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1.
Blood ; 122(7): 1271-83, 2013 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-23838347

RESUMO

Oncogenic transformation requires unlimited self-renewal. Currently, it remains unclear whether a normal capacity for self-renewal is required for acquiring an aberrant self-renewal capacity. Our results in a new conditional transgenic mouse showed that a mixed lineage leukemia (MLL) fusion oncogene, MLL-ENL, at an endogenous-like expression level led to leukemic transformation selectively in a restricted subpopulation of hematopoietic stem cells (HSCs) through upregulation of promyelocytic leukemia zinc finger (Plzf). Interestingly, forced expression of Plzf itself immortalized HSCs and myeloid progenitors in vitro without upregulation of Hoxa9/Meis1, which are well-known targets of MLL fusion proteins, whereas its mutant lacking the BTB/POZ domain did not. In contrast, depletion of Plzf suppressed the MLL-fusion-induced leukemic transformation of HSCs in vitro and in vivo. Gene expression analyses of human clinical samples showed that a subtype of PLZF-high MLL-rearranged myeloid leukemia cells was closely associated with the gene expression signature of HSCs. These findings suggested that MLL fusion protein enhances the self-renewal potential of normal HSCs to develop leukemia, in part through a Plzf-driven self-renewal program.


Assuntos
Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/patologia , Células-Tronco Hematopoéticas/patologia , Fatores de Transcrição Kruppel-Like/metabolismo , Leucemia/etiologia , Células Progenitoras Mieloides/patologia , Proteína de Leucina Linfoide-Mieloide/genética , Proteínas de Fusão Oncogênica/genética , Animais , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Western Blotting , Diferenciação Celular , Proliferação de Células , Transformação Celular Neoplásica/metabolismo , Citometria de Fluxo , Perfilação da Expressão Gênica , Células-Tronco Hematopoéticas/metabolismo , Humanos , Fatores de Transcrição Kruppel-Like/antagonistas & inibidores , Fatores de Transcrição Kruppel-Like/genética , Leucemia/metabolismo , Leucemia/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Células Progenitoras Mieloides/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , Proteína com Dedos de Zinco da Leucemia Promielocítica , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Retroviridae/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa
2.
Plant Cell Environ ; 37(5): 1086-96, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-24125071

RESUMO

Cadmium (Cd) accumulations in a Cd hyper-accumulator fern, Athyrium yokoscense (Ay), and tobacco, Nicotiana tabacum (Nt), were kinetically analysed using the positron-emitting tracer imaging system under two medium conditions (basal and no-nutrient). In Ay, maximumly 50% and 15% of the total Cd accumulated in the distal roots and the shoots under the basal condition, respectively. Interestingly, a portion of the Cd in the distal roots returned to the medium. In comparison with Ay, a little fewer Cd accumulations in the distal roots and clearly higher Cd migration to the shoots were observed in Nt under the basal condition (maximumly 40% and 70% of the total Cd, respectively). The no-nutrient condition down-regulated the Cd migration in both species, although the regulation was highly stricter in Ay than in Nt (almost no migration in Ay and around 20% migration in Nt). In addition, the present work enabled to estimate physical and physiological Cd accumulation capacities in the distal roots, and demonstrated condition-dependent changes especially in Ay. These results clearly suggested occurrences of species-/condition-specific regulations in each observed parts. It is probable that integration of these properties govern the specific Cd tolerance/accumulation in Ay and Nt.


Assuntos
Cádmio/metabolismo , Gleiquênias/metabolismo , Nicotiana/metabolismo , Autorradiografia , Elétrons , Imageamento Tridimensional , Cinética , Raízes de Plantas/metabolismo , Nicotiana/crescimento & desenvolvimento
3.
Int J Food Sci Nutr ; 65(2): 241-4, 2014 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-24144396

RESUMO

Xenobiotic-metabolizing enzymes (XMEs) play an important role in the elimination and detoxification of xenobiotics and drugs. A variety of natural dietary agents are known to protect against cancer by inducing XME. To elucidate the molecular mechanism of XME induction, we examined the effect of dietary eugenol (4-allyl-1-hydroxy-2-methoxybenzene) on xenobiotic metabolism. In this study, rats were administered dietary eugenol for 4 weeks to investigate the various effects of UDP-glucuronosyltransferase (UGT) and cytochrome P450 (CYP) expression. In rats administered dietary eugenol, expression levels of hepatic CYP1A 1 were reduced to 40% than of the controls, while expression of hepatic UGT1A6, UGT1A7 and UGT2B1 increased to 2-3 times than observed in the controls. Hepatic protein levels of UGT1A6 and 2B1 were also elevated in the eugenol-treated rats. These results suggest that the natural compound eugenol improves the xenobiotic-metabolizing systems that suppress and induce the expression of CYP1A1 and UGT, respectively.


Assuntos
Citocromo P-450 CYP1A1/metabolismo , Dieta , Eugenol/farmacologia , Glucuronosiltransferase/metabolismo , Fígado/metabolismo , Extratos Vegetais/farmacologia , Xenobióticos/metabolismo , Animais , Interações Medicamentosas , Eugenol/uso terapêutico , Glutationa Transferase/metabolismo , Masculino , Neoplasias/prevenção & controle , Fitoterapia , Ratos , Ratos Sprague-Dawley
4.
Microbiol Immunol ; 57(10): 684-91, 2013 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-23930694

RESUMO

To determine and compare the extent of contamination caused by antimicrobial-resistant lactic acid bacteria (LAB) in imported and domestic natural cheeses on the Japanese market, LAB were isolated using deMan, Rogosa and Sharpe (MRS) agar and MRS agar supplemented with six antimicrobials. From 38 imported and 24 Japanese cheeses, 409 LAB isolates were obtained and their antimicrobial resistance was tested. The percentage of LAB resistant to dihydrostreptomycin, erythromycin, and/or oxytetracycline isolated from imported cheeses (42.1%) was significantly higher than that of LAB resistant to dihydrostreptomycin or oxytetracycline from cheeses produced in Japan (16.7%; P=0.04). Antimicrobial resistance genes were detected in Enterococcus faecalis (tetL, tetM, and ermB; tetL and ermB; tetM) E. faecium (tetM), Lactococcus lactis (tetS), Lactobacillus (Lb.), casei/paracasei (tetM or tetW), and Lb. rhamnosus (ermB) isolated from seven imported cheeses. Moreover, these E. faecalis isolates were able to transfer antimicrobial resistance gene(s). Although antimicrobial resistance genes were not detected in any LAB isolates from Japanese cheeses, Lb. casei/paracasei and Lb. coryniformis isolates from a Japanese farm-made cheese were resistant to oxytetracycline (minimal inhibitory concentration [MIC], 32 µg/mL). Leuconostoc isolates from three Japanese farm-made cheeses were also resistant to dihydrostreptomycin (MIC, 32 to >512 µg/mL). In conclusion, the present study demonstrated contamination with antimicrobial-resistant LAB in imported and Japanese farm-made cheeses on the Japanese market, but not in Japanese commercial cheeses.


Assuntos
Antibacterianos/farmacologia , Queijo/microbiologia , Farmacorresistência Bacteriana , Lactobacillales/efeitos dos fármacos , Lactobacillales/isolamento & purificação , Técnicas Bacteriológicas , Sulfato de Di-Hidroestreptomicina/farmacologia , Eritromicina/farmacologia , Japão , Lactobacillales/genética , Testes de Sensibilidade Microbiana , Oxitetraciclina/farmacologia
5.
Evolution ; 77(10): 2234-2245, 2023 10 03.
Artigo em Inglês | MEDLINE | ID: mdl-37487052

RESUMO

Complexity in prezygotic mating behavior can contribute to the emergence of sexual incompatibility and reproductive isolation. In this study, we performed behavioral tests with two tidepool copepod species of the genus Tigriopus to explore the possibility of precopulatory behavioral isolation. We found that interspecific mating attempts failed prior to genital contact, and that this failure occurred at different behavioral steps between reciprocal pairings. Our results suggest that prezygotic barriers may exist at multiple points of the behavioral process on both male and female sides, possibly due to interspecific differences in mate-recognition cues used at those "checkpoints." While many copepod species are known to show unique precopulatory mate-guarding behavior, the potential contribution of prezygotic behavioral factors to their isolation is not widely recognized. The pattern of sequential mate-guarding behaviors may have allowed the diversification of precopulatory communication and contributed to the evolutionary diversity of the Tigriopus copepods.


Assuntos
Copépodes , Animais , Masculino , Feminino , Evolução Biológica , Comportamento Sexual Animal , Isolamento Reprodutivo , Reprodução
6.
Biol Pharm Bull ; 35(4): 634-8, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22466573

RESUMO

Two mechanisms have been proposed to explain quinone cytotoxicity: oxidative stress via the redox cycle, and the arylation of intracellular nucleophiles. The redox cycle is catalyzed by intracellular reductases, and therefore the toxicity of redox cycling quinone is considered to be closely associated with the reductase activity. This study examined the relationship between quinone toxicity and the intracellular reductase activity using 3 kinds of hepatic cells; rat primary hepatocytes, HepG2 and H4IIE. The intracellular reductase activity was; primary hepatocyte >>HepG2>H4IIE. The three kinds of cells showed almost the same vulnerability to an arylating quinone, 1,4-naphthoquinone (NQ). However, the susceptibility to a redox cycling quinone, 2,3-dimethoxy-1,4-naphthoquinone (DMNQ) was; primary hepatocyte>HepG2>H4IIE. In addition, the cytotoxicity elicited by DMNQ was significantly attenuated in HepG2 cells and almost completely suppressed in primary hepatocytes by diphenyleneiodonium chloride, a reductase inhibitor. These data suggest that cells with a high reductase activity are susceptible to redox cycling quinones. This study provides essential evidence to assess the toxicity of quinone-based drugs during their developmental processes.


Assuntos
Hepatócitos/efeitos dos fármacos , Oxirredutases/metabolismo , Quinonas/toxicidade , Animais , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Células Hep G2 , Humanos , Masculino , Oxirredução , Ratos , Ratos Wistar
7.
BMC Plant Biol ; 11: 172, 2011 Nov 29.
Artigo em Inglês | MEDLINE | ID: mdl-22123026

RESUMO

BACKGROUND: Rice is a major source of dietary intake of cadmium (Cd) for populations that consume rice as a staple food. Understanding how Cd is transported into grains through the whole plant body is necessary for reducing rice Cd concentrations to the lowest levels possible, to reduce the associated health risks. In this study, we have visualized and quantitatively analysed the real-time Cd dynamics from roots to grains in typical rice cultivars that differed in grain Cd concentrations. We used positron-emitting 107Cd tracer and an innovative imaging technique, the positron-emitting tracer imaging system (PETIS). In particular, a new method for direct and real-time visualization of the Cd uptake by the roots in the culture was first realized in this work. RESULTS: Imaging and quantitative analyses revealed the different patterns in time-varying curves of Cd amounts in the roots of rice cultivars tested. Three low-Cd accumulating cultivars (japonica type) showed rapid saturation curves, whereas three high-Cd accumulating cultivars (indica type) were characterized by curves with a peak within 30 min after 107Cd supplementation, and a subsequent steep decrease resulting in maintenance of lower Cd concentrations in their roots. This difference in Cd dynamics may be attributable to OsHMA3 transporter protein, which was recently shown to be involved in Cd storage in root vacuoles and not functional in the high-Cd accumulating cultivars. Moreover, the PETIS analyses revealed that the high-Cd accumulating cultivars were characterized by rapid and abundant Cd transfer to the shoots from the roots, a faster transport velocity of Cd to the panicles, and Cd accumulation at high levels in their panicles, passing through the nodal portions of the stems where the highest Cd intensities were observed. CONCLUSIONS: This is the first successful visualization and quantification of the differences in whole-body Cd transport from the roots to the grains of intact plants within rice cultivars that differ in grain Cd concentrations, by using PETIS, a real-time imaging method.


Assuntos
Cádmio/metabolismo , Imageamento Tridimensional , Oryza/metabolismo , Raízes de Plantas/metabolismo , Transporte Biológico , Radioisótopos de Cádmio/análise , Brotos de Planta/metabolismo , Tomografia por Emissão de Pósitrons
8.
J Appl Toxicol ; 31(2): 173-8, 2011 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-20803752

RESUMO

Quinone toxicity is induced by two principal mechanisms: arylation/alkylation and a redox cycle. We have previously shown that increases in intracellular levels of superoxide anion and cell death induced by 2,3-dimethoxy-1,4-naphthoquinone (DMNQ), a redox cycling quinone, are enhanced by pretreatment of rat primary hepatocytes with cytochrome P450 inhibitors. This indicates a novel interaction of quinones with cytochrome P450, and is thus worthy of further investigation using an in vivo model. The aim of this study was to examine the effects of cytochrome P450 inhibitors on DMNQ-induced hepatotoxicity in rats. When DMNQ was administered intraperitoneally, the activities of serum alanine aminotransferase and aspartate aminotransferase were found to increase in a dose-dependent manner, indicating that hepatotoxicity was induced by treatment with DMNQ. Pretreatment with the cytochrome P450 inhibitors SKF-525A (SKF), cimetidine and ketoconazole potentiated the DMNQ-induced hepatotoxicity. The blood concentration of DMNQ was not affected by administration of SKF. Pretreatment with the antioxidant α-tocopherol almost completely attenuated the hepatotoxicity induced by DMNQ and by the combination of DMNQ with SKF. Levels of reduced glutathione in the liver were decreased and levels of oxidized glutathione were increased by treatment with DMNQ. These effects were potentiated by pretreatment with SKF. DMNQ-induced lipid peroxidation in the liver was also enhanced by pretreatment with SKF. Taken together, these results indicate that DMNQ-induced hepatotoxicity is augmented by inhibition of cytochrome P450 and that this augmentation is due to the enhancement of oxidative stress.


Assuntos
Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Inibidores das Enzimas do Citocromo P-450 , Fígado/efeitos dos fármacos , Naftoquinonas/toxicidade , Animais , Antioxidantes/uso terapêutico , Doença Hepática Induzida por Substâncias e Drogas/sangue , Doença Hepática Induzida por Substâncias e Drogas/tratamento farmacológico , Cimetidina/farmacologia , Relação Dose-Resposta a Droga , Interações Medicamentosas , Inibidores Enzimáticos/farmacologia , Inibidores Enzimáticos/uso terapêutico , Glutationa/metabolismo , Cetoconazol/farmacologia , Peroxidação de Lipídeos/efeitos dos fármacos , Fígado/metabolismo , Masculino , Naftoquinonas/administração & dosagem , Naftoquinonas/sangue , Naftoquinonas/farmacocinética , Oxirredução , Estresse Oxidativo , Proadifeno/farmacologia , Proadifeno/uso terapêutico , Ratos , Ratos Wistar , Ciclização de Substratos/efeitos dos fármacos , alfa-Tocoferol/uso terapêutico
9.
PLoS One ; 16(3): e0248425, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33705482

RESUMO

The Ten Eleven Translocation 1 (TET1) gene encodes an epigenetic modifying molecule that is involved in demethylation of 5-methylcytosine. In hematological malignancies, loss-of-function mutations of TET2, which is one of the TET family genes including TET1, are frequently found, while the mutations of TET1 are not. However, clinical studies have revealed that TET1 is highly expressed in some cases of the hematological malignancies including acute myeloid leukemia. Indeed, studies by mouse models using conventional Tet1 knockout mice demonstrated that Tet1 is involved in myeloid leukemogenesis by Mixed Lineage Leukemia (MLL) fusion gene or TET2 mutant. Meanwhile, the other study showed that Tet1 is highly expressed in hematopoietic stem cells (HSCs), and that deletion of Tet1 in HSCs enhances potential self-renewal capacity, which is potentially associated with myeloid leukemogenesis. To examine the role of Tet1 in myeloid leukemogenesis more precisely, we generated novel conditional Tet1-knockout mice, which were used to generate the compound mutant mice by crossing with the inducible MLL-ENL transgenic mice that we developed previously. The leukemic immortalization in vitro was not critically affected by conditional ablation of Tet1 in HSCs with the induced expression of MLL-ENL or in hematopoietic progenitor cells retrovirally transduced with MLL-ENL. In addition, the leukemic phenotypes caused by the induced expression of MLL-ENL in vivo was not also critically affected in the compound mutant mouse model by conditional ablation of Tet1, although we found that the expression of Evi1, which is one of critical target genes of MLL fusion gene, in tumor cells was remarkably low under Tet1-ablated condition. These results revealed that Tet1 was dispensable for the myeloid leukemogenesis by MLL-ENL, suggesting that the therapeutic application of Tet1 inhibition may need careful assessment.


Assuntos
Carcinogênese , Proteínas de Ligação a DNA , Regulação Leucêmica da Expressão Gênica , Histona-Lisina N-Metiltransferase , Leucemia Mieloide , Proteína de Leucina Linfoide-Mieloide , Neoplasias Experimentais , Proteínas de Fusão Oncogênica , Proteínas Proto-Oncogênicas , Fatores de Transcrição , Animais , Carcinogênese/genética , Carcinogênese/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Células-Tronco Hematopoéticas/metabolismo , Histona-Lisina N-Metiltransferase/genética , Histona-Lisina N-Metiltransferase/metabolismo , Leucemia Mieloide/genética , Leucemia Mieloide/metabolismo , Camundongos , Camundongos Transgênicos , Proteína de Leucina Linfoide-Mieloide/genética , Proteína de Leucina Linfoide-Mieloide/metabolismo , Neoplasias Experimentais/genética , Neoplasias Experimentais/metabolismo , Proteínas de Fusão Oncogênica/genética , Proteínas de Fusão Oncogênica/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
10.
Plant Sci ; 305: 110822, 2021 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-33691958

RESUMO

Glutathione (GSH) is a tripeptide involved in controlling heavy metal movement in plants. Our previous study showed that GSH, when site-specifically applied to plant roots, inhibits Cd translocation from the roots to shoots in hydroponically cultured oilseed rape (Brassica napus) plants. A factor that led to this inhibitory effect was the activation of Cd efflux from root cells. To further investigate the molecular mechanism triggered by root-applied GSH, Cd movement was non-invasively monitored using a positron-emitting tracer imaging system. The Cd absorption and efflux process in the roots were visualized successfully. The effects of GSH on Cd efflux from root cells were estimated by analyzing imaging data. Reanalysis of image data suggested that GSH applied to roots, at the shoot base, activated Cd return. Cutting the shoot base significantly inhibited Cd efflux from root cells. These experimental results demonstrate that the shoot base plays an important role in distributing Cd throughout the plant body. Furthermore, microarray analysis revealed that about 400 genes in the roots responded to root-applied GSH. Among these, there were genes for transporter proteins related to heavy metal movement in plants and proteins involved in the structure modification of cell walls.


Assuntos
Transporte Biológico/fisiologia , Brassica napus/metabolismo , Cádmio/metabolismo , Glutationa/metabolismo , Metais Pesados/metabolismo , Raízes de Plantas/metabolismo , Brotos de Planta/metabolismo , Produtos Agrícolas/metabolismo
11.
Plant Sci ; 290: 110304, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31779894

RESUMO

Glutathione (GSH) is a thiol-containing compound involved in many aspects of plant metabolism. In the present study, we investigated how enhancing endogenous and exogenous GSH affects cadmium (Cd) movement and distribution in Arabidopsis plants cultured hydroponically. Transgenic Arabidopsis plants with a strong ability to synthesize GSH in roots were generated by transforming the gene encoding the bifunctional γ-glutamylcysteine synthetase-glutathione synthetase enzyme from Streptococcus thermophiles (StGCS-GS). Enhancing endogenous and exogenous GSH decreased the Cd translocation ratio in different ways. Only exogenous GSH significantly inhibited Cd translocation from roots to shoots in wild-type and transgenic Arabidopsis plants. Our study demonstrated that GSH mainly functions outside root cells to inhibit Cd translocation from roots to shoots.


Assuntos
Arabidopsis/metabolismo , Cádmio/metabolismo , Glutationa/metabolismo , Plantas Geneticamente Modificadas/metabolismo , Poluentes do Solo/metabolismo , Arabidopsis/efeitos dos fármacos , Transporte Biológico , Glutationa/farmacologia , Hidroponia , Raízes de Plantas/metabolismo , Plantas Geneticamente Modificadas/efeitos dos fármacos , Plantas Geneticamente Modificadas/genética
12.
Appl Radiat Isot ; 151: 7-12, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31151049

RESUMO

Accurate analysis of N fixation in leguminous crops requires determination of N utilization within an intact plant; however, most approaches require tissue disassembly. We developed a simple and rapid technique to generate high-purity and high-yield [13N]N2 gas and obtained real-time images of N fixation in an intact soybean plant. The purification efficiency was ∼81.6% after decay correction. Our method provides accurate signals of N fixation and allows free changes to the tracer gas composition to suit different experimental designs.


Assuntos
Produtos Agrícolas/metabolismo , Glycine max/metabolismo , Fixação de Nitrogênio , Radioisótopos de Nitrogênio/isolamento & purificação , Nitrogênio/metabolismo , Transporte Biológico , Cromatografia Gasosa
13.
Plant Sci ; 283: 416-423, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31128713

RESUMO

Glutathione (GSH) is a vital compound involved in several plant metabolic pathways. Our previous study indicated that foliar GSH application can increase zinc (Zn) levels in leafy vegetables. The objective of this study was to determine the mode of action of GSH as it relates to Zn transport from roots to shoots. Two types of transgenic Arabidopsis plants with genes for GSH synthesis, including StGCS-GS or AtGSH1 driven by the leaf-specific promoter of chlorophyll a/b-binding protein (pCab3) gene were generated. Both types of transgenic Arabidopsis plants showed significant increases in shoot GSH concentrations compared to the wild type (WT). Monitoring 65Zn movement by positron-emitting tracer imaging system (PETIS) analysis indicated that the 65Zn amount in the shoots of both types of transgenic Arabidopsis plants were higher than that in the WT. GSH concentration in phloem sap was increased significantly in WT with foliar applications of 10 mM GSH (WT-GSH), but not in transgenic Arabidopsis with elevated foliar GSH synthesis. Both types of transgenic Arabidopsis with elevated foliar GSH synthesis and WT-GSH exhibited increased shoot Zn concentrations and Zn translocation ratios. These results suggest that enhancement of endogenous foliar GSH synthesis and exogenous foliar GSH application affect root-to-shoot transport of Zn.


Assuntos
Arabidopsis/metabolismo , Glutationa/metabolismo , Folhas de Planta/metabolismo , Raízes de Plantas/metabolismo , Brotos de Planta/metabolismo , Zinco/metabolismo , Arabidopsis/genética , Transporte Biológico , Genes de Plantas/genética , Floema/metabolismo , Plantas Geneticamente Modificadas , Reação em Cadeia da Polimerase em Tempo Real
14.
FEBS J ; 275(9): 2032-41, 2008 May.
Artigo em Inglês | MEDLINE | ID: mdl-18355319

RESUMO

In eukaryotes, meiosis leads to genetically variable gametes through recombination between homologous chromosomes of maternal and paternal origin. Chromatin organization following meiotic recombination is critical to ensure the correct segregation of homologous chromosomes into gametes. However, the mechanism of chromatin organization after meiotic recombination is unknown. In this study we report that the meiosis-specific recombinase Lim15/Dmc1 interacts with the homologue of the largest subunit of chromatin assembly factor 1 (CAF-1) in the basidiomycete Coprinopsis cinerea (Coprinus cinereus). Using C. cinerea LIM15/DMC1 (CcLIM15) as the bait in a yeast two-hybrid screen, we have isolated the C. cinerea homologue of Cac1, the largest subunit of CAF-1 in Saccharomyces cerevisiae, and named it C. cinerea Cac1-like (CcCac1L). Two-hybrid assays confirmed that CcCac1L binds CcLim15 in vivo. beta-Galactosidase assays revealed that the N-terminus of CcCac1L preferentially interacts with CcLim15. Co-immunoprecipitation experiments showed that these proteins also interact in the crude extract of meiotic cells. Furthermore, we demonstrate that, during meiosis, CcCac1L interacts with proliferating cell nuclear antigen (PCNA), a component of the DNA synthesis machinery recently reported as an interacting partner of Lim15/Dmc1. Taken together, these results suggest a novel role of the CAF-1-PCNA complex in meiotic events. We propose that the CAF-1-PCNA complex modulates chromatin assembly following meiotic recombination.


Assuntos
Proteínas de Ciclo Celular/metabolismo , Montagem e Desmontagem da Cromatina , Proteínas Cromossômicas não Histona/metabolismo , Proteínas de Ligação a DNA/metabolismo , Meiose , Recombinação Genética , Fator 1 de Modelagem da Cromatina , Proteínas Cromossômicas não Histona/química , Proteínas Cromossômicas não Histona/genética , Coprinus/enzimologia , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/genética , Humanos , Modelos Biológicos , Antígeno Nuclear de Célula em Proliferação/genética , Antígeno Nuclear de Célula em Proliferação/metabolismo , Subunidades Proteicas/química , Ressonância de Plasmônio de Superfície , Técnicas do Sistema de Duplo-Híbrido
15.
J Vis Exp ; (139)2018 09 26.
Artigo em Inglês | MEDLINE | ID: mdl-30320760

RESUMO

Copepods of the genus Tigriopus, which are common zooplankton in rocky tide pools, show precopulatory mate-guarding behavior where a male clasps a potential mate to form a pair. While this phenomenon has attracted interest of researchers, methods for its analysis have not been well described. Here we describe procedures for: 1) individual culturing and staging of Tigriopus juveniles and adults, and 2) video-based analysis of their mate-guarding behavior. The culturing method enables experimental control of paring experience of animals as well as the ability to track their development before behavioral tests. The analysis method allows quantitative evaluation of several aspects of the mate-guarding behavior, including capturing attempts by males and swimming trajectory of mate-guarding pairs. Although these methods were originally established for ethological studies on Tigriopus, with proper modifications they can also be applied to studies of other zooplankton in different research fields, such as physiology, toxicology, and ecological genetics.


Assuntos
Copépodes/fisiologia , Comportamento Sexual Animal/fisiologia , Criação de Animais Domésticos , Animais , Feminino , Masculino , Reprodução/fisiologia
16.
Anim Sci J ; 89(1): 219-226, 2018 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-28833797

RESUMO

Airag, (Fermented horse milk) is a traditional milk product in Mongolia. Herders separate foals from their dams and tie them at a milking site during the daytime to produce airag. To evaluate the effects of horse management on the movement of dams, we tracked three dams in a herd in camp 1 during summer and camp 2 during autumn of 2013 and analyzed their movements during the milking (daytime) and non-milking (nighttime) periods in an area famous for its high-quality airag. Dams were gathered every 1.7 ± 0.0 h between 07.46 and 15.47 hours at the milking sites and milked 4.6 ± 0.2 times/day during the study period (86 days). Daily cumulative and maximum linear distances from the milking sites were longer (P < 0.01) during the non-milking period than during the milking period. Daily home ranges were 91 and 26 times greater during the non-milking period (P < 0.001) in camps 1 and 2, respectively. The greater range during the non-milking period would reflect the spatial distributions of water, salt and forage. The dams initially used similar areas and gradually shifted their daily home ranges after several days. This shift suggests that the dams grazed farther afield as forage availability declined around the milking site. For better airag production and sustainable pasture use, our results provide insights useful for evaluating the effects of milking management on vegetation and soil in those pastures, for selecting the appropriate milking times and frequency, and for choosing the right timing to shift milking sites.


Assuntos
Indústria de Laticínios/métodos , Herbivoria/fisiologia , Cavalos/fisiologia , Cavalos/psicologia , Lactação/fisiologia , Locomoção/fisiologia , Animais , Feminino , Sistemas de Informação Geográfica/instrumentação , Mongólia , Estações do Ano , Fatores de Tempo
17.
Front Plant Sci ; 9: 1946, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30687351

RESUMO

In protected strawberry (Fragaria × ananassa Duch.) cultivation, environmental control based on the process of photosynthate translocation is essential for optimizing fruit quality and yield, because the process of photosynthate translocation directly affects dry matter partitioning. We visualized photosynthate translocation to strawberry fruits non-invasively with 11CO2 and a positron-emitting tracer imaging system (PETIS). We used PETIS to evaluate real-time dynamics of 11C-labeled photosynthate translocation from a 11CO2-fed leaf, which was immediately below the inflorescence, to individual fruits on an inflorescence in intact plant. Serial photosynthate translocation images and animations obtained by PETIS verified that the 11C-photosynthates from the source leaf reached the sink fruit within 1 h but did not accumulate homogeneously within a fruit. The quantity of photosynthate translocation as represented by 11C radioactivity varied among individual fruits and their positions on the inflorescence. Photosynthate translocation rates to secondary fruit were faster than those to primary or tertiary fruits, even though the translocation pathway from leaf to fruit was the longest for the secondary fruit. Moreover, the secondary fruit was 25% smaller than the primary fruit. Sink activity (11C radioactivity/dry weight [DW]) of the secondary fruit was higher than those of the primary and tertiary fruits. These relative differences in sink activity levels among the three fruit positions were also confirmed by 13C tracer measurement. Photosynthate translocation rates in the pedicels might be dependent on the sink strength of the adjoining fruits. The present study established 11C-photosynthate arrival times to the sink fruits and demonstrated that the translocated material does not uniformly accumulate within a fruit. The actual quantities of translocated photosynthates from a specific leaf differed among individual fruits on the same inflorescence. To the best of our knowledge, this is the first reported observation of real-time translocation to individual fruits in an intact strawberry plant using 11C-radioactive- and 13C-stable-isotope analyses.

18.
Nucleic Acids Res ; 33(18): 5809-18, 2005.
Artigo em Inglês | MEDLINE | ID: mdl-16221977

RESUMO

Lim15/Dmc1 is a meiosis specific RecA-like protein. Here we propose its participation in meiotic chromosome pairing-related events along with DNA topoisomerase II. Analysis of protein-protein interactions using in vitro binding assays provided evidence that Coprinus cinereus DNA topoisomerase II (CcTopII) specifically interacts with C.cinereus Lim15/Dmc1 (CcLim15). Co-immunoprecipitation experiments also indicated that the CcLim15 protein interacts with CcTopII in vivo. Furthermore, a significant proportion of CcLim15 and CcTopII could be shown to co-localize on chromosomes from the leptotene to the zygotene stage. Interestingly, CcLim15 can potently activate the relaxation/catenation activity of CcTopII in vitro, and CcTopII suppresses CcLim15-dependent strand transfer activity. On the other hand, while enhancement of CcLim15's DNA-dependent ATPase activity by CcTopII was found in vitro, the same enzyme activity of CcTopII was inhibited by adding CcLim15. The interaction of CcLim15 and CcTopII may facilitate pairing of homologous chromosomes.


Assuntos
Proteínas de Ciclo Celular/metabolismo , DNA Topoisomerases Tipo II/metabolismo , Proteínas de Ligação a DNA/metabolismo , Meiose , Sequência de Bases , Proteínas de Ciclo Celular/análise , Proteínas de Ciclo Celular/química , Núcleo Celular/química , Núcleo Celular/enzimologia , Coprinus/enzimologia , DNA Topoisomerases Tipo II/análise , DNA Topoisomerases Tipo II/química , Proteínas de Ligação a DNA/análise , Proteínas de Ligação a DNA/química , Imunoprecipitação , Dados de Sequência Molecular , Deleção de Sequência , Técnicas do Sistema de Duplo-Híbrido
19.
Mol Cell Biol ; 37(13)2017 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-28416638

RESUMO

PLZF is a transcription factor that confers aberrant self-renewal in leukemogenesis, and the PLZF-RARA fusion gene causes acute promyelocytic leukemia (APL) through differentiation block. However, the molecular mechanisms of aberrant self-renewal underlying PLZF-mediated leukemogenesis are poorly understood. To investigate these mechanisms, comprehensive expression profiling of mouse hematopoietic stem/progenitor cells transduced with Plzf was performed, which revealed the involvement of a key transcriptional coactivator, Eya2, a target molecule shared by Plzf and PLZF-RARA, in the aberrant self-renewal. Indeed, PLZF-RARA as well as Plzf rendered those cells immortalized through upregulation of Eya2. Eya2 also led to immortalization without differentiation block, while depletion of Eya2 suppressed clonogenicity in cells immortalized by PLZF-RARA without influence on differentiation and apoptosis. Interestingly, cancer outlier profile analysis of human samples of acute myeloid leukemia (AML) in The Cancer Genome Atlas (TCGA) revealed a subtype of AML that strongly expressed EYA2 In addition, gene set enrichment analysis of human AML samples, including TCGA data, showed that this subtype of AML was more closely associated with the properties of leukemic stem cells in its gene expression signature than other AMLs. Therefore, EYA2 may be a target for molecular therapy in this subtype of AML, including PLZF-RARA APL.


Assuntos
Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/patologia , Regulação Neoplásica da Expressão Gênica , Células-Tronco Hematopoéticas/patologia , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteínas Nucleares/genética , Proteínas de Fusão Oncogênica/metabolismo , Proteínas Tirosina Fosfatases/genética , Animais , Diferenciação Celular , Transformação Celular Neoplásica/metabolismo , Células Cultivadas , Células-Tronco Hematopoéticas/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Leucemia Mieloide Aguda/patologia , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Nucleares/metabolismo , Proteínas de Fusão Oncogênica/genética , Regiões Promotoras Genéticas , Proteínas Tirosina Fosfatases/metabolismo
20.
Plant Methods ; 13: 40, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28533812

RESUMO

BACKGROUND: Positron imaging can be used to non-destructively visualize the dynamics of a positron-emitting radionuclide in vivo, and is therefore a tool for understanding the mechanisms of nutrient transport in intact plants. The transport of zinc, which is one of the most important nutrient elements for plants, has so far been visualized by positron imaging using 62Zn (half-life: 9.2 h), which is manufactured in the limited number of facilities that have a cyclotron. In contrast, the positron-emitting radionuclide 65Zn (half-life: 244 days) is commercially available worldwide. In this study, we examined the possibility of conducting positron imaging of zinc in intact plants using 65Zn. RESULTS: By administering 65Zn and imaging over a long time, clear serial images of 65Zn distributions from the root to the panicle of dwarf rice plants were successfully obtained. CONCLUSIONS: Non-destructive visualization of zinc dynamics in plants was achieved using commercially available 65Zn and a positron imaging system, demonstrating that zinc dynamics can be visualized even in facilities without a cyclotron.

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