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BACKGROUND: The ILD-GAP scoring system is known to be useful in predicting prognosis in patients with interstitial lung disease (ILD). An elevated monocyte count was associated with increased risks of IPF poor prognosis. We examined whether the ILD-GAP scoring system combined with the monocyte ratio (ILD-GAPM) is superior to the conventional ILD-GAP model in predicting ILD prognosis. METHODS: In patients with ILD treated between April 2013 and April 2017, we were retrospectively assessed the relationships between baseline clinical parameters, including age, sex, Charlson Comorbidity Index score (CCIS), ILD diagnosis, blood biomarkers, pulmonary function test results, and disease outcomes. In ILD patients were included idiopathic pulmonary fibrosis (IPF), idiopathic nonspecific interstitial pneumonia (iNSIP), collagen vascular disease-related interstitial pneumonia (CVD-IP), chronic hypersensitivity pneumonitis (CHP), and unclassifiable ILD (UC-ILD). We also assessed the ability to predict prognosis was compared between the ILD-GAP and ILD-GAPM models. RESULTS: A total of 179 patients (mean age, 73 years) were assessed. All of them were taken pulmonary function test, including percentage predicted diffusion capacity for carbon monoxide. ILD patients included 56 IPF cases, 112 iNSIP and CVD-IP cases, 6 CHP cases and 5 UC-ILD cases. ILD-GAPM provided a greater area under the receiver-operating characteristic curve (0.747) than ILD-GAP (0.710) for predicting 3-year ILD-related events. Furthermore, the log-rank test showed that the Kaplan-Meier curves in ILD-GAPM were significantly different by stage (P = 0.015), but not by stage in ILD-GAP (P = 0.074). CONCLUSIONS: The ILD-GAPM model may be a more accurate predictor of prognosis for ILD patients than the ILD-GAP model.
Assuntos
Alveolite Alérgica Extrínseca , Doenças Autoimunes , Doenças Cardiovasculares , Fibrose Pulmonar Idiopática , Doenças Pulmonares Intersticiais , Humanos , Idoso , Monócitos , Prognóstico , Estudos Retrospectivos , Doenças Pulmonares Intersticiais/diagnóstico , Fibrose Pulmonar Idiopática/complicações , Fibrose Pulmonar Idiopática/diagnóstico , Alveolite Alérgica Extrínseca/diagnósticoRESUMO
BACKGROUND: For simultaneous prediction of phenotypic drug susceptibility test (pDST) for multiple anti-tuberculosis drugs, the whole genome sequencing (WGS) data can be analyzed using either catalogue-based approach, wherein one causative mutation suggests resistance, (e.g., WHO catalog) or non-catalogue-based approach using complicated algorithm (e.g., TB-profiler, machine learning). The aim was to estimate the predictive ability of WGS-based tests with pDST as the reference, and to compare the two approaches. METHODS: Following the systematic literature search, the diagnostic test accuracies for 14 drugs were pooled using a random-effect bivariate model. RESULTS: Out of 779 articles, 44 articles with 16,821 specimens for meta-analysis and 13 articles not for meta-analysis were adopted. The areas under summary receiver operating characteristic curve suggested "excellent" (0.97-1.00) for 2 drugs (isoniazid 0.975, rifampicin 0.975), "very good" (0.93-0.97) for 8 drugs (pyrazinamide 0.946, streptomycin 0.952, amikacin 0.968, kanamycin 0.963, capreomycin 0.965, para-aminosalicylic acid 0.959, levofloxacin 0.960, ofloxacin 0.958), and "good" (0.75-0.93) for 4 drugs (ethambutol 0.926, moxifloxacin 0.896, ethionamide 0.878, prothionamide 0.908). The non-catalogue-based and catalogue-based approaches had similar ability for all drugs. CONCLUSION: WGS accurately identifies isoniazid and rifampicin resistance. For most drugs, positive WGS results reliably predict pDST positive. The two approaches had similar ability.
RESUMO
BACKGROUND: This study aimed to determine the effectiveness of liquid biopsy in detecting epidermal growth factor receptor (EGFR) mutations at diagnosis, disease progression, and intermediate stages. METHODS: This prospective, multicenter, observational study included 30 patients with non-small cell lung cancer treated with afatinib, harboring a major EGFR mutation confirmed by tumor tissue biopsy. We collected blood samples for liquid biopsy at diagnosis, intermediate stage, and progressive disease. Tissue and liquid biopsies were examined using Cobas ® EGFR Mutation Test v2. RESULTS: Liquid biopsy detected EGFR mutations in 63.6% of the patients at diagnosis. The presence of metastasis in the extrathoracic, brain, and adrenal glands correlated positively with the detection of EGFR mutations. Patients with positive EGFR mutations at diagnosis had significantly shorter overall and progression-free survival than patients with negative EGFR mutations. Four of the 18 patients (22.2%) who reached progressive disease had positive EGFR T790M mutations. Three of 10 patients (30.0%) with progressive disease were positive and negative for T790M using tumor re-biopsy and liquid biopsy, respectively. The results of EGFR mutation by tissue re-biopsy were the same as those of liquid biopsy in the three patients who were positive for significant EGFR mutations but negative for the T790M mutation using liquid biopsy at progressing disease. Only two patients were positive for major EGFR mutations at intermediate levels. CONCLUSIONS: Liquid biopsy can be a prognostic factor in EGFR-tyrosine kinase inhibitor treatments at diagnosis. Tumor re-biopsy can be omitted in patients with positive EGFR mutations by liquid biopsy at PD.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Afatinib/uso terapêutico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/genética , Resistencia a Medicamentos Antineoplásicos/genética , Receptores ErbB/genética , Humanos , Biópsia Líquida/métodos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Mutação , Estudos Prospectivos , Inibidores de Proteínas Quinases/uso terapêuticoRESUMO
Plants adjust to unfavorable conditions by altering physiological activities, such as gene expression. Although previous studies have identified multiple stress-induced genes, the function of many genes during the stress responses remains unclear. Expression of ERD7 (EARLY RESPONSE TO DEHYDRATION 7) is induced in response to dehydration. Here, we show that ERD7 plays essential roles in both plant stress responses and development. In Arabidopsis, ERD7 protein accumulated under various stress conditions, including exposure to low temperature. A triple mutant of Arabidopsis lacking ERD7 and two closely related homologs had an embryonic lethal phenotype, whereas a mutant lacking the two homologs and one ERD7 allele had relatively round leaves, indicating that the ERD7 gene family has essential roles in development. Moreover, the importance of the ERD7 family in stress responses was evidenced by the susceptibility of the mutant lines to cold stress. ERD7 protein was found to bind to several, but not all, negatively charged phospholipids and was associated with membranes. Lipid components and cold-induced reduction in PIP2 in the mutant line were altered relative to wild type. Furthermore, membranes from the mutant line had reduced fluidity. Taken together, ERD7 and its homologs are important for plant stress responses and development and associated with the modification in membrane lipid composition.
Assuntos
Proteínas de Arabidopsis/fisiologia , Arabidopsis/fisiologia , Membrana Celular/metabolismo , Proteínas de Cloroplastos/fisiologia , Resposta ao Choque Frio , Lipídeos de Membrana/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Membrana Celular/química , Proteínas de Cloroplastos/genética , Proteínas de Cloroplastos/metabolismo , Lipídeos de Membrana/análise , Fosfatos de Fosfatidilinositol/metabolismo , Fosfolipídeos/análise , Fosfolipídeos/metabolismoRESUMO
Sucrose non-fermenting 1-related protein kinases (SnRKs) are important for plant growth and stress responses. This family has three clades: SnRK1, SnRK2 and SnRK3. Although plant SnRKs are thought to be activated by upstream kinases, the overall mechanism remains obscure. Geminivirus Rep-Interacting Kinase (GRIK)1 and GRIK2 phosphorylate SnRK1s, which are involved in sugar/energy sensing, and the grik1-1 grik2-1 double mutant shows growth retardation under regular growth conditions. In this study, we established another Arabidopsis mutant line harbouring a different allele of gene GRIK1 (grik1-2 grik2-1) that grows similarly to the wild-type, enabling us to evaluate the function of GRIKs under stress conditions. In the grik1-2 grik2-1 double mutant, phosphorylation of SnRK1.1 was reduced, but not eliminated, suggesting that the grik1-2 mutation is a weak allele. In addition to high sensitivity to glucose, the grik1-2 grik2-1 mutant was sensitive to high salt, indicating that GRIKs are also involved in salinity signalling pathways. Salt Overly Sensitive (SOS)2, a member of the SnRK3 subfamily, is a critical mediator of the response to salinity. GRIK1 phosphorylated SOS2 in vitro, resulting in elevated kinase activity of SOS2. The salt tolerance of sos2 was restored to normal levels by wild-type SOS2, but not by a mutated form of SOS2 lacking the T168 residue phosphorylated by GRIK1. Activation of SOS2 by GRIK1 was also demonstrated in a reconstituted system in yeast. Our results indicate that GRIKs phosphorylate and activate SnRK1 and other members of the SnRK3 family, and that they play important roles in multiple signalling pathways in vivo.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/enzimologia , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais , Arabidopsis/genética , Arabidopsis/fisiologia , Proteínas de Arabidopsis/genética , Regulação da Expressão Gênica de Plantas , Fosforilação , Proteínas Serina-Treonina Quinases/genética , Tolerância ao SalRESUMO
Alternative splicing (AS) of precursor RNAs enhances transcriptome plasticity and proteome diversity in response to diverse growth and stress cues. Recent work has shown that AS is pervasive across plant species, with more than 60% of intron-containing genes producing different isoforms. Mammalian cell-based assays have discovered various inhibitors of AS. Here, we show that the macrolide pladienolide B (PB) inhibits constitutive splicing and AS in plants. Also, our RNA sequencing (RNA-seq) data revealed that PB mimics abiotic stress signals including salt, drought and abscisic acid (ABA). PB activates the abiotic stress- and ABA-responsive reporters RD29A::LUC and MAPKKK18::uidA in Arabidopsis thaliana and mimics the effects of ABA on stomatal aperture. Genome-wide analysis of AS by RNA-seq revealed that PB perturbs the splicing machinery and leads to a striking increase in intron retention and a reduction in other forms of AS. Interestingly, PB treatment activates the ABA signaling pathway by inhibiting the splicing of clade A PP2C phosphatases while still maintaining to some extent the splicing of ABA-activated SnRK2 kinases. Taken together, our data establish PB as an inhibitor and modulator of splicing and a mimic of abiotic stress signals in plants. Thus, PB reveals the molecular underpinnings of the interplay between stress responses, ABA signaling and post-transcriptional regulation in plants.
Assuntos
Arabidopsis/fisiologia , Compostos de Epóxi/farmacologia , Macrolídeos/farmacologia , Splicing de RNA/efeitos dos fármacos , Transdução de Sinais/genética , Estresse Fisiológico/genética , Ácido Abscísico/metabolismo , Ácido Abscísico/farmacologia , Arabidopsis/efeitos dos fármacos , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Secas , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Íntrons , MAP Quinase Quinase Quinases/genética , MAP Quinase Quinase Quinases/metabolismo , Estômatos de Plantas/efeitos dos fármacos , Plantas Geneticamente Modificadas , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Precursores de RNA/metabolismo , RNA Mensageiro/genética , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismoRESUMO
Arabidopsis glycogen synthase kinase 3 (GSK3)-like kinases have versatile functions in plant development and in responding to abiotic stresses. Although physiological evidence suggested a potential role of GSK3-like kinases in abscisic acid (ABA) signaling, the underlying molecular mechanism was largely unknown. Here we identified members of Snf1-related kinase 2s (SnRK2s), SnRK2.2 and SnRK2.3, that can interact with and be phosphorylated by a GSK3-like kinase, brassinosteroid insensitive 2 (BIN2). bin2-3 bil1 bil2, a loss-of-function mutant of BIN2 and its two closest homologs, BIN2 like 1 (BIL1) and BIN2 like 2 (BIL2), was hyposensitive to ABA in primary root inhibition, ABA-responsive gene expression, and phosphorylating ABA Response Element Binding Factor (ABF) 2 fragment by in-gel kinase assays, whereas bin2-1, a gain-of-function mutation of BIN2, was hypersensitive to ABA, suggesting that these GSK3-like kinases function as positive regulators in ABA signaling. Furthermore, BIN2 phosphorylated SnRK2.3 on T180, and SnRK2.3(T180A) had decreased kinase activity in both autophosphorylation and phosphorylating ABFs. Bikinin, a GSK3 kinase inhibitor, inhibited the SnRK2.3 kinase activity and its T180 phosphorylation in vivo. Our genetic analysis further demonstrated that BIN2 regulates ABA signaling downstream of the PYRABACTIN RESISTANCE1/PYR1-LIKE/REGULATORY COMPONENTS OF ABA RECEPTORS receptors and clade A protein phosphatase 2C but relies on SnRK2.2 and SnRK2.3. These findings provide significant insight into the modulation of ABA signaling by Arabidopsis GSK3-like kinases.
Assuntos
Ácido Abscísico/metabolismo , Proteínas de Arabidopsis/metabolismo , Arabidopsis/fisiologia , Quinase 3 da Glicogênio Sintase/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais/fisiologia , Arabidopsis/metabolismo , Fosforilação , Interferência de RNA , Reação em Cadeia da Polimerase em Tempo Real , Espectrometria de Massas em TandemRESUMO
Plants subjected to a prior dehydration stress were seen to have altered transcriptional responses during a subsequent dehydration stress for up to 5 days after the initial stress. The abscisic acid (ABA) inducible RD29B gene of Arabidopsis thaliana was strongly induced after the first stress and displayed transcriptional memory with transcript levels nine-fold higher during the second dehydration stress. These increased transcript levels were due to an increased rate of transcription and are associated with an altered chromatin template during the recovery interval between the dehydration stresses. Here we use a combination of promoter deletion/substitutions, mutants in the trans-acting transcription factors and their upstream protein kinases, and treatments with exogenous ABA or dehydration stress to advance our understanding of the features required for transcriptional memory of RD29B. ABA Response Elements (ABREs) are sufficient to confer transcriptional memory on a minimal promoter, although there is a context effect from flanking sequences. Different mutations in Snf1 Related Protein Kinase 2 (SnRK2) genes positively and negatively affected the response, suggesting that this effect is important for transcriptional memory. Although exogenous ABA treatments could prime transcriptional memory, a second ABA treatment was not sufficient to activate transcriptional memory. Therefore, we concluded that transcriptional memory requires ABA and an ABA-independent factor that is induced or activated by a subsequent dehydration stress and directly or indirectly results in a more active RD29B chromatin template. These results advance our knowledge of the cis- and trans-acting factors that are required for transcriptional memory of RD29B.
Assuntos
Ácido Abscísico/farmacologia , Arabidopsis/fisiologia , Regulação da Expressão Gênica de Plantas , Reguladores de Crescimento de Plantas/farmacologia , Transdução de Sinais , Arabidopsis/citologia , Arabidopsis/efeitos dos fármacos , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Sequência de Bases , Proteínas e Peptídeos de Choque Frio/genética , Proteínas e Peptídeos de Choque Frio/metabolismo , Desidratação , Genes Reporter , Dados de Sequência Molecular , Mutação , Folhas de Planta , Plantas Geneticamente Modificadas , Regiões Promotoras Genéticas/genética , Plântula , Alinhamento de Sequência , Estresse FisiológicoRESUMO
There is no evidence on the precise role of synaptic Zn2+ signaling on the retention and recall of recognition memory. On the basis of the findings that intracellular Zn2+ signaling in the dentate gyrus is required for object recognition, short-term memory, the present study deals with the effect of spatiotemporally blocking Zn2+ signaling in the dentate gyrus after LTP induction and learning. Three-day-maintained LTP was impaired 1 day after injection of clioquinol into the dentate gyrus, which transiently reduced intracellular Zn2+ signaling in the dentate gyrus. The irreversible impairment was rescued not only by co-injection of ZnCl2 , which ameliorated the loss of Zn2+ signaling, but also by pre-injection of Jasplakinolide, a stabilizer of F-actin, prior to clioquinol injection. Simultaneously, 3-day-old space recognition memory was impaired 1 day after injection of clioquinol into the dentate gyrus, but not by pre-injection of Jasplakinolide. Jasplakinolide also rescued both impairments of 3-day-maintained LTP and 3-day-old memory after injection of ZnAF-2DA into the dentate gyrus, which blocked intracellular Zn2+ signaling in the dentate gyrus. The present paper indicates that the blockade and/or loss of intracellular Zn2+ signaling in the dentate gyrus coincidently impair maintained LTP and recognition memory. The mechanism maintaining LTP via intracellular Zn2+ signaling in dentate granule cells, which may be involved in the formation of F-actin, may retain space recognition memory.
Assuntos
Giro Denteado/metabolismo , Potenciação de Longa Duração/fisiologia , Reconhecimento Psicológico/fisiologia , Transdução de Sinais/fisiologia , Zinco/metabolismo , Animais , Antineoplásicos/farmacocinética , Cálcio/metabolismo , Quelantes/farmacologia , Clioquinol/farmacologia , Giro Denteado/efeitos dos fármacos , Depsipeptídeos/farmacologia , Estimulação Elétrica , Etilenodiaminas/farmacologia , Técnicas In Vitro , Potenciação de Longa Duração/efeitos dos fármacos , Masculino , Aprendizagem em Labirinto/efeitos dos fármacos , Técnicas de Patch-Clamp , Piridinas/farmacologia , Ratos , Ratos Wistar , Reconhecimento Psicológico/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , VigíliaRESUMO
With the tremendous progress of the past decades, molecular plant science is becoming more unified than ever. We now have the exciting opportunity to further connect subdisciplines and understand plants as whole organisms, as will be required to efficiently utilize them in natural and agricultural systems to meet human needs. The subfields of photosynthesis, plant developmental biology and plant stress are used as examples to discuss how plant science can become better integrated. The challenges, strategies and rich opportunities for the integration of the plant sciences are discussed. In recent years, more and more overlap between various subdisciplines has been inadvertently discovered including tradeoffs that may occur in plants engineered for biotechnological applications. Already important, bioinformatics and computational modelling will become even more central to structuring and understanding the ever growing amounts of data. The process of integrating and overlapping fields in plant biology research is advancing, but plant science will benefit from dedicating more effort and urgency to reach across its boundaries.
Assuntos
Botânica/tendências , Fotossíntese , Desenvolvimento Vegetal , Plantas/metabolismo , Estresse Fisiológico , Comunicação Celular , Parede Celular/metabolismo , Cloroplastos/metabolismo , Biologia Computacional , Expressão Gênica , Imunidade Vegetal , Madeira/metabolismoRESUMO
The phytohormone abscisic acid (ABA) regulates the expression of many genes in plants; it has critical functions in stress resistance and in growth and development. Several proteins have been reported to function as ABA receptors, and many more are known to be involved in ABA signalling. However, the identities of ABA receptors remain controversial and the mechanism of signalling from perception to downstream gene expression is unclear. Here we show that by combining the recently identified ABA receptor PYR1 with the type 2C protein phosphatase (PP2C) ABI1, the serine/threonine protein kinase SnRK2.6/OST1 and the transcription factor ABF2/AREB1, we can reconstitute ABA-triggered phosphorylation of the transcription factor in vitro. Introduction of these four components into plant protoplasts results in ABA-responsive gene expression. Protoplast and test-tube reconstitution assays were used to test the function of various members of the receptor, protein phosphatase and kinase families. Our results suggest that the default state of the SnRK2 kinases is an autophosphorylated, active state and that the SnRK2 kinases are kept inactive by the PP2Cs through physical interaction and dephosphorylation. We found that in the presence of ABA, the PYR/PYL (pyrabactin resistance 1/PYR1-like) receptor proteins can disrupt the interaction between the SnRK2s and PP2Cs, thus preventing the PP2C-mediated dephosphorylation of the SnRK2s and resulting in the activation of the SnRK2 kinases. Our results reveal new insights into ABA signalling mechanisms and define a minimal set of core components of a complete major ABA signalling pathway.
Assuntos
Ácido Abscísico/fisiologia , Proteínas de Arabidopsis/fisiologia , Arabidopsis/fisiologia , Regulação da Expressão Gênica de Plantas , Transdução de Sinais , Estresse Fisiológico/fisiologia , Arabidopsis/enzimologia , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Mutação , Fenótipo , Fosforilação , Protoplastos/fisiologiaRESUMO
Abscisic acid (ABA) is a ubiquitous hormone that regulates plant growth, development and responses to environmental stresses. Its action is mediated by the PYR/PYL/RCAR family of START proteins, but it remains unclear how these receptors bind ABA and, in turn, how hormone binding leads to inhibition of the downstream type 2C protein phosphatase (PP2C) effectors. Here we report crystal structures of apo and ABA-bound receptors as well as a ternary PYL2-ABA-PP2C complex. The apo receptors contain an open ligand-binding pocket flanked by a gate that closes in response to ABA by way of conformational changes in two highly conserved beta-loops that serve as a gate and latch. Moreover, ABA-induced closure of the gate creates a surface that enables the receptor to dock into and competitively inhibit the PP2C active site. A conserved tryptophan in the PP2C inserts directly between the gate and latch, which functions to further lock the receptor in a closed conformation. Together, our results identify a conserved gate-latch-lock mechanism underlying ABA signalling.
Assuntos
Ácido Abscísico/metabolismo , Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/fisiologia , Arabidopsis/fisiologia , Modelos Moleculares , Transdução de Sinais/fisiologia , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Sítios de Ligação , Análise Mutacional de DNA , Plantas Geneticamente Modificadas , Ligação Proteica , Estrutura Terciária de ProteínaRESUMO
The role of perforant pathway-dentate granule cell synapses in cognitive behavior was examined focusing on synaptic Zn(2+) signaling in the dentate gyrus. Object recognition memory was transiently impaired when extracellular Zn(2+) levels were decreased by injection of clioquinol and N,N,N',N'-tetrakis-(2-pyridylmethyl) ethylendediamine. To pursue the effect of the loss and/or blockade of Zn(2+) signaling in dentate granule cells, ZnAF-2DA (100 pmol, 0.1 mM/1 µl), an intracellular Zn(2+) chelator, was locally injected into the dentate molecular layer of rats. ZnAF-2DA injection, which was estimated to chelate intracellular Zn(2+) signaling only in the dentate gyrus, affected object recognition memory 1 h after training without affecting intracellular Ca(2+) signaling in the dentate molecular layer. In vivo dentate gyrus long-term potentiation (LTP) was affected under the local perfusion of the recording region (the dentate granule cell layer) with 0.1 mM ZnAF-2DA, but not with 1-10 mM CaEDTA, an extracellular Zn(2+) chelator, suggesting that the blockade of intracellular Zn(2+) signaling in dentate granule cells affects dentate gyrus LTP. The present study demonstrates that intracellular Zn(2+) signaling in the dentate gyrus is required for object recognition memory, probably via dentate gyrus LTP expression.
Assuntos
Giro Denteado/fisiologia , Neurônios/fisiologia , Reconhecimento Psicológico/fisiologia , Sinapses/fisiologia , Zinco/metabolismo , Animais , Cálcio/metabolismo , Fármacos do Sistema Nervoso Central/farmacologia , Quelantes/farmacologia , Clioquinol/farmacologia , Giro Denteado/efeitos dos fármacos , Espaço Extracelular/metabolismo , Íons/metabolismo , Potenciação de Longa Duração/efeitos dos fármacos , Potenciação de Longa Duração/fisiologia , Masculino , Neurônios/efeitos dos fármacos , Testes Neuropsicológicos , Ratos Wistar , Reconhecimento Psicológico/efeitos dos fármacos , Transdução de Sinais , Sinapses/efeitos dos fármacos , Técnicas de Cultura de TecidosRESUMO
Brain zinc homeostasis is strictly controlled under healthy conditions, indicating the importance of zinc for physiological function in the brain. A part of zinc in the brain exists in the synaptic vesicles, is released from a subclass of glutamatergic neurons (i.e., zincergic neurons), and serves as a signal factor (Zn(2+) signal) in the intracellular (cytosol) compartment as well as in the extracellular compartment. Zn(2+) signaling is dynamically linked to glutamate signaling and may be involved in synaptic plasticity, such as long-term potentiaion and cognitive activity. In zincergic synapses, intracellular Zn(2+) signaling in the postsynaptic neurons, which is linked to Zn(2+) release from zincergic neuron terminals, plays a role in cognitive activity. When nonzincergic synapses participate in cognition, on the other hand, it is possible that intracellular Zn(2+) signaling, which is due mainly to Zn(2+) release from the internal stores and/or metallothioneins, also is involved in cognitive activity, because zinc-dependent system such as zinc-binding proteins is usually required for cognitive process. Intracellular Zn(2+) dynamics may be modified via an endocrine system activity, glucocorticoid secretion in both zincergic and nonzincergic neurons, which is linked to a long-lasting change in synaptic efficacy. On the basis of the evidence of cognitive decline caused by the lack and/or the blockade of synaptic Zn(2+) signaling, this article summarizes the involvement of intracellular Zn(2+) signaling in zincergic synapses in cognition and a hypothetical involvement of that in nonzincergic synapses.
Assuntos
Cognição/fisiologia , Líquido Intracelular/metabolismo , Transdução de Sinais/fisiologia , Zinco/metabolismo , Animais , HumanosRESUMO
Osmotic stress associated with drought or salinity is a major factor that limits plant productivity. Protein kinases in the SNF1-related protein kinase 2 (SnRK2) family are activated by osmotic stress, suggesting that the kinases are involved in osmotic stress signaling. However, due to functional redundancy, their contribution to osmotic stress responses remained unclear. In this report, we constructed an Arabidopsis line carrying mutations in all 10 members of the SnRK2 family. The decuple mutant snrk2.1/2/3/4/5/6/7/8/9/10 grew poorly under hyperosmotic stress conditions but was similar to the wild type in culture media in the absence of osmotic stress. The mutant was also defective in gene regulation and the accumulation of abscisic acid (ABA), proline, and inositol 1,4,5-trisphosphate under osmotic stress. In addition, analysis of mutants defective in the ABA-activated SnRK2s (snrk2.2/3/6) and mutants defective in the rest of the SnRK2s (snrk2.1/4/5/7/8/9/10) revealed that SnRK2s are a merging point of ABA-dependent and -independent pathways for osmotic stress responses. These results demonstrate critical functions of the SnRK2s in mediating osmotic stress signaling and tolerance.
Assuntos
Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Arabidopsis/genética , Mutação , Proteínas Serina-Treonina Quinases/genética , Ácido Abscísico/farmacologia , Sequência de Aminoácidos , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Ativação Enzimática/efeitos dos fármacos , Perfilação da Expressão Gênica , Inositol 1,4,5-Trifosfato/metabolismo , Dados de Sequência Molecular , Análise de Sequência com Séries de Oligonucleotídeos , Pressão Osmótica/fisiologia , Reguladores de Crescimento de Plantas/farmacologia , Prolina/metabolismo , Proteínas Quinases/genética , Proteínas Quinases/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Homologia de Sequência de Aminoácidos , Transdução de Sinais/fisiologiaRESUMO
The plasma membrane sodium/proton exchanger Salt-Overly-Sensitive 1 (SOS1) is a critical salt tolerance determinant in plants. The SOS2-SOS3 calcium-dependent protein kinase complex up-regulates SOS1 activity, but the mechanistic details of this crucial event remain unresolved. Here we show that SOS1 is maintained in a resting state by a C-terminal auto-inhibitory domain that is the target of SOS2-SOS3. The auto-inhibitory domain interacts intramolecularly with an adjacent domain of SOS1 that is essential for activity. SOS1 is relieved from auto-inhibition upon phosphorylation of the auto-inhibitory domain by SOS2-SOS3. Mutation of the SOS2 phosphorylation and recognition site impeded the activation of SOS1 in vivo and in vitro. Additional amino acid residues critically important for SOS1 activity and regulation were identified in a genetic screen for hypermorphic alleles.
Assuntos
Arabidopsis/metabolismo , Trocadores de Sódio-Hidrogênio/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Mutação , Fosforilação/fisiologia , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Estrutura Terciária de Proteína , Trocadores de Sódio-Hidrogênio/genéticaRESUMO
Protein phosphorylation is one of the most important posttranslational modifications in cell signaling pathways. Kinases and phosphatases play essential roles in transferring information between sensors and effectors under stress conditions. Several methods have been developed to analyze the phosphorylation mechanisms. Each method has advantages and disadvantages. In vitro kinase assay using recombinant proteins is a method to analyze kinase activities under simplified conditions. It is a good strategy to understand each mechanism one by one, although it is not always suitable to estimate the feature of complex machinery in vivo. In this chapter, the purification of recombinant proteins produced in Escherichia coli followed by assaying a kinase activity using radioactivity is described.
Assuntos
Ensaios Enzimáticos , Escherichia coli , Proteínas Serina-Treonina Quinases , Proteínas Recombinantes , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/genética , Escherichia coli/genética , Escherichia coli/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Ensaios Enzimáticos/métodos , Fosforilação , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Estresse Fisiológico , Arabidopsis/genéticaRESUMO
BACKGROUND: Immune checkpoint inhibitors (ICIs) have improved outcomes in cancer treatment but are also associated with adverse events and financial burdens. Identifying accurate biomarkers is crucial for determining which patients are likely to benefit from ICIs. Current markers, such as PD-L1 expression and tumor mutation burden, exhibit limited predictive accuracy. This study utilizes a Clinical Data Warehouse (CDW) to explore the prognostic significance of novel blood-based factors, such as the neutrophil-to-lymphocyte ratio and red cell distribution width (RDW), to enhance the prediction of ICI therapy benefit. METHODS: This retrospective study utilized an exploratory cohort from the CDW that included a variety of cancers to explore factors associated with pembrolizumab treatment duration, validated in a non-small cell lung cancer (NSCLC) patient cohort from electronic medical records (EMR) and CDW. The CDW contained anonymized data on demographics, diagnoses, medications, and tests for cancer patients treated with ICIs between 2017-2022. Logistic regression identified factors predicting ≤2 or ≥5 pembrolizumab doses as proxies for progression-free survival (PFS), and Receiver Operating Characteristic analysis was used to examine their predictive ability. These factors were validated by correlating doses with PFS in the EMR cohort and re-testing their significance in the CDW cohort with other ICIs. This dual approach utilized the CDW for discovery and EMR/CDW cohorts for validating prognostic biomarkers before ICI treatment. RESULTS: A total of 609 cases (428 in the exploratory cohort and 181 in the validation cohort) from CDW and 44 cases from EMR were selected for study. CDW analysis revealed that elevated red cell distribution width (RDW) correlated with receiving ≤2 pembrolizumab doses (p = 0.0008), with an AUC of 0.60 for predicting treatment duration. RDW's correlation with PFS (r = 0.80, p<0.0001) and its weak association with RDW (r = -0.30, p = 0.049) were confirmed in the EMR cohort. RDW also remained significant in predicting short treatment duration across various ICIs (p = 0.0081). This dual methodology verified pretreatment RDW elevation as a prognostic biomarker for shortened ICI therapy. CONCLUSION: This study suggests the utility of CDWs in identifying prognostic biomarkers for ICI therapy in cancer treatment. Elevated RDW before treatment initiation emerged as a potential biomarker of shorter therapy duration.
Assuntos
Anticorpos Monoclonais Humanizados , Índices de Eritrócitos , Inibidores de Checkpoint Imunológico , Humanos , Inibidores de Checkpoint Imunológico/uso terapêutico , Feminino , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Idoso , Anticorpos Monoclonais Humanizados/uso terapêutico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/sangue , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/sangue , Prognóstico , Neutrófilos , Intervalo Livre de Progressão , Biomarcadores Tumorais/sangue , AdultoRESUMO
Benralizumab, a monoclonal antibody targeting IL-5 receptors, reduces exacerbations and oral corticosteroid requirements for severe, uncontrolled eosinophilic asthma. In Japan, geographic disparities in asthma outcomes suggest differential prescribing and access. This study aimed to quantify regional prescribing variations for benralizumab nationwide. Using Japan's National Database (NDB) of insurance claims (2009-2019), benralizumab standardized claim ratios (SCRs) were calculated for 47 prefectures. Correlations between SCRs and other biologics' SCRs, economic variables like average income, and physician densities were evaluated through univariate analysis and multivariate regressions. Income-related barriers to optimal prescribing were examined. Wide variation emerged in benralizumab SCRs, from 40.1 to 184.2 across prefectures. SCRs strongly correlated with omalizumab (r = 0.61, p < 0.00001) and mepolizumab (r = 0.43, p = 0.0024). Average monthly income also positively correlated with benralizumab SCRs (r = 0.45, p = 0.0016), whereas lifestyle factors were insignificant. Respiratory specialist density modestly correlated with SCRs (r = 0.29, p = 0.047). In multivariate regressions, average income remained the most robust predictor (B = 0.74, p = 0.022). Benralizumab SCRs strongly associate with income metrics more than healthcare infrastructure/population factors. Many regions show low SCRs, constituting apparent prescribing gaps. Access barriers for advanced asthma therapies remain inequitable among Japan's income strata. Addressing affordability alongside specialist allocation can achieve better prescribing quality and asthma outcomes.
Assuntos
Antiasmáticos , Anticorpos Monoclonais Humanizados , Asma , Humanos , Asma/tratamento farmacológico , Asma/economia , Japão , Anticorpos Monoclonais Humanizados/uso terapêutico , Anticorpos Monoclonais Humanizados/economia , Masculino , Antiasmáticos/uso terapêutico , Antiasmáticos/economia , Feminino , Pessoa de Meia-Idade , Adulto , Idoso , Padrões de Prática MédicaRESUMO
BACKGROUND: The Guidelines for the Management of Cough and Sputum (2019) of the Japanese Respiratory Society (JRS) were the first internationally published guidelines for the management of sputum. However, the data used to determine the causative diseases of bloody sputum and hemoptysis in these guidelines were not obtained in Japan. METHODS: A retrospective analysis was performed using the clinical information of patients with bloody sputum or hemoptysis who visited the department of respiratory medicine at a university or core hospital in Japan. RESULTS: Included in the study were 556 patients (median age, 73 years; age range, 21-98 years; 302 males (54.3%)). The main causative diseases were bronchiectasis (102 patients (18.3%)), lung cancer (97 patients (17.4%)), and non-tuberculous mycobacterial disease (89 patients (16%)). Sex and age differences were observed in the frequency of causative diseases of bloody sputum and hemoptysis. The most common cause was lung cancer in males (26%), bronchiectasis in females (29%), lung cancer in patients aged <65 years (19%), and bronchiectasis in those aged >65 years (20%). CONCLUSIONS: The present study is the first to investigate the causative diseases of bloody sputum and hemoptysis using data obtained in Japan. When investigating the causative diseases of bloody sputum and hemoptysis, it is important to take the sex and age of the patients into account.