fatores de Transcrição SOXB1/metabolismo , Células Tumorais CultivadasRESUMO
The equilibrium between proliferation and apoptosis is tightly balanced to maintain tissue homeostasis in normal tissues and even in tumors. Achieving and maintaining such a balance is important for cancer regrowth and spreading after cytotoxic treatments. Caspase-3 activation and tumor cell death following anticancer therapy as well as accompanying cell death pathways are well characterized, but their association to homeostasis of cancerous tissue and tumor progression remains poorly understood. Here we proposed a novel mechanism of cancer spreading induced by caspase-3. RhoGDIß, known as a direct cleavage substrate of caspase-3, is overexpressed in many epithelial cancers. The N-terminal-truncated RhoGDIß (ΔN-RhoGDIß) is accumulated in caspase-3-activated cells. Stable expression of ΔN-RhoGDIß in HeLa cells did not induce apoptosis, but impaired directional cell migration in a wound-healing assay accompanied by a perturbed direction of cell division at the wound edge. Subcellular protein fractionation experiments revealed that ΔN-RhoGDIß but not wild-type RhoGDIß was present in the detergent-soluble cytoplasmic and nuclear fractions and preferentially associated with Cdc42. Furthermore, Cdc42 activity was constitutively inhibited by stable expression of ΔN-RhoGDIß, resulting in increased radiation-induced compensatory proliferation linking to RhoA activation. Thus, ΔN-RhoGDIß dominant-negatively regulates Cdc42 activity and contributes to loss of polarity-related functions. The caspase-3-cleaved RhoGDIß is a possible determinant to promote cancer spreading due to deregulation of directional organization of tumor cell population and inhibition of default equilibrium between proliferation and apoptosis after cytotoxic damage. J. Cell. Physiol. 231: 2493-2505, 2016. © 2016 Wiley Periodicals, Inc.
Assuntos
Movimento Celular/efeitos da radiação , Polaridade Celular/efeitos da radiação , Neoplasias/patologia , Radiação , Inibidor beta de Dissociação do Nucleotídeo Guanina rho/metabolismo , Apoptose/efeitos da radiação , Caspase 3/metabolismo , Divisão Celular/efeitos da radiação , Proliferação de Células/efeitos da radiação , Sobrevivência Celular/efeitos da radiação , Regulação para Baixo/efeitos da radiação , Ativação Enzimática/efeitos da radiação , Genes Dominantes , Células HeLa , Humanos , Modelos Biológicos , Proteínas Mutantes/metabolismo , Metástase Neoplásica , Transporte Proteico/efeitos da radiação , Frações Subcelulares/metabolismo , Raios X , Proteína cdc42 de Ligação ao GTP/metabolismoRESUMO
Arabinogalactan proteins (AGPs) are highly glycosylated, hydroxyproline-rich proteins found at the cell surface of plants, where they play key roles in developmental processes. Brown algae are marine, multicellular, photosynthetic eukaryotes. They belong to the phylum Stramenopiles, which is unrelated to land plants and green algae (Chloroplastida). Brown algae share common evolutionary features with other multicellular organisms, including a carbohydrate-rich cell wall. They differ markedly from plants in their cell wall composition, and AGPs have not been reported in brown algae. Here we investigated the presence of chimeric AGP-like core proteins in this lineage. We report that the genome sequence of the brown algal model Ectocarpus siliculosus encodes AGP protein backbone motifs, in a gene context that differs considerably from what is known in land plants. We showed the occurrence of AGP glycan epitopes in a range of brown algal cell wall extracts. We demonstrated that these chimeric AGP-like core proteins are developmentally regulated in embryos of the order Fucales and showed that AGP loss of function seriously impairs the course of early embryogenesis. Our findings shine a new light on the role of AGPs in cell wall sensing and raise questions about the origin and evolution of AGPs in eukaryotes.
Assuntos
Epitopos/metabolismo , Fucus/crescimento & desenvolvimento , Fucus/genética , Mucoproteínas/metabolismo , Sequência de Aminoácidos , Divisão Celular/efeitos da radiação , Parede Celular/metabolismo , Parede Celular/efeitos da radiação , Fucus/efeitos da radiação , Genes de Plantas , Genoma , Indicadores e Reagentes , Luz , Modelos Biológicos , Mucoproteínas/química , Filogenia , Proteínas de Plantas/química , Proteínas de Plantas/metabolismo , Domínios Proteicos , Homologia de Sequência do Ácido Nucleico , Zigoto/metabolismoRESUMO
Because of their sessile lifestyle, plants are continuously exposed to solar UV-B radiation. Inhibition of leaf growth is one of the most consistent responses of plants upon exposure to UV-B radiation. In this work, we investigated the role of Growth-Regulating Factors (GRFs) and of microRNA miR396 in UV-B-mediated inhibition of leaf growth in Arabidopsis thaliana plants. We demonstrate that miRNA396 is upregulated by UV-B radiation in proliferating tissues and that this induction is correlated with a decrease in GRF1, GRF2, and GRF3 transcripts. Induction of miR396 results in inhibition of cell proliferation, and this outcome is independent of the UV-B photoreceptor UV resistance locus 8, as well as ATM AND RAD3-related and the mitogen-activated protein kinase MPK6, but is dependent on MPK3. Transgenic plants expressing an artificial target mimic directed against miR396 (MIM396) with a decrease in the endogenous microRNA activity or plants expressing miR396-resistant copies of several GRFs are less sensitive to this inhibition. Consequently, at intensities that can induce DNA damage in Arabidopsis plants, UV-B radiation limits leaf growth by inhibiting cell division in proliferating tissues, a process mediated by miR396 and GRFs.