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Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease that results from many diverse genetic causes. Although therapeutics specifically targeting known causal mutations may rescue individual types of ALS, these approaches cannot treat most cases since they have unknown genetic etiology. Thus, there is a pressing need for therapeutic strategies that rescue multiple forms of ALS. Here, we show that pharmacological inhibition of PIKFYVE kinase activates an unconventional protein clearance mechanism involving exocytosis of aggregation-prone proteins. Reducing PIKFYVE activity ameliorates ALS pathology and extends survival of animal models and patient-derived motor neurons representing diverse forms of ALS including C9ORF72, TARDBP, FUS, and sporadic. These findings highlight a potential approach for mitigating ALS pathogenesis that does not require stimulating macroautophagy or the ubiquitin-proteosome system.
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Esclerose Lateral Amiotrófica , Fosfatidilinositol 3-Quinases , Animais , Esclerose Lateral Amiotrófica/tratamento farmacológico , Esclerose Lateral Amiotrófica/genética , Esclerose Lateral Amiotrófica/metabolismo , Neurônios Motores , Mutação , Proteína FUS de Ligação a RNA/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Modelos Animais de DoençasRESUMO
Certain obligate parasites induce complex and substantial phenotypic changes in their hosts in ways that favor their transmission to other trophic levels. However, the mechanisms underlying these changes remain largely unknown. Here we demonstrate how SAP05 protein effectors from insect-vectored plant pathogenic phytoplasmas take control of several plant developmental processes. These effectors simultaneously prolong the host lifespan and induce witches' broom-like proliferations of leaf and sterile shoots, organs colonized by phytoplasmas and vectors. SAP05 acts by mediating the concurrent degradation of SPL and GATA developmental regulators via a process that relies on hijacking the plant ubiquitin receptor RPN10 independent of substrate ubiquitination. RPN10 is highly conserved among eukaryotes, but SAP05 does not bind insect vector RPN10. A two-amino-acid substitution within plant RPN10 generates a functional variant that is resistant to SAP05 activities. Therefore, one effector protein enables obligate parasitic phytoplasmas to induce a plethora of developmental phenotypes in their hosts.
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Arabidopsis/crescimento & desenvolvimento , Arabidopsis/parasitologia , Interações Hospedeiro-Parasita/fisiologia , Parasitos/fisiologia , Proteólise , Ubiquitinas/metabolismo , Sequência de Aminoácidos , Animais , Arabidopsis/genética , Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/metabolismo , Engenharia Genética , Humanos , Insetos/fisiologia , Modelos Biológicos , Fenótipo , Fotoperíodo , Filogenia , Phytoplasma/fisiologia , Desenvolvimento Vegetal , Brotos de Planta/crescimento & desenvolvimento , Plantas Geneticamente Modificadas , Complexo de Endopeptidases do Proteassoma/metabolismo , Estabilidade Proteica , Reprodução , Nicotiana , Fatores de Transcrição/metabolismo , Transcrição GênicaRESUMO
SARS-CoV-2 RNA interacts with host factors to suppress interferon responses and simultaneously induces cytokine release to drive the development of severe coronavirus disease 2019 (COVID-19). However, how SARS-CoV-2 hijacks host RNAs to elicit such imbalanced immune responses remains elusive. Here, we analyzed SARS-CoV-2 RNA in situ structures and interactions in infected cells and patient lung samples using RIC-seq. We discovered that SARS-CoV-2 RNA forms 2,095 potential duplexes with the 3' UTRs of 205 host mRNAs to increase their stability by recruiting RNA-binding protein YBX3 in A549 cells. Disrupting the SARS-CoV-2-to-host RNA duplex or knocking down YBX3 decreased host mRNA stability and reduced viral replication. Among SARS-CoV-2-stabilized host targets, NFKBIZ was crucial for promoting cytokine production and reducing interferon responses, probably contributing to cytokine storm induction. Our study uncovers the crucial roles of RNA-RNA interactions in the immunopathogenesis of RNA viruses such as SARS-CoV-2 and provides valuable host targets for drug development.
Assuntos
COVID-19 , Humanos , COVID-19/genética , SARS-CoV-2/genética , SARS-CoV-2/metabolismo , RNA Viral/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Interferons/genética , CitocinasRESUMO
The coronavirus disease 2019 (COVID-19) pandemic is a global public health crisis. However, little is known about the pathogenesis and biomarkers of COVID-19. Here, we profiled host responses to COVID-19 by performing plasma proteomics of a cohort of COVID-19 patients, including non-survivors and survivors recovered from mild or severe symptoms, and uncovered numerous COVID-19-associated alterations of plasma proteins. We developed a machine-learning-based pipeline to identify 11 proteins as biomarkers and a set of biomarker combinations, which were validated by an independent cohort and accurately distinguished and predicted COVID-19 outcomes. Some of the biomarkers were further validated by enzyme-linked immunosorbent assay (ELISA) using a larger cohort. These markedly altered proteins, including the biomarkers, mediate pathophysiological pathways, such as immune or inflammatory responses, platelet degranulation and coagulation, and metabolism, that likely contribute to the pathogenesis. Our findings provide valuable knowledge about COVID-19 biomarkers and shed light on the pathogenesis and potential therapeutic targets of COVID-19.
Assuntos
Infecções por Coronavirus/sangue , Infecções por Coronavirus/patologia , Plasma/metabolismo , Pneumonia Viral/sangue , Pneumonia Viral/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Betacoronavirus , Biomarcadores/sangue , Proteínas Sanguíneas/metabolismo , COVID-19 , Infecções por Coronavirus/classificação , Infecções por Coronavirus/metabolismo , Feminino , Humanos , Aprendizado de Máquina , Masculino , Pessoa de Meia-Idade , Pandemias/classificação , Pneumonia Viral/classificação , Pneumonia Viral/metabolismo , Proteômica , Reprodutibilidade dos Testes , SARS-CoV-2RESUMO
Neurons in the subthalamic nucleus (STN) become hyperactive following nerve injury and promote pain-related responses in mice. Considering that the anterior cingulate cortex (ACC) is involved in pain and emotion processing and projects to the STN, we hypothesize that ACC neurons may contribute to hyperactivity in STN neurons in chronic pain. In the present study, we showed that ACC neurons enhanced activity in response to noxious stimuli and to alterations in emotional states and became hyperactive in chronic pain state established by spared nerve injury of the sciatic nerve (SNI) in mice. In naïve mice, STN neurons were activated by noxious stimuli, but not by alterations in emotional states. Pain responses in STN neurons were attenuated in both naïve and SNI mice when ACC neurons were inhibited. Furthermore, optogenetic activation of the ACC-STN pathway induced bilateral hyperalgesia and depression-like behaviors in naive mice; conversely, inhibition of this pathway is sufficient to attenuate hyperalgesia and depression-like behaviors in SNI mice and naïve mice subjected to stimulation of STN neurons. Finally, mitigation of pain-like and depression-like behaviors in SNI mice by inhibition of the ACC-STN projection was eliminated by activation of STN neurons. Our results demonstrate that hyperactivity in the ACC-STN pathway may be an important pathophysiology in comorbid chronic pain and depression. Thus, the ACC-STN pathway may be an intervention target for the treatment of the comorbid chronic pain and depression.
Assuntos
Dor Crônica , Camundongos , Masculino , Animais , Giro do Cíngulo/fisiologia , Hiperalgesia , Depressão , Neurônios/fisiologiaRESUMO
The comorbidity of chronic pain and depression poses tremendous challenges for the treatment of either one because they exacerbate each other with unknown mechanisms. As the posterior insular cortex (PIC) integrates multiple somatosensory and emotional information and is implicated in either chronic pain or depression, we hypothesize that the PIC and its projections may contribute to the pathophysiology of comorbid chronic pain and depression. We show that PIC neurons were readily activated by mechanical, thermal, aversive, and stressful and appetitive stimulation in naive and neuropathic pain male mice subjected to spared nerve injury (SNI). Optogenetic activation of PIC neurons induced hyperalgesia and conditioned place aversion in naive mice, whereas inhibition of these neurons led to analgesia, conditioned place preference (CPP), and antidepressant effect in both naive and SNI mice. Combining neuronal tracing, optogenetics, and electrophysiological techniques, we found that the monosynaptic glutamatergic projections from the PIC to the basolateral amygdala (BLA) and the ventromedial nucleus (VM) of the thalamus mimicked PIC neurons in pain modulation in naive mice; in SNI mice, both projections were enhanced accompanied by hyperactivity of PIC, BLA, and VM neurons and inhibition of these projections led to analgesia, CPP, and antidepressant-like effect. The present study suggests that potentiation of the PICâBLA and PICâVM projections may be important pathophysiological bases for hyperalgesia and depression-like behavior in neuropathic pain and reversing the potentiation may be a promising therapeutic strategy for comorbid chronic pain and depression.
Assuntos
Dor Crônica , Neuralgia , Camundongos , Masculino , Animais , Hiperalgesia , Dor Crônica/complicações , Depressão , Córtex Insular , Tonsila do Cerebelo/metabolismo , Neuralgia/metabolismo , Comorbidade , Tálamo , Antidepressivos/uso terapêuticoRESUMO
BACKGROUND: Pulmonary hypertension (PH) is a progressive cardiopulmonary disease with a high mortality rate. Although growing evidence has revealed the importance of dysregulated energetic metabolism in the pathogenesis of PH, the underlying cellular and molecular mechanisms are not fully understood. In this study, we focused on ME1 (malic enzyme 1), a key enzyme linking glycolysis to the tricarboxylic acid cycle. We aimed to determine the role and mechanistic action of ME1 in PH. METHODS: Global and endothelial-specific ME1 knockout mice were used to investigate the role of ME1 in hypoxia- and SU5416/hypoxia (SuHx)-induced PH. Small hairpin RNA and ME1 enzymatic inhibitor (ME1*) were used to study the mechanism of ME1 in pulmonary artery endothelial cells. Downstream key metabolic pathways and mediators of ME1 were identified by metabolomics analysis in vivo and ME1-mediated energetic alterations were examined by Seahorse metabolic analysis in vitro. The pharmacological effect of ME1* on PH treatment was evaluated in PH animal models induced by SuHx. RESULTS: We found that ME1 protein level and enzymatic activity were highly elevated in lung tissues of patients and mice with PH, primarily in vascular endothelial cells. Global knockout of ME1 protected mice from developing hypoxia- or SuHx-induced PH. Endothelial-specific ME1 deletion similarly attenuated pulmonary vascular remodeling and PH development in mice, suggesting a critical role of endothelial ME1 in PH. Mechanistic studies revealed that ME1 inhibition promoted downstream adenosine production and activated A2AR-mediated adenosine signaling, which leads to an increase in nitric oxide generation and a decrease in proinflammatory molecule expression in endothelial cells. ME1 inhibition activated adenosine production in an ATP-dependent manner through regulating malate-aspartate NADH (nicotinamide adenine dinucleotide plus hydrogen) shuttle and thereby balancing oxidative phosphorylation and glycolysis. Pharmacological inactivation of ME1 attenuated the progression of PH in both preventive and therapeutic settings by promoting adenosine production in vivo. CONCLUSIONS: Our findings indicate that ME1 upregulation in endothelial cells plays a causative role in PH development by negatively regulating adenosine production and subsequently dysregulating endothelial functions. Our findings also suggest that ME1 may represent as a novel pharmacological target for upregulating protective adenosine signaling in PH therapy.
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Excess bone loss due to increased osteoclastogenesis is a significant clinical problem. Intraflagellar transport (IFT) proteins have been reported to regulate cell growth and differentiation. The role of IFT80, an IFT complex B protein, in osteoclasts (OCs) is completely unknown. Here, we demonstrate that deletion of IFT80 in the myeloid lineage led to increased OC formation and activity accompanied by severe bone loss in mice. IFT80 regulated OC formation by associating with Casitas B-lineage lymphoma proto-oncogene-b (Cbl-b) to promote protein stabilization and proteasomal degradation of tumor necrosis factor (TNF) receptor-associated factor 6 (TRAF6). IFT80 knockdown resulted in increased ubiquitination of Cbl-b and higher TRAF6 levels, thereby hyperactivating the receptor activator of nuclear factor-κß (NF-κß) ligand (RANKL) signaling axis and increased OC formation. Ectopic overexpression of IFT80 rescued osteolysis in a calvarial model of bone loss. We have thus identified a negative function of IFT80 in OCs.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal , Reabsorção Óssea , Proteínas de Transporte , Osteoclastos , Osteogênese , Proteínas Proto-Oncogênicas c-cbl , Fator 6 Associado a Receptor de TNF , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Reabsorção Óssea/genética , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Diferenciação Celular , Modelos Animais de Doenças , Deleção de Genes , Camundongos , Osteoclastos/citologia , Osteoclastos/metabolismo , Osteogênese/genética , Proteólise , Proteínas Proto-Oncogênicas c-cbl/metabolismo , Ligante RANK/genética , Ligante RANK/metabolismo , Fator 6 Associado a Receptor de TNF/metabolismo , UbiquitinaçãoRESUMO
Conventional Li-ion battery intercalation cathodes leverage charge compensation that is formally associated with redox on the transition metal. Employing the anions in the charge compensation mechanism, so-called anion redox, can yield higher capacities beyond the traditional limitations of intercalation chemistry. Here, we aim to understand the structural considerations that enable anion oxidation and focus on processes that result in structural changes, such as the formation of persulfide bonds. Using a Li-rich metal sulfide as a model system, we present both first-principles simulations and experimental data that show that cation vacancies are required for anion oxidation. First-principles simulations show that the oxidation of sulfide to persulfide only occurs when a neighboring vacancy is present. To experimentally probe the role of vacancies in anion redox processes, we introduce vacancies into the Li2TiS3 phase while maintaining a high valency of Ti. When the cation sublattice is fully occupied and no vacancies can be formed through transition metal oxidation, the material is electrochemically inert. Upon introduction of vacancies, the material can support high degrees of anion redox, even in the absence of transition metal oxidation. The model system offers fundamental insights to deepen our understanding of structure-property relationships that govern reversible anion redox in sulfides and demonstrates that cation vacancies are required for anion oxidation, in which persulfides are formed.
RESUMO
Macrocyclic peptides are promising scaffolds for the covalent ligand discovery. However, platforms enabling the direct identification of covalent macrocyclic ligands in a high-throughput manner are limited. In this study, we present an mRNA display platform allowing selection of covalent macrocyclic inhibitors using 1,3-dibromoacetone-vinyl sulfone (DBA-VS). Testcase selections on TEV protease resulted in potent covalent inhibitors with diverse cyclic structures, among which cTEV6-2, a macrocyclic peptide with a unique C-terminal cyclization, emerged as the most potent covalent inhibitor of TEV protease described to-date. This study outlines the workflow for integrating chemical functionalizationâinstallation of a covalent warheadâwith mRNA display and showcases its application in targeted covalent ligand discovery.
Assuntos
RNA Mensageiro , RNA Mensageiro/antagonistas & inibidores , Ciclização , Sulfetos/química , Sulfetos/farmacologia , Peptídeos Cíclicos/química , Peptídeos Cíclicos/farmacologia , Peptídeos Cíclicos/síntese química , Compostos Macrocíclicos/química , Compostos Macrocíclicos/farmacologia , Compostos Macrocíclicos/síntese química , Sulfonas/química , Sulfonas/farmacologia , Descoberta de Drogas , Inibidores de Proteases/farmacologia , Inibidores de Proteases/química , Inibidores de Proteases/síntese química , Estrutura MolecularRESUMO
Diabetes is closely associated with K+ disturbances during disease progression and treatment. However, it remains unclear whether K+ imbalance occurs in diabetes with normal kidney function. In this study, we examined the effects of dietary K+ intake on systemic K+ balance and renal K+ handling in streptozotocin (STZ)-induced diabetic mice. The control and STZ mice were fed low or high K+ diet for 7 days to investigate the role of dietary K+ intake in renal K+ excretion and K+ homeostasis and to explore the underlying mechanism by evaluating K+ secretion-related transport proteins in distal nephrons. K+-deficient diet caused excessive urinary K+ loss, decreased daily K+ balance, and led to severe hypokalemia in STZ mice compared with control mice. In contrast, STZ mice showed an increased daily K+ balance and elevated plasma K+ level under K+-loading conditions. Dysregulation of the NaCl cotransporter (NCC), epithelial Na+ channel (ENaC), and renal outer medullary K+ channel (ROMK) was observed in diabetic mice fed either low or high K+ diet. Moreover, amiloride treatment reduced urinary K+ excretion and corrected hypokalemia in K+-restricted STZ mice. On the other hand, inhibition of SGLT2 by dapagliflozin promoted urinary K+ excretion and normalized plasma K+ levels in K+-supplemented STZ mice, at least partly by increasing ENaC activity. We conclude that STZ mice exhibited abnormal K+ balance and impaired renal K+ handling under either low or high K+ diet, which could be primarily attributed to the dysfunction of ENaC-dependent renal K+ excretion pathway, despite the possible role of NCC.NEW & NOTEWORTHY Neither low dietary K+ intake nor high dietary K+ intake effectively modulates renal K+ excretion and K+ homeostasis in STZ mice, which is closely related to the abnormality of ENaC expression and activity. SGLT2 inhibitor increases urinary K+ excretion and reduces plasma K+ level in STZ mice under high dietary K+ intake, an effect that may be partly due to the upregulation of ENaC activity.
Assuntos
Diabetes Mellitus Experimental , Canais Epiteliais de Sódio , Potássio na Dieta , Potássio , Animais , Diabetes Mellitus Experimental/metabolismo , Potássio/metabolismo , Potássio/urina , Masculino , Potássio na Dieta/metabolismo , Canais Epiteliais de Sódio/metabolismo , Camundongos Endogâmicos C57BL , Inibidores do Transportador 2 de Sódio-Glicose/farmacologia , Canais de Potássio Corretores do Fluxo de Internalização/metabolismo , Canais de Potássio Corretores do Fluxo de Internalização/genética , Camundongos , Nefropatias Diabéticas/metabolismo , Nefropatias Diabéticas/etiologia , Nefropatias Diabéticas/fisiopatologia , Rim/metabolismo , Rim/efeitos dos fármacos , Rim/fisiopatologia , Hipopotassemia/metabolismo , Amilorida/farmacologia , Eliminação Renal/efeitos dos fármacos , Homeostase , Membro 3 da Família 12 de Carreador de Soluto/metabolismo , Membro 3 da Família 12 de Carreador de Soluto/genética , Glucosídeos/farmacologia , Estreptozocina , Compostos Benzidrílicos , Transportador 2 de Glucose-SódioRESUMO
Autism spectrum disorder (ASD) is thought to result from susceptibility genotypes and environmental risk factors. The offspring of women who experience pregnancy infection have an increased risk for autism. Maternal immune activation (MIA) in pregnant animals produces offspring with autistic behaviors, making MIA a useful model for autism. However, how MIA causes autistic behaviors in offspring is not fully understood. Here, we show that NKCC1 is critical for mediating autistic behaviors in MIA offspring. We confirmed that MIA induced by poly(I:C) infection during pregnancy leads to autistic behaviors in offspring. We further demonstrated that MIA offspring showed significant microglia activation, excessive dendritic spines, and narrow postsynaptic density (PSD) in their prefrontal cortex (PFC). Then, we discovered that these abnormalities may be caused by overexpression of NKCC1 in MIA offspring's PFCs. Finally, we ameliorated the autistic behaviors using PFC microinjection of NKCC1 inhibitor bumetanide (BTN) in MIA offspring. Our findings may shed new light on the pathological mechanisms for autism caused by pregnancy infection.
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BACKGROUND: Circß-catenin, our first reported circRNA, has been reported to mediate tumorigenesis in various cancers. However, its biological functions and underlying mechanisms in colorectal cancer (CRC) remain unknown. METHODS: The qRT-PCR examination was used to detect the expression of circß-catenin, miR-197-3p, and CTNND1 in cells and human tissues. Western blot was conducted to detect the protein expression levels. The biological function of circß-catenin was verified by MTT, colony formation, wound healing, and transwell assays. The in vivo effects of circß-catenin were verified by nude mice xenograft and metastasis models. The regulatory network of circß-catenin/miR-197-3p/CTNND1 was confirmed via dual-luciferase reporter and RIP assays. RESULTS: In the present study, circß-catenin was found to promote CRC cell proliferation and metastasis in vitro and in vivo. Mechanistically, circß-catenin served as miRNA decoy to directly bind to miR-197-3p, then antagonized the repression of the target gene CTNND1, and eventually promoted the malignant phenotype of CRC. More interestingly, the inverted repeated Alu pairs termed AluJb1/2 and AluY facilitated the biogenesis of circß-catenin, which could be partially reversed by EIF4A3 binding to Alu element AluJb2. CONCLUSIONS: Our findings illustrated a novel mechanism of circß-catenin in modulating CRC tumorigenesis and metastasis, which provides a potential therapeutic target for CRC patients.
Assuntos
Proliferação de Células , Neoplasias Colorretais , Progressão da Doença , Fator de Iniciação 4A em Eucariotos , Camundongos Nus , MicroRNAs , RNA Circular , beta Catenina , MicroRNAs/genética , Humanos , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Neoplasias Colorretais/metabolismo , RNA Circular/genética , Animais , Camundongos , beta Catenina/metabolismo , beta Catenina/genética , Proliferação de Células/genética , Fator de Iniciação 4A em Eucariotos/genética , Fator de Iniciação 4A em Eucariotos/metabolismo , delta Catenina , Regulação Neoplásica da Expressão Gênica , Linhagem Celular Tumoral , Masculino , Feminino , Movimento Celular/genética , Camundongos Endogâmicos BALB CRESUMO
This study aimed to explore the expression, function, and mechanisms of TBC1D10B in colon cancer, as well as its potential applications in the diagnosis and treatment of the disease.The expression levels of TBC1D10B in colon cancer were assessed by analyzing the TCGA and CCLE databases. Immunohistochemistry analysis was conducted using tumor and adjacent non-tumor tissues from 68 colon cancer patients. Lentiviral infection techniques were employed to silence and overexpress TBC1D10B in colon cancer cells. The effects on cell proliferation, migration, and invasion were evaluated using CCK-8, EDU, wound healing, and Transwell invasion assays. Additionally, GSEA enrichment analysis was used to explore the association of TBC1D10B with biological pathways related to colon cancer. TBC1D10B was significantly upregulated in colon cancer and closely associated with patient prognosis. Silencing of TBC1D10B notably inhibited proliferation, migration, and invasion of colon cancer cells and promoted apoptosis. Conversely, overexpression of TBC1D10B enhanced these cellular functions. GSEA analysis revealed that TBC1D10B is enriched in the AKT/PI3K/mTOR signaling pathway and highly correlated with PAK4. The high expression of TBC1D10B in colon cancer is associated with poor prognosis. It influences cancer progression by regulating the proliferation, migration, and invasion capabilities of colon cancer cells, potentially acting through the AKT/PI3K/mTOR signaling pathway. These findings provide new targets and therapeutic strategies for the treatment of colon cancer.
Assuntos
Apoptose , Movimento Celular , Proliferação de Células , Neoplasias do Colo , Proteínas Ativadoras de GTPase , Fosfatidilinositol 3-Quinases , Proteínas Proto-Oncogênicas c-akt , Transdução de Sinais , Serina-Treonina Quinases TOR , Quinases Ativadas por p21 , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Apoptose/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Neoplasias do Colo/genética , Neoplasias do Colo/patologia , Neoplasias do Colo/metabolismo , Progressão da Doença , Regulação Neoplásica da Expressão Gênica , Proteínas Ativadoras de GTPase/metabolismo , Proteínas Ativadoras de GTPase/genética , Quinases Ativadas por p21/metabolismo , Quinases Ativadas por p21/genética , Fosfatidilinositol 3-Quinases/metabolismo , Fosfatidilinositol 3-Quinases/genética , Prognóstico , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Serina-Treonina Quinases TOR/metabolismo , Serina-Treonina Quinases TOR/genéticaRESUMO
Covalent organic frameworks (COFs) are attractive adsorbents for sample pretreatment due to their unique structure and properties. However, the selectivity of COFs for the extraction of hazardous compounds is still limited due to the lack of specific interactions between COFs and targets. Herein, we report a pore size adjustment strategy for room-temperature synthesis of molecularly imprinted COF (MICOF) for selective extraction of zearalenone (ZEN) in complex food samples. The three-dimensional building block tetra(4-aminophenyl) methane was used as a functional monomer, while dialdehyde monomers with different numbers of benzene ring were used to adjust the pore size of MICOF to match with the size of ZEN molecules. The prepared MICOF gave the largest adsorption capacity of 177.2 mg g-1 and the highest imprinting factor of 10.1 for ZEN so far. MICOF was used as the adsorbent for dispersed solid-phase extraction in combination with high-performance liquid chromatography for the determination of trace ZEN in cereals. The high selectivity of the developed method allows simple aqueous standard calibration for the matrix effect-free determination of ZEN in food samples. The limit of detection and the recoveries of the developed method were 0.21 µg kg-1 and 93.7-101.4%, respectively. The precision for the determination of ZEN was less than 3.8% (RSD, n = 6). The developed method is promising for the selective determination of ZEN in complex matrices.
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Estruturas Metalorgânicas , Nanosferas , Zearalenona , Estruturas Metalorgânicas/química , Zearalenona/análise , Grão Comestível/química , Temperatura , Cromatografia Líquida de Alta Pressão/métodos , Extração em Fase Sólida/métodos , AdsorçãoRESUMO
The extensive accumulation of polyethylene terephthalate (PET) has become a critical environmental issue. PET hydrolases can break down PET into its building blocks. Recently, we identified a glacial PET hydrolase GlacPETase sharing less than 31% amino acid identity with any known PET hydrolases. In this study, the crystal structure of GlacPETase was determined at 1.8 Å resolution, revealing unique structural features including a distinctive N-terminal disulfide bond and a specific salt bridge network. Site-directed mutagenesis demonstrated that the disruption of the N-terminal disulfide bond did not reduce GlacPETase's thermostability or its catalytic activity on PET. However, mutations in the salt bridges resulted in changes in melting temperature ranging from -8°C to +2°C and the activity on PET ranging from 17.5% to 145.5% compared to the wild type. Molecular dynamics simulations revealed that these salt bridges stabilized the GlacPETase's structure by maintaining their surrounding structure. Phylogenetic analysis indicated that GlacPETase represented a distinct branch within PET hydrolases-like proteins, with the salt bridges and disulfide bonds in this branch being relatively conserved. This research contributed to the improvement of our comprehension of the structural mechanisms that dictate the thermostability of PET hydrolases, highlighting the diverse characteristics and adaptability observed within PET hydrolases.IMPORTANCEThe pervasive problem of polyethylene terephthalate (PET) pollution in various terrestrial and marine environments is widely acknowledged and continues to escalate. PET hydrolases, such as GlacPETase in this study, offered a solution for breaking down PET. Its unique origin and less than 31% identity with any known PET hydrolases have driven us to resolve its structure. Here, we report the correlation between its unique structure and biochemical properties, focusing on an N-terminal disulfide bond and specific salt bridges. Through site-directed mutagenesis experiments and molecular dynamics simulations, the roles of the N-terminal disulfide bond and salt bridges were elucidated in GlacPETase. This research enhanced our understanding of the role of salt bridges in the thermostability of PET hydrolases, providing a valuable reference for the future engineering of PET hydrolases.
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Hidrolases , Polietilenotereftalatos , Polietilenotereftalatos/metabolismo , Filogenia , Estabilidade Enzimática , Hidrolases/metabolismo , Dissulfetos , TemperaturaRESUMO
BACKGROUND: The burden and characteristics of respiratory viral infections in children hospitalized for acute respiratory tract infections (ARTIs) during the post-COVID-19 pandemic era are unclear. We analyzed the epidemiological and clinical characteristics of pediatric patients hospitalized with common respiratory virus infections before and after relaxation of non-pharmaceutical interventions in Hangzhou, China and evaluated the diagnostic value of the six-panel respiratory pathogen detection system. METHODS: Six types of respiratory viruses were detected in respiratory samples from children with suspected ARTIs by multiplex real-time quantitative polymerase chain reaction (RT-qPCR). Changes in virus detection rates and epidemiological and clinical characteristics, obtained from electronic health records, were analyzed. Binary logistic regression was used to identify respiratory tract infections risk factors. Multiplex RT-qPCR and targeted next-generation sequencing results were compared in random samples. RESULTS: Among the 11,056 pediatric samples, 3228 tested positive for one or more of six common respiratory pathogens. RSV and PIV-3 detection rates differed significantly across age groups (both P < 0.001), and were more common in younger children. PIV-1 was more common in infants, toddlers, and preschoolers than in school-age children (P < 0.001). FluB was predominantly detected in school-age children (P < 0.001). RSV-, ADV-, and PIV-1-positivity rates were higher in 2022 than in 2023. Seasonal viral patterns differed across years. RSV (OR 9.156. 95% CI 5.905-14.195) and PIV-3 (OR 1.683, 95% CI 1.133-2.501) were risk factors for lower respiratory tract infections. RSV-positivity was associated with severe pneumonia (P = 0.044). PIV-3 (OR 0.391, 95% CI 0.170-0.899), summer season (OR 1.982, 95% CI 1.117-3.519), and younger age (OR 0.938, 95% CI 0.893-0.986) influenced pneumonia severity. Multiplex RT-qPCR showed good diagnostic performance. CONCLUSION: After changes in COVID-19 prevention and control strategies, six common respiratory viruses in children were prevalent in 2022-2023, with different seasonal epidemic characteristics and age proclivities. RSV and PIV-3 cause lower, and FluA, FluB, and ADV more typically cause upper respiratory tract infections. Infancy and summer season influence severe pneumonia risk. Multiplex RT-qPCR is valuable for accurate and timely detection of respiratory viruses in children, which facilitates management, treatment, and prevention of ARTIs.
Assuntos
Reação em Cadeia da Polimerase Multiplex , Infecções Respiratórias , Humanos , Infecções Respiratórias/epidemiologia , Infecções Respiratórias/virologia , Infecções Respiratórias/diagnóstico , Criança , Pré-Escolar , Lactente , Feminino , Masculino , China/epidemiologia , Adolescente , COVID-19/epidemiologia , COVID-19/diagnóstico , COVID-19/virologia , Recém-Nascido , Vírus/isolamento & purificação , Vírus/genética , Vírus/classificação , Viroses/epidemiologia , Viroses/virologia , Viroses/diagnóstico , SARS-CoV-2/genética , SARS-CoV-2/isolamento & purificação , Hospitalização , Fatores de Risco , Reação em Cadeia da Polimerase em Tempo Real , Sequenciamento de Nucleotídeos em Larga Escala , Estudos Epidemiológicos , Estações do AnoRESUMO
Low-cost sodium ion batteries are of great significance in large-scale energy storage applications. With its high energy density and simple synthesis process, layered transition-metal oxides have become one of the most likely sodium ion battery cathode materials to replace lithium ion batteries in the energy storage market. Here, we report a prilling and MoS2 coating strategy to prepare the spherical cathode material. The spherical micronano particles shorten the diffusion path of Na+, restrain the complexity phase transitions, and enhance the tap density of the materials. In addition, the MoS2 coating improves the electrical conductivity of the material and the structural stability of the cathode material in air. The initial specific discharge capacity is 148.4 mA h g-1 at 0.1 C, which can be maintained at 128.9 mA h g-1 after exposure to air for 10 days. This method dramatically improves the energy density and structural stability of the cathode material, which provides a new scheme for preparing high-performance sodium ion batteries.
RESUMO
Yellow catfish (Pelteobagrus fulvidraco) is an important economic species of freshwater fish, widely distributed in China. Recently, viral diseases of yellow catfish have been identified in Chian (Hubei province), arising more attention to the viral immunity in P. fulvidraco. Tumor necrosis factor (TNF) receptor-associated factor NF-κB activator (TANK)-binding kinase 1 (TBK1) plays an essential role in IFN production and innate antiviral immunity. In the present study, we characterized the P. fulvidraco TBK1 (PfTBK1) and reported its function in interferon response. The full-length open reading frame (ORF) is 2184 bp encoding a protein with 727 amino acids, which is composed of four conserved domains, including KD, ULD, CCD1, and CCD2, similar to TBK1 in other species. Pftbk1 was widely expressed in all detected tissues by qPCR and was not inducible by the spring viremia of carp virus (SVCV), a single-strand RNA virus. In addition, the cellular distribution indicated that PfTBK1 was only located in the cytoplasm. Moreover, PfTBK1 induced strong IFN promoter activities through the Jak-stat pathway, and PfTBK1 interacted with and significantly phosphorylated IFN regulatory factor 3/7 (IRF3/7) in P. fulvidraco, promoting the nuclear translocation of pfIRF3 and PfIRF7, and PfTBK1 upregulated IFN response by PfTBK1-PfIRF3/7 axis. Above all, PfTBK1 triggered IFN response and strongly inhibited the replication of SVCV in EPC cells through induction of IFN downstream IFN-stimulated genes (ISGs). Summarily, this work reveals that PfTBK1 plays a positive regulatory role in IFN induction through the TBK1-IRF3/7 axis, laying a foundation for further exploring the molecular mechanism of the antiviral process in P. fulvidraco.
Assuntos
Peixes-Gato , Interferons , Animais , Interferons/metabolismo , Transdução de Sinais , Fator Regulador 3 de Interferon/genética , Peixes-Gato/genética , Peixes-Gato/metabolismo , Janus Quinases , Fatores de Transcrição STAT , Imunidade Inata/genéticaRESUMO
Grifola frodosa polysaccharides, especially ß-D-glucans, possess significant anti-tumor, antioxidant and immunostimulatory activities. However, the synthesis mechanism remains to be elucidated. A newly discovered glycosyltransferase UGT88A1 was found to extend glucan chains in vitro. However, the role of UGT88A1 in the growth and polysaccharide synthesis of G. frondosa in vivo remains unclear. In this study, the overexpression of UGT88A1 improved mycelial growth, increased polysaccharide production, and decreased cell wall pressure sensitivity. Biomass and polysaccharide production decreased in the silenced strain, and the pressure sensitivity of the cell wall increased. Overexpression and silencing of UGT88A1 both affected the monosaccharide composition and surface morphology of G. frondosa polysaccharides and influenced the antioxidant activity of polysaccharides from different strains. The messenger RNA expression of glucan synthase (GLS), UTP-glucose-1-phosphate uridylyltransferase (UGP), and UDP-xylose-4-epimerase (UXE) related to polysaccharide synthesis, and genes related to cell wall integrity increased in the overexpression strain. Overall, our study indicates that UGT88A1 plays an important role in the growth, stress, and polysaccharide synthesis of G. frondosa, providing a reference for exploring the pathway of polysaccharide synthesis and metabolic regulation. KEY POINTS: â¢UGT88A1 plays an important role in the growth, stress response, and polysaccharide synthesis in G. frondosa. â¢UGT88A1 affected the monosaccharide composition, surface morphology and antioxidant activity of G. frondosa polysaccharides. â¢UGT88A1 regulated the mRNA expression of genes related to polysaccharide synthesis and cell wall integrity.