RESUMO
Exoskeletons have decreased physical effort and increased comfort in activities of daily living (ADL) such as walking, squatting, and running. However, this assistance is often activity specific and does not accommodate a wide variety of different activities. To overcome this limitation and increase the scope of exoskeleton application, an automatic human activity recognition (HAR) system is necessary. We developed two deep-learning models for HAR using one-dimensional-convolutional neural network (CNN) and a hybrid model using CNNs and long-short term memory (LSTM). We trained both models using the data collected from a single three-axis accelerometer placed on the chest of ten subjects. We were able to classify five different activities, standing, walking on level ground, walking on an incline, running, and squatting, with an accuracy of 98.1% and 97.8%, respectively. A two subject real-time validation trial was also conducted to validate the real-time applicability of the system. The real-time accuracy was measured at 96.6% and 97.2% for the CNN and the hybrid model, respectively. The high classification accuracy in the test and real-time evaluation suggests that a single sensor could distinguish human activities using machine-learning-based models.
Assuntos
Atividades Cotidianas , Redes Neurais de Computação , Humanos , Aprendizado de Máquina , Atividades Humanas , AcelerometriaRESUMO
Pollen tube growth is essential for successful double fertilization, which is critical for grain yield in crop plants. Rapid alkalinization factors (RALFs) function as ligands for signal transduction during fertilization. However, functional studies on RALF in monocot plants are lacking. Herein, we functionally characterized two pollen-specific RALFs in rice (Oryza sativa) using multiple clustered regularly interspaced palindromic repeats (CRISPR)/CRISPR-associated protein 9-induced loss-of-function mutants, peptide treatment, expression analyses, and tag reporter lines. Among the 41 RALF members in rice, OsRALF17 was specifically expressed at the highest level in pollen and pollen tubes. Exogenously applied OsRALF17 or OsRALF19 peptide inhibited pollen tube germination and elongation at high concentrations but enhanced tube elongation at low concentrations, indicating growth regulation. Double mutants of OsRALF17 and OsRALF19 (ralf17/19) exhibited almost full male sterility with defects in pollen hydration, germination, and tube elongation, which was partially recovered by exogenous treatment with OsRALF17 peptide. This study revealed that two partially functionally redundant OsRALF17 and OsRALF19 bind to Oryza sativa male-gene transfer defective 2 (OsMTD2) and transmit reactive oxygen species signals for pollen tube germination and integrity maintenance in rice. Transcriptomic analysis confirmed their common downstream genes, in osmtd2 and ralf17/19. This study provides new insights into the role of RALF, expanding our knowledge of the biological role of RALF in regulating rice fertilization.
Assuntos
Oryza , Tubo Polínico , Tubo Polínico/genética , Pólen/genética , Transdução de Sinais , PeptídeosRESUMO
In flowering plants, double fertilization between male and female gametophytes, which are separated by distance, largely depends on the unique pattern of the male gametophyte (pollen): two non-motile sperm cells suspended within a tube-producing vegetative cell. A morphological screen to elucidate the genetic control governing the strategic patterning of pollen has led to the isolation of a sticky generative cell (sgc) mutant that dehisces abnormal pollen with the generative cell immobilized at the pollen wall. Analyses revealed that the sgc mutation is specifically detrimental to pollen development, causing ectopic callose deposition that impedes the timely internalization and differentiation of the generative cell. We found that the SGC gene encodes the highly conserved domain of unknown function 707 (DUF707) gene that is broadly expressed but is germline specific during pollen development. Additionally, transgenic plants co-expressing fluorescently fused SGC protein and known organelle markers showed that SGC localizes in the endoplasmic reticulum, Golgi apparatus and vacuoles in pollen. A yeast two-hybrid screen with an SGC bait identified a thaumatin-like protein that we named GCTLP1, some homologs of which bind and/or digest ß-1,3-glucans, the main constituent of callose. GCTLP1 is expressed in a germline-specific manner and colocalizes with SGC during pollen development, indicating that GCTLP1 is a putative SGC interactor. Collectively, our results show that SGC suppresses callose deposition in the nascent generative cell, thereby allowing the generative cell to fully internalize into the vegetative cell and correctly differentiate as the germline progenitor, with the potential involvement of the GCTLP1 protein, during pollen development in Arabidopsis.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Glucanos/metabolismo , Pólen/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Diferenciação Celular/genética , Diferenciação Celular/fisiologia , Regulação da Expressão Gênica de Plantas/genética , Regulação da Expressão Gênica de Plantas/fisiologia , Glucanos/genética , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/metabolismo , Pólen/fisiologiaRESUMO
The highly specialized haploid male gametophyte-pollen consist of two sperm cells and a large vegetative cell. Successful fertilization requires proper growth timing and rupture of the pollen tube until it delivers sperm cells, which occur immediately after a pollen grain hydrates. Although a tight regulation on polar cell-wall expansion of the pollen tube is fundamentally important, the underlying molecular mechanism remains largely unknown, especially in crop plants. Here, we characterized the function of male-gene transfer defective 2 (OsMTD2) gene in rice (Oryza sativa), which belongs to the plant-specific receptor-like kinase, the CrRLK1L family. We demonstrated that OsMTD2 is an essential male factor participating in pollen-tube elongation based on genetic evidence and physiological observations. Because of unavailability of homozygous mutant via conventional methods, we used CRISPR-Cas9 system to obtain homozygous knockout mutant of OsMTD2. We were able to identify phenotypic changes including male sterility due to early pollen-tube rupture in the mutant. We observed that the production of reactive oxygen species (ROS) was dramatically reduced in mutants of OsMTD2 pollen grain and tubes with defective pectin distribution. Transcriptome analysis of osmtd2-2 versus wild-type anthers revealed that genes involved in defense responses, metabolic alteration, transcriptional and protein modification were highly upregulated in the osmtd2-2 mutant. Through yeast-two-hybrid screening, we found that OsMTD2 kinase interacts with E3 ligase SPL11. Taken together, we propose that OsMTD2 has crucial functions in promoting pollen-tube elongation through cell-wall modification, possibly by modulating ROS homeostasis during pollen-tube growth.
Assuntos
Oryza/fisiologia , Proteínas de Plantas/metabolismo , Tubo Polínico/fisiologia , Espécies Reativas de Oxigênio/metabolismo , Membrana Celular/metabolismo , Regulação da Expressão Gênica de Plantas , Germinação , Mutação , Proteínas de Plantas/genética , Plantas Geneticamente Modificadas , Pólen/genética , Processamento de Proteína Pós-Traducional , Técnicas do Sistema de Duplo-HíbridoRESUMO
Successful delivery of sperm cells to the embryo sac in higher plants is mediated by pollen tube growth. The molecular mechanisms underlying pollen germination and tube growth in crop plants remain rather unclear, although these mechanisms are crucial to plant reproduction and seed formation. By screening pollen-specific gene mutants in rice (Oryza sativa), we identified a T-DNA insertional mutant of Germinating modulator of rice pollen (GORI) that showed a one-to-one segregation ratio for wild type (WT) to heterozygous. GORI encodes a seven-WD40-motif protein that is homologous to JINGUBANG/REN4 in Arabidopsis. GORI is specifically expressed in rice pollen, and its protein is localized in the nucleus, cytosol and plasma membrane. Furthermore, a homozygous mutant, gori-2, created through CRISPR-Cas9 clearly exhibited male sterility with disruption of pollen tube germination and elongation. The germinated pollen tube of gori-2 exhibited decreased actin filaments and altered pectin distribution. Transcriptome analysis revealed that 852 pollen-specific genes were downregulated in gori-2 compared with the WT, and Gene Ontology enrichment analysis indicated that these genes are strongly associated with cell wall modification and clathrin coat assembly. Based on the molecular features of GORI, phenotypical observation of the gori mutant and its interaction with endocytic proteins and Rac GTPase, we propose that GORI plays key roles in forming endo-/exocytosis complexes that could mediate pollen tube growth in rice.
Assuntos
Oryza/metabolismo , Proteínas de Plantas/metabolismo , Tubo Polínico/metabolismo , Regulação da Expressão Gênica de Plantas , Germinação/genética , Germinação/fisiologia , Oryza/genética , Proteínas de Plantas/genética , Tubo Polínico/genética , RNA-SeqRESUMO
Aflatoxins are carcinogenic secondary metabolites produced by Aspergillus section Flavi that contaminates a wide variety of food and feed products and is responsible for serious health and economic consequences. Fermented foods are prepared with a wide variety of substrates over a long fermentation time and are thus vulnerable to contamination by aflatoxin-producing fungi, leading to the production of aflatoxin B1. The mitigation and control of aflatoxin is currently a prime focus for developing safe aflatoxin-free food. This review summarizes the role of major aflatoxin-degrading enzymes such as laccase, peroxidase, and lactonase, and microorganisms in the context of their application in food. A putative mechanism of enzyme-mediated aflatoxin degradation and toxicity evaluation of the degraded products are also extensively discussed to evaluate the safety of degradation processes for food applications. The review also describes aflatoxin-degrading microorganisms isolated from fermented products and investigates their applicability in food as aflatoxin preventing agents. Furthermore, a summary of recent technological advancements in protein engineering, nanozymes, in silico and statistical optimization approaches are explored to improve the industrial applicability of aflatoxin-degrading enzymes.
Assuntos
Aflatoxinas , Aflatoxina B1/metabolismo , Aflatoxina B1/toxicidade , Aflatoxinas/análise , Aspergillus/metabolismo , Contaminação de Alimentos/análise , Contaminação de Alimentos/prevenção & controle , Fungos/metabolismo , Inativação MetabólicaRESUMO
Natural products derived carbon quantum dots (CQDs) catch huge attention owing to their distinctive properties of smaller size, water dispersibility, high photostability, lower cost, tunable emission, biocompatibility, least toxicity, electrical conductivity, optical and catalytic properties, and easy modification. Herein high fluorescent CQDs were prepared using Borassus flabellifer (ice apple) as a carbon source utilizing the simplistic one-step hydrothermal method. The prepared CQDs possessed excellent photoluminescence, high photostability, and stability in an aqueous solution and harbored large of quantum yield and strong stability in high pH conditions with the characteristic strong blue fluorescence emission. With these superior properties, the CQDs have been used as sensing probes for the detection of Fe3+ ions having excellent selectivity and sensitivity with a 2.01 µM limit of detection. The CQDs decorated probe was found effective in detecting Fe3+ ions in the tap and drinking mineral water, suggesting the applicability of the prepared sensor. The developed sensor exhibited advantages, including simple, low-cost, label-free, rapid, and good sensitivity and selectivity towards Fe3+ ions, with a great application for detection of such ions in real water.
Assuntos
Pontos Quânticos , Carbono/química , Endosperma , Corantes Fluorescentes/química , Íons , Pontos Quânticos/química , ÁguaRESUMO
Monophosphoryl lipid A (MPLA) is a toll-like receptor 4 ligand that promotes immune activation in mice and humans, without undesired inflammation. Immunotherapy by the combining immune checkpoint blockade and MPLA has shown promising anti-cancer effects in both mice and humans. In this study, we explored how MPLA enhanced the anti-cancer effects of anti-PD-L1 antibodies (Abs). Anti-cancer immunity induced by the combination of anti-PD-L1 Abs and MPLA failed in CD4 and CD8 cell-depleted mice. Moreover, the combination treatment of anti-PD-L1 Abs and MPLA synergistically enhanced the activation of plasmacytoid dendritic cells (pDCs) in the mouse in vivo, while conventional DCs were not. In addition, mice treated with anti-PD-L1 Abs and MPLA were not protected from B16 melanoma by blockade of interferon-alpha receptor (IFNAR). The combination of anti-PD-L1 Abs and MPLA also promoted human peripheral blood pDC activation and induced IFN-α-dependent T cell activation. Therefore, these results demonstrate that MPLA enhances anti-PD-L1 Ab-mediated anti-cancer immunity through the activation and IFN-α production of pDCs.
Assuntos
Antígeno B7-H1/antagonistas & inibidores , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/imunologia , Inibidores de Checkpoint Imunológico/farmacologia , Lipídeo A/análogos & derivados , Animais , Biomarcadores , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Citocinas/metabolismo , Células Dendríticas/metabolismo , Feminino , Humanos , Imunofenotipagem , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/metabolismo , Lipídeo A/farmacologia , Ativação Linfocitária/efeitos dos fármacos , Ativação Linfocitária/imunologia , Melanoma Experimental , CamundongosRESUMO
Paeonol exerts various pharmacological effects owing to its antiangiogenic, antioxidant, and antidiabetic activities. We aimed to investigate the transport mechanism of paeonol across the inner blood-retinal barrier both in vitro and in vivo. The carotid artery single injection method was used to investigate the retina uptake index of paeonol. The retina uptake index (RUI) value of [³H]paeonol was dependent on both concentration and pH. This value decreased significantly in the presence of imperatorin, tramadol, and pyrilamine when compared to the control. However, para-aminohippuric acid, choline, and taurine had no effect on the RUI value. Conditionally immortalized rat retina capillary endothelial cells (TR-iBRB cell lines) were used as an in vitro model of the inner blood-retinal barrier (iBRB). The uptake of [³H]paeonol by the TR-iBRB cell lines was found to be time-, concentration-, and pH-dependent. However, the uptake was unaffected by the absence of sodium or by membrane potential disruption. Moreover, in vitro structural analog studies revealed that [³H]paeonol uptake was inhibited in the presence of organic cationic compounds including imperatorin, clonidine and tramadol. This is consistent with the results obtained in vivo. In addition, transfections with OCTN1, 2 or plasma membrane monoamine transporter (PMAT) small interfering RNA did not affect paeonol uptake in TR-iBRB cell lines. Upon pre-incubation of these cell lines with high glucose (HG) media, [3H]paeonol uptake decreased and mRNA expression levels of angiogenetic factors, such as hypoxia inducible factor-1 (HIF-1) and vascular endothelial growth factor (VEGF) increased. However, after the pretreatment of unlabeled paeonol in HG conditions, the mRNA levels of VEGF and HIF-1 were comparatively reduced, and the [3H]paeonol uptake rate was restored. After being exposed to inflammatory conditions induced by glutamate, TNF-α, and LPS, paeonol and propranolol pretreatment significantly increased the uptake of both [3H]paeonol and [3H]propranolol in TR-iBRB cell lines compared to their respective controls. Our results demonstrate that the transport of paeonol to the retina across the iBRB may involve the proton-coupled organic cation antiporter system, and the uptake of paeonol is changed by HG conditions.
Assuntos
Acetofenonas/metabolismo , Barreira Hematorretiniana/efeitos dos fármacos , Glucose/farmacologia , Doenças Retinianas/metabolismo , Animais , Transporte Biológico , Linhagem Celular , Modelos Animais de Doenças , Masculino , Ratos , Ratos Sprague-Dawley , Doenças Retinianas/patologiaRESUMO
Correction for 'Specific detection of Cronobacter sakazakii in powdered infant formula using ssDNA aptamer' by Hye Ri Kim et al., Analyst, 2021, 146, 3534-3542, DOI: .
RESUMO
Cronobacter sakazakii (C. sakazakii) is a foodborne pathogen associated with bacterial meningitis, sepsis, and necrotizing enterocolitis in premature and immuno-compromised infants. C. sakazakii is typically acquired by ingesting contaminated powdered infant formula (PIF). The growing demand for a safe food supply requires rapid detection of foodborne pathogens for delivering safe-to-consume food to consumers. In the present study, we isolated C. sakazakii-specific aptamers using a centrifugation-based partitioning method (CBPM) instead of systematic evolution of ligands by exponential enrichment (SELEX) process. Unlike SELEX, the CBPM reduces the evolution-loop time to obtain enriched probes, allowing the isolation of target-specific aptamers in a shorter time. The two aptamers (SC25 and SC45) isolated using the CBPM showed high affinity and specificity for C. sakazakii (Kd: 34 and 66 nM). Among the two aptamers, SC25 aptamer detected efficiently C. sakazakii in PIF with less cross-reactivity. Our results indicate that the isolated aptamers could be used for detecting C. sakazakii in PIF and reducing the overall testing time compared with the conventional C. sakazakii detection method.
Assuntos
Cronobacter sakazakii , Cronobacter sakazakii/genética , DNA de Cadeia Simples , Microbiologia de Alimentos , Humanos , Lactente , Fórmulas Infantis , Recém-Nascido , Oligonucleotídeos , PósRESUMO
Prosthesis features that enhance balance are desirable to people with transtibial amputation. Ankle inversion/eversion compliance is intended to improve balance on uneven ground, but its effects remain unclear on level ground. We posited that increasing ankle inversion/eversion stiffness during level-ground walking would reduce balance-related effort by assisting in recovery from small disturbances in frontal-plane motions. We performed a pilot test with an ankle-foot prosthesis emulator programmed to apply inversion/eversion torques in proportion to the deviation from a nominal inversion/eversion position trajectory. We applied a range of stiffnesses to clearly understand the effect of the stiffness on balance-related effort, hypothesizing that positive stiffness would reduce effort while negative stiffness would increase effort. Nominal joint angle trajectories were calculated online as a moving average over several steps. In experiments with K3 ambulators with unilateral transtibial amputation (N = 5), stiffness affected step-width variability, average step width, margin of stability, intact-foot center of pressure variability, and user satisfaction (p ≤ 0.05, Friedman's test), but not intact-limb evertor average, intact-limb evertor variability, and metabolic rate (p ≥ 0.38, Friedman's test). Compared to zero stiffness, high positive stiffness reduced step-width variability by 13%, step width by 3%, margin of stability by 3%, and intact-foot center of pressure variability by 14%, whereas high negative stiffness had opposite effects and decreased satisfaction by 63%. The results of this pilot study suggest that positive ankle inversion stiffness can reduce active control requirements during level walking.
Assuntos
Membros Artificiais , Fenômenos Biomecânicos , Marcha , Projetos PilotoRESUMO
A fluorometric immunoassay is described for the determination of patulin, a highly toxic fungal metabolite. A rabbit anti-patulin-bovine serum albumin (BSA) IgG conjugate was prepared and used to compose immunoliposomes and immunomagnetic nanoparticles. The immunomagnetic nanoparticles are then added to the sample to form the patulin-antibody composites which can be magnetically separated. The immunoliposomes are then added to form a sandwich. After magnetic separation of the composites and adding n-octyl-ß-D-glucopyranoside, the fluorophore sulforhodamine B (SRB) is released. Its fluorescence intensity was then measured at excitation/emission wavelengths of 550/585 nm. The immunoliposome-based immunomagnetic nanoparticle assay can detect 8 µg L-1 of patulin in apple juice without the need for extraction, separation, and purification. The detection limit falls within the European regulatory limit for infants and children's products (10 µg L-1). The method is rapid, enviroment-friendly, and reliable. Graphical abstractSchematic of the immunoassay.An immunoliposome-patulin-immunomagnetic nanoparticle sandwich complex is formed that can be separated from the sample by magnetic force. The fluorescence of sulforhodamine B carried inside of immunoliposome was measured to determine patulin. The method can detect 8 µg L-1 of patulin in apple juice.
Assuntos
Fluorometria , Imunoensaio , Separação Imunomagnética , Nanopartículas/química , Patulina/análise , Animais , Imunoglobulina G/química , Lipossomos/química , Tamanho da Partícula , Coelhos , Soroalbumina Bovina/química , Propriedades de SuperfícieRESUMO
Nowadays, viruses of foodborne origin such as norovirus and hepatitis A are considered major causes of foodborne gastrointestinal illness with widespread distribution worldwide. A number of foodborne outbreaks associated with food products of animal and non-animal origins, which often involve multiple cases of variety of food streams, have been reported. Although several viruses, including rotavirus, adenovirus, astrovirus, parvovirus, and other enteroviruses, significantly contribute to incidence of gastrointestinal diseases, systematic information on the role of food in transmitting such viruses is limited. Most of the outbreak cases caused by infected food handlers were the source of 53% of total outbreaks. Therefore, prevention and hygiene measures to reduce the frequency of foodborne virus outbreaks should focus on food workers and production site of food products. Pivotal strategies, such as proper investigation, surveillance, and reports on foodborne viral illnesses, are needed in order to develop more accurate measures to detect the presence and pathogenesis of viral infection with detailed descriptions. Moreover, molecular epidemiology and surveillance of food samples may help analysis of public health hazards associated with exposure to foodborne viruses. In this present review, we discuss different aspects of foodborne viral contamination and its impact on human health. This review also aims to improve understanding of foodborne viral infections as major causes of human illness as well as provide descriptions of their control and prevention strategies and rapid detection by advanced molecular techniques. Further, a brief description of methods available for the detection of viruses in food and related matrices is provided.
Assuntos
Microbiologia de Alimentos , Doenças Transmitidas por Alimentos/prevenção & controle , Animais , Surtos de Doenças/prevenção & controle , Manipulação de Alimentos/métodos , Doenças Transmitidas por Alimentos/epidemiologia , Doenças Transmitidas por Alimentos/virologia , Humanos , PrevalênciaRESUMO
Cronobacter species are foodborne pathogens that can affect the human central nervous system. Survivors of Cronobacter infections often suffer from severe neurological impairments, including hydrocephalus, quadriplegia, and developmental delays in all ages, especially in infants and the immunocompromised. Moreover, Cronobacter species pose a high risk in powdered infant formula (PIF) because PIF is a major source of nutrition for infants worldwide. To develop a rapid and sensitive detection method for Cronobacter species in PIF, immunoliposomes and immunomagnetic nanoparticles were synthesized, after which an immunoliposome-based immunomagnetic concentration and separation assay was developed and applied to PIF for the detection of Cronobacter species. The detection limits of the developed assay were 5.9 × 103 ± 0.7-4.8 × 104 ± 0.2 CFU/mL for Cronobacter species in pure culture with no cross-reactivity with 13 other tested non-Cronobacter strains. Additionally, the developed assay could provide results in 3 h when the contaminated level was higher than 104 CFU/25 g PIF and in 9 h when the contaminated level was 10 CFU/25 g PIF. The developed immunoliposome-based immunomagnetic concentration and separation assay is rapid, sensitive, and simple and thus has great potential for use in the detection of Cronobacter species in PIF.
Assuntos
Cronobacter/isolamento & purificação , Contaminação de Alimentos/análise , Separação Imunomagnética/métodos , Fórmulas Infantis/microbiologia , Cronobacter/classificação , Cronobacter/genética , Humanos , Separação Imunomagnética/instrumentação , Fórmulas Infantis/análise , Pós/análiseRESUMO
Lotus (Nelumbo nucifera) root has been used as an edible vegetable in East Asia for thousands of years. The present research was aimed to explore the physicochemical, nutritional and microbiological safety of lotus root fermented sugar syrup as a fermented food supplement or condiment for human health benefits. In this study, the physicochemical, nutritional and microbiological safety properties of lotus root syrup fermented with 57° Brix brown sugar at different time periods until 6 months (180 days) was investigated. There was a significant improvement as compared to 57° Brix brown sugar broth (as a control) in the total acceptability and physicochemical properties of lotus root sugar syrup samples such as pH and color improvement. The red color values of 180 days lotus root fermented sugar syrup samples were significantly enhanced (6.85 ± 0.58) when compared with the control (0.20 ± 0.15). In addition, the total protein content was increased from 8.27 ± 0.86 to 392.33 ± 7.19 µg/mL, along with the increase in fermentation time reaching to the level of consumption acceptability. All the lotus root fermented sugar syrup samples were subjected to microbiological analysis. It was found that the coliform, Bacillus cereus, Escherichia coli, Salmonella and Staphylococcus aureus counts were not detected in majority of the samples, confirming the high degree of hygiene processing of lotus root fermented sugar syrup samples for its use as a food supplement or condiment.
RESUMO
The acute phase (AP) reactant serum amyloid A (SAA), an HDL apolipoprotein, exhibits pro-inflammatory activities, but its physiological function(s) are poorly understood. Functional differences between SAA1.1 and SAA2.1, the two major SAA isoforms, are unclear. Mice deficient in either isoform were used to investigate plasma isoform effects on HDL structure, composition, and apolipoprotein catabolism. Lack of either isoform did not affect the size of HDL, normally enlarged in the AP, and did not significantly change HDL composition. Plasma clearance rates of HDL apolipoproteins were determined using native HDL particles. The fractional clearance rates (FCRs) of apoA-I, apoA-II, and SAA were distinct, indicating that HDL is not cleared as intact particles. The FCRs of SAA1.1 and SAA2.1 in AP mice were similar, suggesting that the selective deposition of SAA1.1 in amyloid plaques is not associated with a difference in the rates of plasma clearance of the isoforms. Although the clearance rate of SAA was reduced in the absence of the HDL receptor, scavenger receptor class B type I (SR-BI), it remained significantly faster compared with that of apoA-I and apoA-II, indicating a relatively minor role of SR-BI in SAA's rapid clearance. These studies enhance our understanding of SAA metabolism and SAA's effects on AP-HDL composition and catabolism.
Assuntos
HDL-Colesterol/metabolismo , Lipoproteínas HDL/sangue , Isoformas de Proteínas/genética , Proteína Amiloide A Sérica/genética , Reação de Fase Aguda/metabolismo , Animais , Apolipoproteína A-I/sangue , Apolipoproteína A-I/química , Apolipoproteína A-II/sangue , Apolipoproteína A-II/química , Apolipoproteína A-II/metabolismo , Humanos , Lipoproteínas HDL/química , Camundongos , Isoformas de Proteínas/química , Receptores Depuradores Classe B/sangue , Receptores Depuradores Classe B/química , Proteína Amiloide A Sérica/química , Proteína Amiloide A Sérica/metabolismoRESUMO
The THO/TREX complex mediates transport of nascent mRNAs from the nucleus towards the cytoplasm in animals, and has a role in small interfering RNA-dependent processes in plants. Here we describe five mutant alleles of Arabidopsis thaliana THO2, which encodes a core subunit of the plant THO/TREX complex. tho2 mutants present strong developmental defects resembling those in plants compromised in microRNA (miRNA) activity. In agreement, not only were the levels of siRNAs reduced in tho2 mutants, but also those of mature miRNAs. As a consequence, a feedback mechanism is triggered, increasing the amount of miRNA precursors, and finally causing accumulation of miRNA-targeted mRNAs. Yeast two-hybrid experiments and confocal microscopy showed that THO2 does not appear to interact with any of the known miRNA biogenesis components, but rather with the splicing machinery, implying an indirect role of THO2 in small RNA biogenesis. Using an RNA immunoprecipitation approach, we found that THO2 interacts with miRNA precursors, and that tho2 mutants fail to recruit such precursors into the miRNA-processing complex, explaining the reduction in miRNA production in this mutant background. We also detected alterations in the splicing pattern of genes encoding serine/arginine-rich proteins in tho2 mutants, supporting a previously unappreciated role of the THO/TREX complex in alternative splicing.
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Proteínas de Arabidopsis/metabolismo , Arabidopsis/genética , MicroRNAs/metabolismo , Proteínas de Ligação a RNA/metabolismo , Processamento Alternativo , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Regulação da Expressão Gênica de Plantas , MicroRNAs/genética , Complexos Multiproteicos/genética , Complexos Multiproteicos/metabolismo , Mutação , Plantas Geneticamente Modificadas , RNA Interferente Pequeno/metabolismo , Proteínas de Ligação a RNA/genéticaRESUMO
Aminoacyl-tRNA synthetases (ARSs) acylate transfer (t)RNAs with amino acids. Charging tRNAs with the right amino acids is the first step in translation; therefore, the accurate and error-free functioning of ARSs is an essential prerequisite for translational fidelity. A recent study found that methionine (Met) can be incorporated into non-Met residues of proteins through methionylation of non-cognate tRNAs under conditions of oxidative stress. However, it was not understood how this mis-methionylation is achieved. Here, we report that methionyl-tRNA synthetase (MRS) is phosphorylated at Ser209 and Ser825 by extracellular signal-related kinase (ERK1/2) under conditions of stress caused by reactive oxygen species (ROS), and that this phosphorylated MRS shows increased affinity for non-cognate tRNAs with lower affinity for tRNA(Met), leading to an increase in Met residues in cellular proteins. The expression of a mutant MRS containing the substitutions S209D and S825D, mimicking dual phosphorylation, reduced ROS levels and cell death. This controlled inaccuracy of MRS seems to serve as a defense mechanism against ROS-mediated damage at the cost of translational fidelity.
Assuntos
Metionina tRNA Ligase/metabolismo , Estresse Oxidativo/fisiologia , Acilação , Células HEK293 , Células HeLa , Humanos , Metionina tRNA Ligase/genética , Estresse Oxidativo/genética , Fosforilação , Biossíntese de Proteínas , RNA de Transferência/genética , RNA de Transferência/metabolismo , Espécies Reativas de Oxigênio/metabolismoRESUMO
This study quantitatively determined contents of amino acids and free sugars in soybean fermented samples (Doenjang) using amino acid autoanalyzer and HPLC, respectively. As a result, hreonine, serine, glycine, alanine and lysine categorized as total sweet amino acids and free sugars (sucrose, maltose, glucose, galactose and fructose) were found in the range of 32.7 to 117.1 and 5.85 to 23.66mg/100 g, respectively. However, average of total sugar content was noted to be 71.63mg%. Three samples possessing the best sensory characteristics were selected for further study. The mean values of four sensory attributes such as sweet, sour, savory and bitter tastes of Doenjang samples were found to be 709.53, 210.25, 241.90 and 276.05. These findings confirm nutritive properties of fermented food products of Doenjang with improved quality and utilization.