RESUMO
Carnation (Dianthus caryophyllus L.) is one of the most famous and ethylene-sensitive cut flowers worldwide, but how ethylene interacts with other plant hormones and factors to regulate petal senescence in carnation is largely unknown. Here we found that a gene encoding WRKY family transcription factor, DcWRKY33, was significantly upregulated upon ethylene treatment. Silencing and overexpression of DcWRKY33 could delay and accelerate the senescence of carnation petals, respectively. Abscisic acid (ABA) and H2 O2 treatments could also accelerate the senescence of carnation petals by inducing the expression of DcWRKY33. Further, DcWRKY33 can bind directly to the promoters of ethylene biosynthesis genes (DcACS1 and DcACO1), ABA biosynthesis genes (DcNCED2 and DcNCED5), and the reactive oxygen species (ROS) generation gene DcRBOHB to activate their expression. Lastly, relationships are existed between ethylene, ABA and ROS. This study elucidated that DcWRKY33 promotes petal senescence by activating genes involved in the biosynthesis of ethylene and ABA and accumulation of ROS in carnation, supporting the development of new strategies to prolong the vase life of cut carnation.
Assuntos
Dianthus , Syzygium , Ácido Abscísico/metabolismo , Dianthus/genética , Espécies Reativas de Oxigênio/metabolismo , Syzygium/metabolismo , Etilenos/metabolismo , Flores , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
Carnation (Dianthus caryophyllus L.) is a respiratory climacteric flower, comprising one of the most important cut flowers that is extremely sensitive to plant hormone ethylene. Ethylene signaling core transcription factor DcEIL3-1 plays a key role in ethylene induced petal senescence in carnation. However, how the dose of DcEIL3-1 is regulated in the carnation petal senescence process is still not clear. Here, we screened out two EBF (EIN3 Binding F-box) genes, DcEBF1 and DcEBF2, which showed quick elevation by ethylene treatment according to the ethylene induced carnation petal senescence transcriptome. Silencing of DcEBF1 and DcEBF2 accelerated, whereas overexpression of DcEBF1 and DcEBF2 delayed, ethylene induced petal senescence in carnation by influencing DcEIL3-1 downstream target genes but not DcEIL3-1 itself. Furthermore, DcEBF1 and DcEBF2 interact with DcEIL3-1 to degrade DcEIL3-1 via an ubiquitination pathway in vitro and in vivo. Finally, DcEIL3-1 binds to the promoter regions of DcEBF1 and DcEBF2 to activate their expression. In conclusion, the present study reveals the mutual regulation between DcEBF1/2 and DcEIL3-1 during ethylene induced petal senescence in carnation, which not only expands our understanding about ethylene signal regulation network in the carnation petal senescence process, but also provides potential targets with respect to breeding a cultivar of long-lived cut carnation.
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Dianthus , Syzygium , Dianthus/genética , Syzygium/metabolismo , Melhoramento Vegetal , Etilenos/metabolismo , Flores/genética , Flores/metabolismoRESUMO
Dynamic DNA methylation regulatory networks are involved in many biological processes. However, how DNA methylation patterns change during flower senescence and their relevance with gene expression and related molecular mechanism remain largely unknown. Here, we used whole genome bisulfite sequencing to reveal a significant increase of DNA methylation in the promoter region of genes during natural and ethylene-induced flower senescence in carnation (Dianthus caryophyllus L.), which was correlated with decreased expression of DNA demethylase gene DcROS1. Silencing of DcROS1 accelerated while overexpression of DcROS1 delayed carnation flower senescence. Moreover, among the hypermethylated differentially expressed genes during flower senescence, we identified two amino acid biosynthesis genes, DcCARA and DcDHAD, with increased DNA methylation and reduced expression in DcROS1 silenced petals, and decreased DNA methylation and increased expression in DcROS1 overexpression petals, accompanied by decreased or increased amino acids content. Silencing of DcCARA and DcDHAD accelerates carnation flower senescence. We further showed that adding corresponding amino acids could largely rescue the senescence phenotype of DcROS1, DcCARA and DcDHAD silenced plants. Our study not only demonstrates an essential role of DcROS1-mediated remodeling of DNA methylation in flower senescence but also unravels a novel epigenetic regulatory mechanism underlying DNA methylation and amino acid biosynthesis during flower senescence.
Assuntos
Dianthus , Syzygium , Dianthus/genética , Syzygium/metabolismo , Senescência Vegetal , Metilação de DNA/genética , Aminoácidos/metabolismo , Flores/genética , Flores/metabolismoRESUMO
Petal senescence is controlled by a complex regulatory network. Epigenetic regulation like histone modification influences chromatin state and gene expression. However, the involvement of histone methylation in regulating petal senescence remains poorly understood. Here, we found that the trimethylation of histone H3 at Lysine 4 (H3K4me3) is increased during ethylene-induced petal senescence in carnation (Dianthus caryophyllus L.). H3K4me3 levels were positively associated with the expression of transcription factor DcWRKY75, ethylene biosynthetic genes 1-aminocyclopropane-1-carboxylic acid (ACC) synthase (DcACS1), and ACC oxidase (DcACO1), and senescence associated genes (SAGs) DcSAG12 and DcSAG29. Further, we identified that carnation ARABIDOPSIS HOMOLOG OF TRITHORAX1 (DcATX1) encodes a histone lysine methyltransferase which can methylate H3K4. Knockdown of DcATX1 delayed ethylene-induced petal senescence in carnation, which was associated with the down-regulated expression of DcWRKY75, DcACO1, and DcSAG12, whereas overexpression of DcATX1 exhibited the opposite effects. DcATX1 promoted the transcription of DcWRKY75, DcACO1, and DcSAG12 by elevating the H3K4me3 levels within their promoters. Overall, our results demonstrate that DcATX1 is a H3K4 methyltransferase that promotes the expression of DcWRKY75, DcACO1, DcSAG12 and potentially other downstream target genes by regulating H3K4me3 levels, thereby accelerating ethylene-induced petal senescence in carnation. This study further indicates that epigenetic regulation is important for plant senescence processes.
Assuntos
Dianthus , Dianthus/genética , Dianthus/metabolismo , Histona Metiltransferases/genética , Histona Metiltransferases/metabolismo , Histonas/metabolismo , Epigênese Genética , Etilenos/metabolismoRESUMO
OBJECTIVE: Placenta accreta spectrum disorder (PAS) is a serious obstetric complication associated with significant maternal morbidity and mortality. Prophylactic balloon occlusion (PBO), as an intravascular interventional therapies, has emerged as a potential management strategy for controlling massive hemorrhage in patients with PAS. However, current evidence about the clinical application of PBO in PAS patients are still controversial. This study aimed to evaluate the effectiveness and safety of PBO in the management of PAS. METHODS: A retrospective cohort study including PAS patients underwent cesarean delivery was conducted in a tertiary hospital from January 2015 to March 2022. Included PAS patients were further divided into balloon and control groups by whether PBO was performed. Groups were compared for demographic characteristics, intraoperative and postoperative parameters, maternal and neonatal outcomes, PBO-related complication and follow up outcomes. Additionally, multivariate-logistic regression analysis was performed to determine the definitive associations between PBO and risk of massive hemorrhage and hysterectomy. RESULTS: A total of 285 PAS patients met the inclusion criteria were included, of which 57 PAS patients underwent PBO (PBO group) and 228 women performed cesarean section (CS) without PBO (control group). Irrespective of the differences of baseline characteristics between the two groups, PBO intervention did not reduce the blood loss, hysterectomy rate and postoperative hospital stay, but it prolonged the operation time and increased the cost of hospitalization (All P < 0.05) Additionally, there were no significant differences in postoperative complications, neonatal outcomes, and follow-up outcomes(All P > 0.05). In particular, patients undergoing PBO were more likely to develop the venous thrombosis postoperatively (P = 0.001). However, multivariate logistic regression analysis showed that PBO significantly decreased the risk of massive hemorrhage (OR 0.289, 95%CI:0.109-0.766, P = 0.013). The grade of PAS and MRI with S2 invasion were the significant risk factors affecting massive hemorrhage(OR:6.232 and OR:5.380, P<0.001). CONCLUSION: PBO has the potential to reduce massive hemorrhage in PAS patients undergoing CS. Obstetricians should, however, be aware of potential complications arising from the PBO. Additionally, MRI with S2 invasion and PAS grade will be useful to identify PAS patients who at high risk and may benefit from PBO. In brief, PBO seem to be a promising alternative for management of PAS, yet well-designed randomized controlled trials are needed to convincingly demonstrate its benefits and triage the necessity of PBO.
Assuntos
Oclusão com Balão , Placenta Acreta , Recém-Nascido , Gravidez , Humanos , Feminino , Cesárea , Placenta Acreta/cirurgia , Estudos Retrospectivos , Perda Sanguínea Cirúrgica/prevenção & controle , Histerectomia , PlacentaRESUMO
Bladder cancer is a highly prevalent malignancy that presents significant difficulties in the management of wounds following surgery. The present study investigated the critical necessity to optimize wound healing techniques in patients undergoing bladder cancer surgery by contrasting conventional approaches with advanced modalities in order to promote recovery and mitigate complications. The study assessed the efficacy of conventional and advanced wound healing methods in these patients, taking into account the complex interaction of patient-specific factors and surgical complexities. A cross-sectional analysis was performed on 120 patients who underwent bladder cancer surgery at the first affiliated hospital of Wenzhou Medical University. In addition to medical record evaluations and direct wound assessments, patient interviews were utilized to gather information regarding demographics, surgical specifics, wound healing methodologies and postoperative results. Survival analysis and logistic regression were utilized in statistical analysis, with potential confounding variables such as age, comorbidities and type of surgery being accounted for. Advanced wound healing techniques, such as negative pressure wound therapy, tissue-engineered products, bioactive dressings and platelet-rich plasma (PRP), exhibited distinct advantage in comparison with conventional suturing. The aforementioned techniques, especially PRP, resulted in expedited wound healing, decreased rates of complications (p < 0.05) and enhanced secondary outcomes, including curtailed hospital stays and decreased rates of readmissions. PRP therapy, in particular, demonstrated significant improvements with the faster mean time to wound healing of 9 ± 2 days and lower complication incidence of 2 (6.7%) (p < 0.05), indicating its superior efficacy. A subgroup analysis revealed that younger patients, males and those undergoing laparoscopic surgery exhibited superior outcomes (p < 0.05). The results were further supported by logistic regression and Cox proportional hazards models, which further indicated that sophisticated techniques, notably PRP therapy with a hazard ratio of 3.00 (2.00-4.50) and adjusted odds ratio of 0.20 (0.09-0.43), were effective in improving postoperative recovery. The research clarified the significant advantages that advanced wound healing techniques offered in postoperative care of patients diagnosed with bladder cancer. By customizing these methods to suit the unique requirements of individual patients and specific circumstances of surgical procedures, they can significantly enhance the recuperation process after surgery and set a new standard for patient care.
Assuntos
Plasma Rico em Plaquetas , Neoplasias da Bexiga Urinária , Masculino , Humanos , Estudos Transversais , Cicatrização , Neoplasias da Bexiga Urinária/cirurgiaRESUMO
OBJECTIVE: To investigate the therapeutic utility of psychological nursing interventions for prostatic hyperplasia clients while they are receiving therapy. METHODS: Clinical data of 110 patients with prostate group hyperplasia who underwent treatment in our hospital were collected and analysed retrospectively, and the selected period was from October 2021 to October 2023. The 110 cases of prostate group hyperplasia patients were divided into a research group and a control group according to the different methods of care, and each group had 55 cases each. The research group received psychological nursing intervention based on the conventional nursing care given to the control group. The total treatment compliance rate and contentment with nursing were contrasted between the research and control groups, and changes in the Self Rating Anxiety Scale (SAS) score, Self Rating Depression Scale (SDS) score, Health Survey Short Form score, and sleep problems were observed between the research group and the control group. RESULTS: The research group's overall compliance rate was 94.55% (52/53), a substantial increase over the control group's rate, 69.09% (38/55), P < 0.01. Following nursing, the research group's SAS and SDS scores were considerably more reduced than those of the control group, and both groups' scores were substantially lower than they were prior to nursing (P < 0.05). CONCLUSION: This retrospective study found that psychological nursing intervention applied to patients with prostatic hyperplasia can effectively improve the patient's compliance with treatment, effectively reduce the occurrence of negative emotions, improve the patient's quality of life, and improve sleep problems. In addition, psychological nursing intervention can effectively alleviate the tension between nurses and patients, and is worthy of clinical application.
RESUMO
A large proportion of miscarriages are classified as unexplained miscarriages since no cause is identified. No reliable biomarkers or treatments are available for these pregnancy losses. While our transcriptomic sequencing has revealed substantial upregulation of miR-146b-5p in unexplained miscarriage villous tissues, its role and associated molecular processes have yet to be fully characterized. Our work revealed that relative to samples from normal pregnancy, miR-146b-5p was significantly elevated in villous tissues from unexplained miscarriage patients and displayed promising diagnostic potential. Moreover, miR-146b-5p agomir contributed to higher rates of embryonic resorption in ICR mice. When overexpressed in HTR-8/SVneo cells, miR-146b-5p attenuated the proliferative, invasive, and migratory activity of these cells while suppressing the expression of MMP9 and immune inflammation-associated cytokines, including IL1B, IL11, CXCL1, CXCL8, and CXCL12. Conversely, inhibition of its expression enhanced proliferation, migration, and invasion abilities. Mechanistically, IL-1 receptor-associated kinase-1 and a disintegrin and metalloproteinase 19 were identified as miR-146b-5p targets regulating trophoblast function, and silencing IL-1 receptor-associated kinase-1 had similar effects as miR-146b-5p overexpression, while IL-1 receptor-associated kinase-1 overexpression could partially reverse the inhibitory impact of this microRNA on trophoblasts. miR-146b-5p may inhibit trophoblast proliferation, migration, invasion, and implantation-associated inflammation by downregulating IL-1 receptor-associated kinase-1 and a disintegrin and metalloproteinase 19, participating in the pathogenesis of miscarriage and providing a critical biomarker and a promising therapeutic target for unexplained miscarriage.
Assuntos
Aborto Espontâneo , MicroRNAs , Camundongos , Animais , Gravidez , Feminino , Humanos , Aborto Espontâneo/genética , Quinases Associadas a Receptores de Interleucina-1/genética , Quinases Associadas a Receptores de Interleucina-1/metabolismo , Quinases Associadas a Receptores de Interleucina-1/farmacologia , Desintegrinas/metabolismo , Desintegrinas/farmacologia , Camundongos Endogâmicos ICR , MicroRNAs/genética , MicroRNAs/metabolismo , Trofoblastos/metabolismo , Inflamação/metabolismo , Proliferação de Células/fisiologia , Metaloproteases/metabolismo , Movimento Celular , Proteínas ADAM/metabolismoRESUMO
Petal senescence is the final stage of flower development. Transcriptional regulation plays key roles in this process. However, whether and how post-transcriptional regulation involved is still largely unknown. Here, we identified an ethylene-induced NAC family transcription factor DcNAP in carnation (Dianthus caryophyllus L.). One allele, DcNAP-dTdic1, has an insertion of a dTdic1 transposon in its second exon. The dTdic1 transposon disrupts the structure of DcNAP and causes alternative splicing, which transcribes multiple domain-deleted variants (DcNAP2 and others). Conversely, the wild type allele DcNAP transcribes DcNAP1 encoding an intact NAC domain. Silencing DcNAP1 delays and overexpressing DcNAP1 accelerates petal senescence in carnation, while silencing and overexpressing DcNAP2 have the opposite effects, respectively. Further, DcNAP2 could interact with DcNAP1 and interfere the binding and activation activity of DcNAP1 to the promoters of its downstream target ethylene biosynthesis genes DcACS1 and DcACO1. Lastly, ethylene signalling core transcriptional factor DcEIL3-1 can activate the expression of DcNAP1 and DcNAP2 in the same way by binding their promoters. In summary, we discovered a novel mechanism by which DcNAP regulates carnation petal senescence at the post-transcriptional level. It may also provide a useful strategy to manipulate the NAC domains of NAC transcription factors for crop genetic improvement.
Assuntos
Dianthus , Syzygium , Dianthus/genética , Syzygium/metabolismo , Flores , Etilenos/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
AIM: Cervical cancer is one of common diseases among women. There are limited therapies for patients with metastatic or recurrent cervical cancer. This study sought to explore the role of monoacylglycerol lipase (MAGL), an important metabolic enzyme, in cervical cancer progression. METHODS: In in vitro experiments, MAGL expression was inhibited by si-MAGL or JZL184 in cervical cancer cells. Quantitative real-time polymerase chain reaction and western blotting were performed to measure the expression of target molecules. Proliferation of cervical cancer cells was assessed by CCK-8 and colony formation assays. Apoptosis and cell cycle progression were evaluated by flow cytometry. The migration and invasion were detected by transwell assay. The in vivo tumor growth was detected in nude mice. TUNEL was utilized to observe apoptotic cells in tumor tissues. RESULTS: MAGL was upregulated in cervical cancer tissues and cells. Further, MAGL inhibition suppressed the growth of cervical cancer cells in vitro and in vivo. In addition, apoptosis and G1-phase cell cycle arrest were induced by MAGL knockdown. MAGL silencing-mediated upregulation of Bax and cleaved caspase-3, and downregulation of Bcl-2 was responsible for triggering apoptosis. More importantly, the migration and invasion of cervical cancer cells were restrained by MAGL depletion. CONCLUSIONS: MAGL drives the progression of cervical cancer, which can be a promising candidate to identify effective therapy for cervical cancer.
Assuntos
Monoacilglicerol Lipases , Neoplasias do Colo do Útero , Animais , Apoptose , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Feminino , Humanos , Camundongos , Camundongos Nus , Invasividade Neoplásica , Recidiva Local de NeoplasiaRESUMO
BACKGROUND: We designed a meta-analysis to evaluate the clinical significance and efficacy of circulating noncoding RNAs (ncRNAs) in the early prediction of preeclampsia. METHODS: PubMed, Embase and the Cochrane Library were used to search for literature. The combined prediction performance was evaluated by calculating the area under the summary receiver operator characteristic (SROC) curve. The potential sources of heterogeneity were analysed by meta-regression analysis and subgroup analysis. All statistical analyses and mapping were performed by RevMan 5.3 and Stata 12.0. RESULTS: A total of 41 studies from 14 articles, including 557 preeclampsia patients and 842 controls, were included in our meta-analysis. All studies collected blood before onset. NcRNAs in blood performed relatively well in predicting preeclampsia. The combined sensitivity was 0.71, the specificity was 0.84, and the area under the SROC curve (AUC) was 0.86. Peripheral blood mononuclear cell (PBMC) samples showed the best diagnostic accuracy. The combined AUC was 0.93. Combined detection was better than single detection, and miRNA was better than circRNA. The heterogeneity of the study was determined by sample size, lncRNA characteristics, lncRNA source and race. CONCLUSION: Circulating ncRNAs can be valuable biomarkers used as candidates for noninvasive early predictive biomarkers of preeclampsia and have great clinical application prospects. The clinical value of ncRNAs needs to be tested by further multicentre, comprehensive and prospective studies, and the test criteria should be established.
Assuntos
Ácidos Nucleicos Livres/sangue , Pré-Eclâmpsia/diagnóstico , RNA Longo não Codificante/sangue , Biomarcadores/sangue , Feminino , Humanos , Pré-Eclâmpsia/sangue , Gravidez , Sensibilidade e EspecificidadeRESUMO
A three-dimensional molecularly imprinted overoxidized polypyrrole/MnO2 /carbon felt (MIOPPy/MnO2 /CF) composites were fabricated, which results in increased active area of the surface of the molecularly imprinted polymers film and greatly improved electrochemical chiral recognition effect for tryptophan isomers. The composites were characterized by field emission scanning electron microscope, transmission electron microscope, X-ray diffractometer, UV-visible spectra, N2 adsorption-desorption isotherms, and electrochemical methods. The manuscript provides a proof of concept and shows promising potential of the prepared composites for chiral recognition.
RESUMO
Nucleic acid-based diagnostics are widely used for clinical applications due to their powerful recognition of biomolecule properties. Isolation and purification of nucleic acids such as DNA and RNA in the diagnostic system have been severely hampered in point-of-care testing because of low recovery yields, degradation of nucleic acids due to the use of chaotropic detergent and high temperature, and the requirement of large instruments such as centrifuges and thermal controllers. Here, we report a novel large instrument- and detergent-free assay via binary nanomaterial for ultrasensitive nucleic acid isolation and detection from cells (eukaryotic and prokaryotic). This binary nanomaterial couples a zinc oxide nanomultigonal shuttle (ZnO NMS) for cell membrane rupture without detergent and temperature control and diatomaceous earth with dimethyl suberimidate complex (DDS) for the capture and isolation of nucleic acids (NA) from cells. The ZnO NMS was synthesized to a size of 500 nm to permit efficient cell lysis at room temperature within 2 min using the biological, chemical, and physical properties of the nanomaterial. By combining the ZnO NMS with the DDS and proteinase K, the nucleic acid extraction could be completed in 15 min with high quantity and quality. For bacterial cells, DNA isolation with the binary nanomaterial yielded 100 times more DNA, than a commercial spin column based reference kit, as determined by the NanoDrop spectrophotometer. We believe that this binary nanomaterial will be a useful tool for rapid and sensitive nucleic acid isolation and detection without large instruments and detergent in the field of molecular diagnostics.
Assuntos
Nanoestruturas/química , Ácidos Nucleicos/análise , Reação em Cadeia da Polimerase/métodos , Bactérias/genética , DNA Bacteriano/análise , DNA Bacteriano/isolamento & purificação , Detergentes/química , Dimetil Suberimidato/química , Endopeptidase K/metabolismo , Células HCT116 , Humanos , Ácidos Nucleicos/isolamento & purificação , Tamanho da Partícula , Testes Imediatos , Temperatura , Óxido de Zinco/químicaRESUMO
Electrophilic carbonyl compounds are highly cytotoxic and genotoxic. Aldo-keto reductase 1B10 (AKR1B10) is an enzyme catalyzing reduction of carbonyl compounds to less toxic alcoholic forms. This study presents novel evidence that AKR1B10 protects colon cells from DNA damage induced by electrophilic carbonyl compounds. AKR1B10 is specifically expressed in epithelial cells of the human colon, but this study found that AKR1B10 expression was lost or markedly diminished in colorectal cancer, precancerous tissues, and a notable portion of normal adjacent tissues (NAT). SiRNA-mediated silencing of AKR1B10 in colon cancer cells HCT-8 enhanced cytotoxicity of acrolein and HNE, whereas ectopic expression of AKR1B10 in colon cancer cells RKO prevented the host cells against carbonyl cytotoxicity. Furthermore, siRNA-mediated AKR1B10 silencing led to DNA breaks and activation of γ-H2AX protein, a marker of DNA double strand breaks, particularly in the exposure of HNE (10 µM). In the AKR1B10 silenced HCT-8 cells, hypoxanthine-guanine phosphoribosyl transferase (HPRT) mutant frequency increased by 26.8 times at basal level and by 33.5 times in the presence of 10 µM HNE when compared to vector control cells. In these cells, the cyclic acrolein-deoxyguanosine adducts levels were increased by over 10 times. These findings were confirmed by pharmacological inhibition of AKR1B10 activity by Epalrestat. Taken together, these data suggest that AKR1B10 is a critical protein that protects host cells from DNA damage induced by electrophilic carbonyl compounds. AKR1B10 deficiency in the colon may be an important pathogenic factor in disease progression and carcinogenesis. © 2016 Wiley Periodicals, Inc.
Assuntos
Acroleína/toxicidade , Aldeído Redutase/metabolismo , Aldeídos/toxicidade , Neoplasias Colorretais/induzido quimicamente , Neoplasias Colorretais/metabolismo , Dano ao DNA/efeitos dos fármacos , Mutagênicos/toxicidade , Acroleína/metabolismo , Aldeído Redutase/análise , Aldeído Redutase/genética , Aldeídos/metabolismo , Aldo-Ceto Redutases , Linhagem Celular Tumoral , Colo/metabolismo , Colo/patologia , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Inativação Gênica , Humanos , Mutagênicos/metabolismo , Reto/metabolismo , Reto/patologiaRESUMO
RNA editing is a posttranscriptional process that covalently alters the sequence of RNA molecules and plays important biological roles in both animals and land plants. In flowering plants, RNA editing converts specific cytidine residues to uridine in both plastid and mitochondrial transcripts. Previous studies identified pentatricopeptide repeat (PPR) motif-containing proteins as site-specific recognition factors for cytidine targets in RNA sequences. However, the regulatory mechanism underlying RNA editing was largely unknown. Here, we report that protoporphyrinogen IX oxidase 1 (PPO1), an enzyme that catalyzes protoporphyrinogen IX into protoporphyrin IX in the tetrapyrrole biosynthetic pathway, plays an unexpected role in editing multiple sites of plastid RNA transcripts, most of which encode subunits of the NADH dehydrogenase-like complex (NDH), in the reference plant Arabidopsis thaliana. We identified multiple organellar RNA editing factors (MORFs), including MORF2, MORF8, and MORF9, that interact with PPO1. We found that two conserved motifs within the 22-aa region at the N terminus of PPO1 are essential for its interaction with MORFs, its RNA editing function, and subsequently, its effect on NDH activity. However, transgenic plants lacking key domains for the tetrapyrrole biosynthetic activity of PPO1 exhibit normal RNA editing. Furthermore, MORF2 and MORF9 interact with three PPRs or related proteins required for editing of ndhB and ndhD sites. These results reveal that the tetrapyrrole biosynthetic enzyme PPO1 is required for plastid RNA editing, acting as a regulator that promotes the stability of MORF proteins through physical interaction.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/enzimologia , Arabidopsis/genética , Plastídeos/enzimologia , Plastídeos/genética , Protoporfirinogênio Oxidase/metabolismo , Edição de RNA/genética , Tetrapirróis/biossíntese , Proteínas de Arabidopsis/genética , Sequência de Bases , Clorofila/biossíntese , Flavina-Adenina Dinucleotídeo/metabolismo , Dados de Sequência Molecular , NADH Desidrogenase/metabolismo , Fenótipo , Ligação Proteica , Protoporfirinogênio Oxidase/genética , Plântula/crescimento & desenvolvimento , Especificidade por SubstratoRESUMO
AIMS/HYPOTHESIS: Tankyrase (TNKS) is a ubiquitously expressed molecular scaffold that is implicated in diverse processes. The catalytic activity of TNKS modifies substrate proteins through poly-ADP-ribosylation (PARsylation) and is responsive to cellular energetic state. Global deficiency of the TNKS protein in mice accelerates glucose utilisation and raises plasma adiponectin levels. The aim of this study was to investigate whether the PARsylation activity of TNKS in adipocytes plays a role in systemic glucose homeostasis. METHODS: To inhibit TNKS-mediated PARsylation, we fed mice with a diet containing the TNKS-specific inhibitor G007-LK. To genetically inactivate TNKS catalysis in adipocytes while preserving its function as a molecular scaffold, we used an adipocyte-selective Cre transgene to delete TNKS exons that encoded the catalytic domain at the C-terminus. Tissue-specific insulin sensitivity in mice was investigated using hyperinsulinaemic-euglycaemic clamps. To model adipose-liver crosstalk ex vivo, we applied adipocyte-conditioned media to hepatocytes and assessed the effect on gluconeogenesis. RESULTS: The TNKS inhibitor G007-LK improved glucose tolerance and insulin sensitivity and promptly increased plasma adiponectin levels. In female mice, but not in male mice, adipocyte-selective genetic inactivation of TNKS catalysis improved hepatic insulin sensitivity and post-transcriptionally increased plasma adiponectin levels. Both pharmacological and genetic TNKS inhibition in female mouse-derived adipocytes induced a change in secreted factors to decrease gluconeogenesis in primary hepatocytes. CONCLUSIONS/INTERPRETATION: Systemic glucose homeostasis is regulated by the PARsylation activity of TNKS in adipocytes. This regulation is mediated in part by adipocyte-secreted factors that modulate hepatic glucose production. Pharmacological TNKS inhibition could potentially be used to improve glucose tolerance.
Assuntos
Tecido Adiposo/efeitos dos fármacos , Tecido Adiposo/enzimologia , Glucose/metabolismo , Tanquirases/metabolismo , Animais , Glicemia/efeitos dos fármacos , Metabolismo dos Carboidratos/efeitos dos fármacos , Feminino , Masculino , Camundongos , Sulfonas/farmacologia , Tanquirases/antagonistas & inibidores , Triazóis/farmacologiaRESUMO
Compared with small heat shock proteins (sHSPs) in other organisms, those in plants are the most abundant and diverse. However, the molecular mechanisms by which sHSPs are involved in cell protection remain unknown. Here, we characterized the role of HSP21, a plastid nucleoid-localized sHSP, in chloroplast development under heat stress. We show that an Arabidopsis thaliana knockout mutant of HSP21 had an ivory phenotype under heat stress. Quantitative real-time RT-PCR, run-on transcription, RNA gel blot, and polysome association analyses demonstrated that HSP21 is involved in plastid-encoded RNA polymerase (PEP)-dependent transcription. We found that the plastid nucleoid protein pTAC5 was an HSP21 target. pTAC5 has a C4-type zinc finger similar to that of Escherichia coli DnaJ and zinc-dependent disulfide isomerase activity. Reduction of pTAC5 expression by RNA interference led to similar phenotypic effects as observed in hsp21. HSP21 and pTAC5 formed a complex that was associated mainly with the PEP complex. HSP21 and pTAC5 were associated with the PEP complex not only during transcription initiation, but also during elongation and termination. Our results suggest that HSP21 and pTAC5 are required for chloroplast development under heat stress by maintaining PEP function.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/fisiologia , Proteínas de Transporte/metabolismo , Cloroplastos/metabolismo , Proteínas de Choque Térmico/metabolismo , Resposta ao Choque Térmico , Arabidopsis/efeitos dos fármacos , Arabidopsis/genética , Arabidopsis/efeitos da radiação , Proteínas de Arabidopsis/química , Proteínas de Transporte/química , Cloroplastos/efeitos dos fármacos , Cloroplastos/genética , Cloroplastos/efeitos da radiação , DNA de Cloroplastos/metabolismo , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Regulação da Expressão Gênica de Plantas/efeitos da radiação , Genes de Plantas/genética , Proteínas de Choque Térmico/química , Resposta ao Choque Térmico/efeitos dos fármacos , Resposta ao Choque Térmico/genética , Resposta ao Choque Térmico/efeitos da radiação , Luz , Mutação/genética , Fenótipo , Plantas Geneticamente Modificadas , Ligação Proteica/efeitos dos fármacos , Ligação Proteica/efeitos da radiação , Isomerases de Dissulfetos de Proteínas/metabolismo , Estrutura Terciária de Proteína , Transporte Proteico/efeitos dos fármacos , Transporte Proteico/efeitos da radiação , Plântula/efeitos dos fármacos , Plântula/crescimento & desenvolvimento , Plântula/metabolismo , Plântula/efeitos da radiação , Deleção de Sequência , Frações Subcelulares/efeitos dos fármacos , Frações Subcelulares/metabolismo , Frações Subcelulares/efeitos da radiação , Zinco/farmacologiaRESUMO
The ingestion of fruits containing perfluoroalkyl acids (PFAAs) presents potential hazards to human health. This study aimed to fill knowledge gaps concerning the tissue-specific distribution patterns and bioaccumulation behavior of PFAAs and their isomers, alternatives, and precursors (collectively as per-/polyfluoroalkyl substances, PFASs) within citrus trees growing in contaminated fields. It also assessed the potential contribution of precursor degradation to human exposure risk of PFASs. High concentrations of total target PFASs (∑PFASstarget, 92.45-7496.16 ng/g dw) and precursors measured through the total oxidizable precursor (TOP) assay (130.80-13979.21 ng/g dw) were found in citrus tree tissues, and short-chain PFASs constituted the primary components. The total PFASs concentrations followed the order of leaves > fruits > branches, bark > wood, and peel > pulp > seeds. The average contamination burden of peel (∑PFASstarget: 57.75%; precursors: 71.15%) was highest among fruit tissues. Bioaccumulation factors (BAFs) and translocation potentials of short-chain, branched, or carboxylate-based PFASs exceeded those of their relatively hydrophobic counterparts, while ether-based PFASs showed lower BAFs than similar PFAAs in above-ground tissues of citrus trees. In the risk assessment of residents consuming contaminated citruses, precursor degradation contributed approximately 36.07% to total PFASs exposure, and therefore should not be ignored.
Assuntos
Ácidos Alcanossulfônicos , Fluorocarbonos , Poluentes Químicos da Água , Humanos , Árvores , Bioacumulação , Fluorocarbonos/análise , Poluentes Químicos da Água/química , Medição de Risco , Ácidos Alcanossulfônicos/análise , Monitoramento AmbientalRESUMO
Plants have evolved sophisticated mechanisms to regulate gene expression to activate immune responses against pathogen infections. However, how the translation system contributes to plant immunity is largely unknown. The evolutionarily conserved thiolation modification of transfer RNA (tRNA) ensures efficient decoding during translation. Here, we show that tRNA thiolation is required for plant immunity in Arabidopsis. We identify a cgb mutant that is hyper-susceptible to the pathogen Pseudomonas syringae. CGB encodes ROL5, a homolog of yeast NCS6 required for tRNA thiolation. ROL5 physically interacts with CTU2, a homolog of yeast NCS2. Mutations in either ROL5 or CTU2 result in loss of tRNA thiolation. Further analyses reveal that both transcriptome and proteome reprogramming during immune responses are compromised in cgb. Notably, the translation of salicylic acid receptor NPR1 is reduced in cgb, resulting in compromised salicylic acid signaling. Our study not only reveals a regulatory mechanism for plant immunity but also uncovers an additional biological function of tRNA thiolation.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Saccharomyces cerevisiae/genética , Arabidopsis/metabolismo , Mutação , RNA de Transferência/genética , RNA de Transferência/metabolismo , Imunidade Vegetal/genética , Ácido Salicílico/metabolismo , Regulação da Expressão Gênica de Plantas , Doenças das Plantas/genéticaRESUMO
BACKGROUND: Haemoglobin H (HbH) disease is caused by a disorder of α-globin synthesis, and it results in a wide range of clinical symptoms. M6A methylation modification may be one of the mechanisms of heterogeneity. Therefore, this article explored the role of methyltransferase like 16 (METTL16) in HbH disease. METHOD: The results of epigenetic transcriptome microarray were analysed and verified through bioinformatic methods and qRT-PCR, respectively. The overexpression or knock down of METTL16 in K562 cells was examined to determine its role in reactive oxygen species (ROS), cell cycle processes or iron overload. YTH domain family protein 3 (YTHDF3) was knocked down in K562 cells and K562 cells overexpressing METTL16 via siRNA to investigate its function. In addition, haemoglobin expression was detected through benzidine staining. qRT-PCR, WB, methylated RNA Immunoprecipitation (MeRIP) and (RNA Immunoprecipitation) RIP experiments were conducted to explore the mechanism of intermolecular interaction. RESULTS: METTL16, YTHDF3 and solute carrier family 5 member 3 (SLC5A3) mRNA and the methylation level of SLC5A3 mRNA were downregulated in HbH patients. Insulin-like growth factor 2 mRNA-binding protein 3 (IGF2BP3) mRNA expression was negatively correlated with HGB content among patients with HbH-CS disease. Overexpression of METTL16 increased ROS and intracellular iron contents in K562 cells, changed the K562 cell cycle, reduced hemin-induced haemoglobin synthesis, increased the expressions of SLC5A3 and HBG and increased SLC5A3 mRNA methylation levels. Knockdown of METTL16 reduced ROS and intracellular iron contents in K562 cells. Hemin treatment of K562 cells for more than 14 days reduced the protein expressions of METTL16 and SLC5A3 and SLC5A3 mRNA methylation levels. Knockdown of YTHDF3 rescued the intracellular iron content changes induced by the overexpression of METTL16. The RIP experiment revealed that SLC5A3 mRNA can be enriched by METTL16 antibody. CONCLUSION: METTL16 may affect the expression of SLC5A3 by changing its m6A modification level and regulating ROS synthesis, intracellular iron and cycle of red blood cells. Moreover, METTL16 possibly affects the expression of haemoglobin through IGF2BP3, which regulates the clinical phenotype of HbH disease.