RESUMO
The role of autophagy in pulmonary microvascular endothelial cells (PMVECs) is controversial in LPS-induced acute lung injury (ALI). Mixed lineage kinase domain-like pseudokinase (MLKL) has recently been reported to maintain cell survival by facilitating autophagic flux in response to starvation rather than its well-recognized role in necroptosis. Using a mouse PMVEC and LPS-induced ALI model, we showed that in PMVECs, MLKL was phosphorylated (p-MLKL) and autophagic flux was accelerated at the early stage of LPS stimulation (1-3 h), manifested by increases in concentrations of lipidated MAP1LC3B/LC3B (microtubule-associated protein 1 light chain 3 ß; LC3-II), decreases in concentrations of SQSTM1/p62 (sequestosome 1), and fusion of the autophagosome and lysosome by pHluorin-mKate2-human LC3 assay, which were all reversed by either MLKL inhibitor or siRNA MLKL. In mice, the inhibition of MLKL increased vascular permeability and aggravated mouse ALI upon 3-hour LPS stimulation. The p-MLKL induced by short-term LPS formed multimers to facilitate the closure of the phagophore by HaloTag-LC3 autophagosome completion assay. The charged multivesicular body protein 2A (CHMP2A) is essential in the process of phagophore closure into the nascent autophagosome. In agreement with the p-MLKL change, CHMP2A concentrations markedly increased during 1-3-hour LPS stimulation. CHMP2A knockdown blocked autophagic flux upon LPS stimulation, whereas CHMP2A overexpression boosted autophagic flux and attenuated mouse ALI even in the presence of MLKL inhibitor. We propose that the activated MLKL induced by short-term LPS facilitates autophagic flux by accelerating the closure of the phagophore via CHMP2A, thus protecting PMVECs and alleviating LPS-induced ALI.
Assuntos
Lesão Pulmonar Aguda , Células Endoteliais , Humanos , Lesão Pulmonar Aguda/metabolismo , Autofagia/genética , Proteínas de Transporte/metabolismo , Células Endoteliais/metabolismo , Lipopolissacarídeos , Pulmão/metabolismo , Proteínas Quinases/genéticaRESUMO
MAIN CONCLUSION: The rice SnRK2 members SAPK4, SAPK5, SAPK7 and SAPK10 are positive regulators involved in the regulation of rice flowering, while other single mutants exhibited no effect on rice flowering. The rice SnRK2 family, comprising 10 members known as SAPK (SnRK2-Associated Protein Kinase), is pivotal in the abscisic acid (ABA) pathway and crucial for various biological processes, such as drought resistance and salt tolerance. Additionally, these members have been implicated in the regulation of rice heading date, a key trait influencing planting area and yield. In this study, we utilized gene editing technology to create mutants in the Songjing 2 (SJ2) background, enabling a comprehensive analyze the role of each SAPK member in rice flowering. We found that SAPK1, SAPK2, and SAPK3 may not directly participate in the regulatory network of rice heading date, while SAPK4, SAPK5, and SAPK7 play positive roles in rice flowering regulation. Notably, polygene deletion resulted in an additive effect on delaying flowering. Our findings corroborate the previous studies indicating the positive regulatory role of SAPK10 in rice flowering, as evidenced by delayed flowering observed in sapk9/10 double mutants. Moving forward, our future research will focus on analyzing the molecular mechanisms underlying SAPKs involvement in rice flowering regulation, aiming to enhance our understanding of the rice heading date relationship network and lay a theoretical foundation for breeding efforts to alter rice ripening dates.
Assuntos
Flores , Regulação da Expressão Gênica de Plantas , Oryza , Proteínas de Plantas , Oryza/genética , Oryza/crescimento & desenvolvimento , Oryza/fisiologia , Oryza/enzimologia , Flores/genética , Flores/crescimento & desenvolvimento , Flores/fisiologia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Mutação , Edição de Genes , Estresse Fisiológico/genética , Proteínas Quinases/genética , Proteínas Quinases/metabolismo , Ácido Abscísico/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismoRESUMO
Rice flowering time determines its geographical distribution and yield traits. As a short-day plant, rice can grow in the northern long-day conditions due to the functional mutations of many photosensitive genes. In this study, to identify novel genes or alleles that regulate flowering time in high latitude region, two cultivar, Dongnong 413 (DN413) and Yukimochi (XN) showing extreme early flowering were used for investigation. DN413 is around 4.0 days earlier than XN, and both cultivars can be grown in II (2500 â-2700 â) to III (2300 â-2500 â) accumulated temperature zones. We found that the two cultivars shared the same genotype of heading date genes, including Hd1/2/4/5/6/16/17/18, Ehd2, DTH2, SE5, Hd3a. Importantly, a novel Ehd3 allele characterized by a A1146C substitution was identified, which results in the E382D substitution, hereafter the 382 position E is defined as Hap_E and the 382 position D is defined as Hap_D. Association analysis showed that Hap_E is earlier flowering than Hap_D. Subsequently, we construct DN413 Hap_D line by three times back-crossing DN413 with XN, and found the heading date of DN413 Hap_D was 1.7-3.5 days later than DN413. Moreover, Hap_E and Hap_D of Ehd3 were transformed into ehd3 mutant, respectively, and the Ehd3pro:Ehd3D/ehd3 flowered later than that Ehd3pro:Ehd3E/ehd3 by around 4.3 days. Furthermore, we showed Ehd3 functions as a transcriptional suppressor and the substitution of Asp-382 lost the inhibition activity in protoplasts. Finally, a CAPS marker was developed and used for genotyping and marker assistant breeding. Collectively, we discovered a novel functional allele of Ehd3, which can used as a valuable breeding target. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-024-01472-x.
RESUMO
Panicle exsertion is one of the crucial agronomic traits in rice (Oryza sativa). Shortening of panicle exsertion often leads to panicle enclosure and severely reduces seed production. Gibberellin (GA) plays important roles in regulating panicle exsertion. However, the underlying mechanism and the relative regulatory network remain elusive. Here, we characterized the oswrky78 mutant showing severe panicle enclosure, and found that the defect of oswrky78 is caused by decreased bioactive GA contents. Biochemical analysis demonstrates that OsWRKY78 can directly activate GA biosynthesis and indirectly suppress GA metabolism. Moreover, we found OsWRKY78 can interact with and be phosphorylated by mitogen-activated protein kinase (MAPK) kinase OsMAPK6, and this phosphorylation can enhance OsWRKY78 stability and is necessary for its biological function. Taken together, these results not only reveal the critical function of OsWRKY78, but also reveal its mechanism via mediating crosstalk between MAPK and the GA signaling pathway in regulating panicle exsertion.
Assuntos
Giberelinas , Oryza , Giberelinas/metabolismo , Oryza/genética , Oryza/metabolismo , Transdução de Sinais/genética , Sementes/metabolismo , FenótipoRESUMO
Primordial germ cell 7 (PGC7)(Dppa3 or Stella) is a small inherently disordered protein that is mainly expressed in oocytes and plays a vital role in the regulation of DNA methylation reprogramming in imprinted loci through interaction with other proteins. Most of PGC7-deficient zygotes are blocked at two-cell stage with an increased tri-methylation at lysine 27 of histone H3 (H3K27me3) level in the nucleus. Our previous work has indicated that PGC7 interacts with yin-yang1 (YY1) that is essential for the recruitment of enhancer of zeste homolog 2 (EZH2)-containing Polycomb repressive complex 2 (PRC2) to H3K27me3 modification sites. Here, we found that the presence of PGC7 weakened the interaction between YY1 and PRC2 without disrupting the assembly of core subunits of the PRC2 complex. In addition, PGC7 promoted AKT to phosphorylate serine 21 of EZH2, resulting in inhibition of EZH2 activity and the dissociation of EZH2 from YY1, thereby decreasing H3K27me3 level. In zygotes, the PGC7-deficient and AKT inhibitor MK2206 both promoted EZH2 to enter the pronuclei but without disturbing the subcellular localization of YY1 and caused an increase in the level of H3K27me3 in the pronuclei, as well as inhibition of the expression of zygote-activating genes regulated by H3K27me3 in two-cell embryos. In summary, PGC7 could affect zygotic genome activation during early embryonic development by regulating the level of H3K27me3 through regulation of PRC2 recruitment, EZH2 activity, and subcellular localization.NEW & NOTEWORTHY PGC7 and YY1 interaction inhibits recruitment of PRC2 by YY1. PGC7 promotes AKT and EZH2 interaction to increase pEZH2-S21 level, which weakens YY1 and EZH2 interaction, thereby decreasing H3K27me3 level. In zygotes, the PGC7-deficient and AKT inhibitor MK2206 promote EZH2 to enter the pronuclei, and increase H3K27me3 level in the pronuclei, as well as inhibition of the expression of zygote-activating genes regulated by H3K27me3 in two-cell embryos, which ultimately affects early embryo development.
Assuntos
Histonas , Complexo Repressor Polycomb 2 , Complexo Repressor Polycomb 2/genética , Complexo Repressor Polycomb 2/metabolismo , Histonas/genética , Histonas/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Metilação de DNA , Células Germinativas/metabolismoRESUMO
Soybean (Glycine max) is highly sensitive to photoperiod, which affects flowering time and plant architecture and thus limits the distribution range of elite soybean cultivars. The major maturity gene E1 confers the most prominent effect on photoperiod sensitivity, but its downstream signaling pathway remains largely unknown. Here, we confirm that the encoded E1 protein is a transcriptional repressor. The expression of seven GmMDE genes (Glycine max MADS-box genes downregulated by E1) was suppressed when E1 was overexpressed and promoted when E1 was knocked out through clustered regularly-interspaced short palindromic repeats (CRISPR)/CRISPR associated protein 9 (Cas9)-mediated mutagenesis. These GmMDEs exhibited similar tissue specificity and expression patterns, including in response to photoperiod, E1 expression, and E1 genotype. E1 repressed GmMDE promoter activity. Results for two GmMDEs showed that E1 epigenetically silences their expression by directly binding to their promoters to increase H3K27me3 levels. The overexpression of GmMDE06 promoted flowering and post-flowering termination of stem growth. The late flowering phenotype of E1-overexpressing soybean lines was reversed by the overexpression of GmMDE06, placing GmMDE06 downstream of E1. The overexpression of GmMDE06 increased the expression of the soybean FLOWERING LOCUS T orthologs GmFT2a and GmFT5a, leading to feedback upregulation of GmMDE, indicating that GmMDE and GmFT2a/GmFT5a form a positive regulatory feedback loop promoting flowering. GmMDE06 also promoted post-flowering termination of stem growth by repressing the expression of the shoot identity gene Dt1. The E1-GmMDEs-GmFT2a/5a-Dt1 signaling pathway illustrates how soybean responds to photoperiod by modulating flowering time and post-flowering stem termination.
Assuntos
Glycine max , Fotoperíodo , Florígeno/metabolismo , Flores/fisiologia , Regulação da Expressão Gênica de Plantas , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Glycine max/metabolismoRESUMO
Rice is a chilling-sensitive plant, and extremely low temperatures seriously decrease rice production. Several genes involved in chilling stress have been reported in rice; however, the chilling signaling in rice remains largely unknown. Here, we investigated the chilling tolerance phenotype of overexpression of constitutive active OsMAPK6 (CAMAPK6-OE) and OsMAPK6 mutant dsg1, and demonstrated that OsMAPK6 positively regulated rice chilling tolerance. It was shown that, under cold stress, the survival rate of dsg1 was significantly lower than that of WT, whereas CAMAPK6-OE display higher survival rate than WT. Physiological assays indicate that ion leakage and dead cell in dsg1 was much more severe than those in WT and CAMAPK6-OE. Consistently, expression of chilling responsive genes in dsg1, including OsCBFs and OsTPP1, was significantly lower than that of in WT and CAMAPK6-OE. Biochemical analyses revealed that chilling stress promotes phosphorylation of OsMAPK6. Besides, we found that OsMAPK6 interacts with and phosphorylates two key regulators in rice cold signaling, OsIPA1 and OsICE1, and then enhance their protein stability. Overall, our results revealed a cold-induced OsMAPK6-OsICE1/OsIPA1 signaling cascade by which OsMAPK6 was involved in rice chilling tolerance, which provides novel insights to understand rice cold response at seedling stage.
Assuntos
Oryza , Plântula , Plântula/genética , Plântula/metabolismo , Oryza/metabolismo , Resposta ao Choque Frio/genética , Temperatura Baixa , Fosforilação , Regulação da Expressão Gênica de Plantas , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismoRESUMO
DNA methylation is an epigenetic modification that plays a vital role in a variety of biological processes, including the regulation of gene expression, cell differentiation, early embryonic development, genomic imprinting, and X chromosome inactivation. PGC7 is a maternal factor that maintains DNA methylation during early embryonic development. One mechanism of action has been identified by analyzing the interactions between PGC7 and UHRF1, H3K9 me2, or TET2/TET3, which reveals how PGC7 regulates DNA methylation in oocytes or fertilized embryos. However, the mechanism by which PGC7 regulates the post-translational modification of methylation-related enzymes remains to be elucidated. This study focused on F9 cells (embryonic cancer cells), which display high levels of PGC7 expression. We found that both knockdown of Pgc7 and inhibition of ERK activity resulted in increased genome-wide DNA methylation levels. Mechanistic experiments confirmed that inhibition of ERK activity led to the accumulation of DNMT1 in the nucleus, ERK phosphorylated DNMT1 at ser717, and DNMT1 Ser717-Ala mutation promoted the nuclear localization of DNMT1. Moreover, knockdown of Pgc7 also caused downregulation of ERK phosphorylation and promoted the accumulation of DNMT1 in the nucleus. In conclusion, we reveal a new mechanism by which PGC7 regulates genome-wide DNA methylation via phosphorylation of DNMT1 at ser717 by ERK. These findings may provide new insights into treatments for DNA methylation-related diseases.
Assuntos
Metilação de DNA , Epigênese Genética , Núcleo Celular/metabolismo , DNA (Citosina-5-)-Metiltransferase 1/genética , DNA (Citosina-5-)-Metiltransferases/genética , Impressão Genômica , Processamento de Proteína Pós-Traducional , Proteínas Cromossômicas não HistonaRESUMO
The gene dosage at the imprinted Dlk1-Dio3 locus is critical for cell growth and development. A relatively high gene expression within the Dlk1-Dio3 region, especially the active expression of Gtl2, has been identified as the only reliable marker for cell pluripotency. The DNA methylation state of the IG-DNA methylated regions (DMR), which is located upstream of the Gtl2 gene, dominantly contributes to the control of gene expression in the Dlk1-Dio3 locus. However, the precise mechanism underlying the regulation of DNA methylation in the IG-DMR remains largely unknown. Here, we use the F9 embryonal carcinoma cell line, a low pluripotent cell model, to identify the mechanism responsible for DNA methylation in the IG-DMR, and find that the interaction of PGC7 with UHRF1 is involved in maintaining DNA methylation and inducing DNA hypermethylation in the IG-DMR region. PGC7 and UHRF1 cooperatively bind in the IG-DMR to regulate the methylation of DNA and histones in this imprinted region. PGC7 promotes the recruitment of DNMT1 by UHRF1 to maintain DNA methylation in the IG-DMR locus. The interaction between PGC7 and UHRF1 strengthens their binding to H3K9me3 and leads to further enrichment of H3K9me3 in the IG-DMR by recruiting the specific histone methyltransferase SETDB1. Consequently, the abundance of H3K9me3 promotes DNMT3A to bind to the IG-DMR and increases DNA methylation level in this region. In summary, we propose a new mechanism of DNA methylation regulation in the IG-DMR locus and provide further insight into the understanding of the difference in Gtl2 expression levels between high and low pluripotent cells.
Assuntos
Metilação de DNA , RNA Longo não Codificante , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/metabolismo , DNA/metabolismo , Impressão Genômica , Histona Metiltransferases/genética , Histona Metiltransferases/metabolismo , Histonas/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/genética , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismoRESUMO
Flowering time is a fundamental factor determining the global distribution and final yield of rice (Oryza sativa). Although diverse flowering time genes have been reported in this crop, the transcriptional regulation of its key flowering genes are poorly understood. Here, we report that a basic leucine zipper transcription factor, bZIP71, functions as a flowering repressor. The overexpression of bZIP71 delays flowering, while the bzip71 mutant flowers early in both long-day and short-day conditions. A genetic analysis showed that the regulation of flowering by bZIP71 might be independent of Heading date 2 (Hd2), Hd4, and Hd5. Importantly, bZIP71 directly associates with the Early heading date 1 (Ehd1) promoter and represses its transcription, and genetically the function of bZIP71 is impaired in the ehd1 mutant. Moreover, bZIP71 interacts with major components of polycomb repressive complex 2 (PRC2), SET domain group protein 711 (SDG711), and Fertilization independent endosperm 2 (FIE2), through which bZIP71 regulates the H3K27me3 level of Ehd1. Taken together, we present a transcriptional regulatory mechanism in which bZIP71 enhances the H3K27me3 level of Ehd1 and transcriptionally represses its expression, which not only offers a novel insight into a flowering pathway, but also provides a valuable putative target for the genetic engineering and breeding of elite rice cultivars.
Assuntos
Oryza , Flores/fisiologia , Regulação da Expressão Gênica de Plantas/genética , Histonas/metabolismo , Oryza/metabolismo , Fotoperíodo , Melhoramento Vegetal , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismoRESUMO
Excessive activation of NLRP3 inflammasome is associated with the pathogenesis of inflammatory diseases. Pristimerin (Pri) is a quinonoid triterpene derived from traditional Chinese medical herb Celastraceae and Hippocrateaceae. Pri has shown antifungal, antibacterial, antioxidant, and anticancer activities. In this study we investigated whether NLRP3 inflammasome was associated with the anti-inflammatory activity of Pri. We showed that Pri (0.1-0.4 µM) dose-dependently blocked caspase-1 activation and IL-1ß maturation in LPS-primed mouse bone-marrow-derived macrophages (BMDMs). Pri specifically inhibited NLRP3 inflammasome activation, had no visible effects on NLRC4 and AIM2 inflammasome activation. Furthermore, we demonstrated that Pri blocked the assembly of the NLRP3 inflammasome via disturbing the interaction between NEK7 and NLRP3; the α, ß-unsaturated carbonyl moiety of Pri was essential for NLRP3 inflammasome inactivation. In LPS-induced systemic inflammation mouse model and MSU-induced mouse peritonitis model, preinjection of Pri (500 µg/kg, ip) produced remarkable therapeutic effects via inhibition of NLRP3 inflammasome in vivo. In HFD-induced diabetic mouse model, administration of Pri (100 µg· kg-1 ·d-1, ip, for 6 weeks) reversed HFD-induced metabolic disorders via suppression of NLRP3 inflammasome activation. Taken together, our results demonstrate that Pri acts as a NLRP3 inhibitor, suggesting that Pri might be useful for the treatment of NLRP3-associated diseases.
Assuntos
Anti-Inflamatórios/uso terapêutico , Inflamassomos/antagonistas & inibidores , Inflamação/prevenção & controle , Doenças Metabólicas/prevenção & controle , Proteína 3 que Contém Domínio de Pirina da Família NLR/antagonistas & inibidores , Triterpenos Pentacíclicos/uso terapêutico , Animais , Células Cultivadas , Inflamação/induzido quimicamente , Lipopolissacarídeos , Fígado/patologia , Masculino , Doenças Metabólicas/patologia , Camundongos Endogâmicos C57BL , Peritonite/induzido quimicamente , Peritonite/prevenção & controle , Ácido ÚricoRESUMO
Aberrant activation of signal transducer and activator of transcription 3 (STAT3) plays a critical role in many types of cancers. As a result, STAT3 has been identified as a potential target for cancer therapy. In this study we identified 10,11-dehydrocurvularin (DCV), a natural-product macrolide derived from marine fungus, as a selective STAT3 inhibitor. We showed that DCV (2-8 µM) dose-dependently inhibited the proliferation, migration and invasion of human breast cancer cell lines MDA-MB-231 and MDA-MB-468, and induced cell apoptosis. In the two breast cancer cell lines, DCV selectively inhibited the phosphorylation of STAT3 Tyr-705, but did not affect the upstream components JAK1 and JAK2, as well as dephosphorylation of STAT3. Furthermore, DCV treatment strongly inhibited IFN-γ-induced STAT3 phosphorylation but had no significant effect on IFN-γ-induced STAT1 and STAT5 phosphorylation in the two breast cancer cell lines. We demonstrated that the α, ß-unsaturated carbonyl moiety of DCV was essential for STAT3 inactivation. Cellular thermal shift assay (CETSA) further revealed the direct engagement of DCV with STAT3. In nude mice bearing breast cancer cell line MDA-MB-231 xenografts, treatment with DCV (30 mg·kg-1·d-1, ip, for 14 days) markedly suppressed the tumor growth via inhibition of STAT3 activation without observed toxicity. Our results demonstrate that DCV acts as a selective STAT3 inhibitor for breast cancer intervention.
Assuntos
Antineoplásicos/uso terapêutico , Neoplasias da Mama/tratamento farmacológico , Fator de Transcrição STAT3/antagonistas & inibidores , Zearalenona/análogos & derivados , Animais , Antineoplásicos/farmacologia , Antineoplásicos/toxicidade , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Núcleo Celular/metabolismo , Proliferação de Células/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Camundongos Endogâmicos BALB C , Camundongos Nus , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT3/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto , Zearalenona/farmacologia , Zearalenona/uso terapêutico , Zearalenona/toxicidadeRESUMO
Pristimerin, a triterpenoid, has exhibited potential anti-inflammatory and anti-tumor activities. Nevertheless, the role and mechanism of pristimerin in intestinal inflammation and colon cancer require further investigation. Here, we found that pristimerin protected mice from dextran sulfate sodium (DSS)-induced colitis, restoring epithelial damage and reducing tissue inflammation and inflammatory cell infiltration. In addition, pristimerin dramatically reduced the number and size of the tumors in a azoxymethane (AOM)/DSS-induced colitis-associated colorectal cancer (CAC) model. Furthermore, we found that pristimerin suppressed Wnt/ß-catenin signaling by RNA-Seq. Pristimerin inhibited Wnt/ß-catenin signaling via activation of GSK3ß, thereby suppressing Wnt target gene expression in colon cancer HCT116 and HT-29 cells. In HCT116 colon cancer xenografts and APCmin/+ mice, which undergo spontaneous intestinal tumorigenesis, administration of pristimerin reduced the tumor progression and decreased the expression of phosphorylated GSK3ß Ser 9, ß-catenin, cyclin D1 and c-Myc. These results suggest that pristimerin is a potent agent for preventing colon inflammation and carcinogenesis.
Assuntos
Antineoplásicos/uso terapêutico , Neoplasias Colorretais/tratamento farmacológico , Inflamação/tratamento farmacológico , Triterpenos/uso terapêutico , Via de Sinalização Wnt/efeitos dos fármacos , Animais , Modelos Animais de Doenças , Feminino , Glicogênio Sintase Quinase 3 beta/antagonistas & inibidores , Células HCT116 , Células HT29 , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Transplante de Neoplasias , Triterpenos PentacíclicosRESUMO
In our previous study, AtDBP1 encoding a DBP factor was identified as a putative abiotic stress candidate gene. DBP factors are important regulators that participate in both transcriptional regulation and post-translational regulation, but their roles in abiotic stress are still not well-understood. So we conducted a detailed study on the function of AtDBP1 in abiotic stress. It is found that expression of AtDBP1 could be induced by drought and salt, and the induction by salt was inhibited in ABA-deficient mutant aba2-3, indicating the expression of AtDBP1 was ABA-inducible. Overexpression of AtDBP1 resulted in a rapid stomatal closure, and elevated expression of drought/salt-responsive genes, which should help Arabidopsis to enhance the drought and salt tolerance. Unexpectedly, overexpression of AtDBP1 decreased the drought and salt tolerance of Arabidopsis. Further analysis suggested that AtDBP1 is involved in cuticle wax and cuticle membrane regulation. Overexpression of AtDBP1 showed increased cuticular conductance due to a decreased cuticle wax accumulation and cuticle membrane thickness. The cuticular wax provides an essential barrier for decreasing nonstomatal water loss during drought stress, so overexpression of AtDBP1 showed decreased drought tolerance possibly ascribed to the change of cuticle membrane structure. Our previous study elucidated that AtDBP1 was also involved in flowering time regulation. Taken together, the results above indicated that AtDBP1 was involved in both plant development and stress regulation. The mechanism of AtDBP1 in this study indicates that genes involved in both plant development and stress regulation might be not suitable for production application in breeding. Collectively, our results provide some new ideas on purposefully increasing the abiotic stress without influence on plant growth and development.
Assuntos
Proteínas de Arabidopsis/metabolismo , Proteínas de Ligação a DNA/metabolismo , Fosfoproteínas Fosfatases/metabolismo , Tolerância ao Sal/genética , Estresse Fisiológico/genética , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Ligação a DNA/genética , Secas , Regulação da Expressão Gênica de Plantas/genética , Permeabilidade , Fosfoproteínas Fosfatases/genética , Monoéster Fosfórico Hidrolases/genética , Monoéster Fosfórico Hidrolases/metabolismo , Folhas de Planta/metabolismo , Fatores de Transcrição/genéticaRESUMO
Chronic myeloid leukemia (CML) is a lethal malignancy, and the progress toward long-term survival has stagnated in recent decades. Pristimerin, a quinone methide triterpenoid isolated from the Celastraceae and Hippocrateaceae families, is well-known to exert potential anticancer activities. In this study, we investigated the effects and the mechanisms of action on CML. We found that pristimerin inhibited cell proliferation of K562 CML cells by causing G1 phase arrest. Furthermore, we demonstrated that pristimerin triggered autophagy and apoptosis. Intriguingly, pristimerin-induced cell death was restored by an autophagy inhibitor, suggesting that autophagy is cross-linked with pristimerin-induced apoptosis. Further studies revealed that pristimerin could produce excessive reactive oxygen species (ROS), which then induce JNK activation. These findings provide clear evidence that pristimerin might be clinical benefit to patients with CML.
Assuntos
Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Triterpenos/farmacologia , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Células K562 , Triterpenos Pentacíclicos , Espécies Reativas de Oxigênio/metabolismo , Triterpenos/químicaRESUMO
Retinoic acid (RA) plays a key role in pluripotent cell differentiation. In F9 embryonic carcinoma cells, RA can induce differentiation towards somatic lineages via the Ras-extracellular signal-regulated kinase (Ras/Erk) pathway, but the mechanism through which it induces the Erk1/2 phosphorylation is unclear. Here, we show that miR-485 is a positive regulator that targets α/ß-hydrolase domain-containing protein 2 (Abhd2), which can result in Erk1/2 phosphorylation and triggers differentiation. RA up-regulates miR-485 and concurrently down-regulates Abhd2. We verified that Abhd2 is targeted by miR-485 and they both can influence the phosphorylation of Erk1/2. In summary, RA can mediate cell differentiation by phosphorylating Erk1/2 via miR-485 and Abhd2.
Assuntos
Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Hidrolases/genética , MicroRNAs/genética , Interferência de RNA , Tretinoína/farmacologia , Animais , Biomarcadores , Células-Tronco de Carcinoma Embrionário , Humanos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Camundongos , FosforilaçãoRESUMO
The ability to self-renew is one of the most important properties of embryonic stem (ES) cells. Pluripotin (SC1), a small molecule with high activity and low toxicity, promotes self-renewal in mouse ES cells. SC1 can noticeably change the morphology of retinoic acid (RA)-induced F9 embryonic carcinoma cells (F9 cells). However, in the long term, RA and SC1 together cause cell apoptosis. When being added after 18-24 h of RA-induced F9 cell differentiation, SC1 transitorily activated Nanog and Oct4. Both Nanog and Oct4 were downregulated when SC1 and RA were added simultaneously. On the other hand, Klf4 was continually activated when SC1 was added between 6 and 24 h. Phosphorylated Erk1/2 protein levels were reduced from 6 to 24 h, whereas unphosphorylated Erk1 protein levels remained unchanged. A higher concentration of SC1 promoted cell self-renewal by strengthening the inhibition of Erk1/2 protein phosphorylation in F9 cells. Furthermore, SC1 and RA affect global DNA methylation by influencing the expressions of methylation-associated proteins, including Dnmt3b, Dnmt3l, Tet1, Tet2, and Tet3. In conclusion, SC1 inhibits the differentiation of RA-induced F9 cells mainly by reducing the levels of phosphorylated Erk1/2 and enhancing the expression of Klf4, although it also reduces DNA methylation, which may have an additional effect on ES cell differentiation.
Assuntos
Apoptose/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Pirazóis/farmacologia , Pirimidinas/farmacologia , Tretinoína/farmacologia , Animais , Antineoplásicos/farmacologia , Apoptose/genética , Carcinoma Embrionário/genética , Carcinoma Embrionário/metabolismo , Carcinoma Embrionário/patologia , Diferenciação Celular/genética , Linhagem Celular Tumoral , DNA (Citosina-5-)-Metiltransferases/genética , DNA (Citosina-5-)-Metiltransferases/metabolismo , Metilação de DNA/efeitos dos fármacos , Metilação de DNA/genética , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Fator 4 Semelhante a Kruppel , Camundongos , Proteína Homeobox Nanog/genética , Proteína Homeobox Nanog/metabolismo , Fator 3 de Transcrição de Octâmero/genética , Fator 3 de Transcrição de Octâmero/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismoRESUMO
It is important for companies to increase the efficiency of drilling as well as prolong the lifetime of the drilling tool. Since some previous investigations indicated that a superposition of well drilling with an additional vibration increases the drilling efficiency, this paper introduces a novel well drill which is assisted with additional vibrations by means of piezoelectric sandwich bending vibration transducer. The proposed drill uses bending vibrations in two different directions to from an elliptical trajectory movement, which can help the drill to break the surface of hard material more efficiently and clean away the lithic fragments more easily. The proposed well drill with bending vibration transducer is designed to have a resonance frequency of the first bending vibration mode of about 1779 Hz. The motion equation of the particle on the edge of the drill bit is developed and analyzed. The vibration trajectory of the particle on the edge of the drill bit is calculated by using finite element method. A prototype of the proposed drill using bending vibrations is fabricated and tested to verify the aim of drilling efficiency increase. The feed speed of the vibration assisted drilling is tested to be about 0.296 mm/s when the excitation voltage of the transducer is 300 V, while this speed decreases to about 0.195 mm/s when no vibration is added. This comparison shows that the feed speed of the vibration assisted drilling is about 52% higher than that of the normal drilling, which means the proposed drill has a better efficiency and it is important to consider vibration superimposition in well drilling. In addition, the surface of the drill hole gained by the vibration assisted drilling is smoother than that of the normal drilling, which makes the clearance easier.
RESUMO
A four-feet walking-type rotary piezoelectric actuator with minute step motion was proposed. The proposed actuator used the rectangular motions of four driving feet to push the rotor step-by-step; this operating principle was different with the previous non-resonant actuators using direct-driving, inertial-driving, and inchworm-type mechanisms. The mechanism of the proposed actuator was discussed in detail. Transient analyses were accomplished by ANSYS software to simulate the motion trajectory of the driving foot and to find the response characteristics. A prototype was manufactured to verify the mechanism and to test the mechanical characteristics. A minimum resolution of 0.095 μrad and a maximum torque of 49 N·mm were achieved by the prototype, and the output speed was varied by changing the driving voltage and working frequency. This work provides a new mechanism for the design of a rotary piezoelectric actuator with minute step motion.
RESUMO
BACKGROUND/AIMS: Self-renewal is one of the most important features of embryonic stem (ES) cells. SC1 is a small molecule modulator that effectively maintains the self-renewal of mouse ES cells in the absence of leukemia inhibitory factor (LIF), serum and feeder cells. However, the mechanism by which SC1 maintains the undifferentiated state of mouse ES cells remains unclear. METHODS: In this study, microarray and small RNA deep-sequencing experiments were performed on mouse ES cells treated with or without SC1 to identify the key genes and microRNAs that contributed to self-renewal. RESULTS: SC1 regulates the expressions of pluripotency and differentiation factors, and antagonizes the retinoic acid (RA)-induced differentiation in the presence or absence of LIF. SC1 inhibits the MEK/ERK pathway through Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis and pathway reporting experiments. Small RNA deep-sequencing revealed that SC1 significantly modulates the expression of multiple microRNAs with crucial functions in ES cells. The expression of miR124-3p is upregulated in SC1-treated ES cells, which significantly inhibits the MEK/ERK pathway by targeting Grb2, Sos2 and Egr1. CONCLUSION: SC1 enhances the self-renewal capacity of mouse ES cells by modulating the expression of key regulatory genes and pluripotency-associated microRNAs. SC1 significantly upregulates miR124-3p expression to further inhibit the MEK/ ERK pathway by targeting Grb2, Sos2 and Egr1.