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Targeting tumor-infiltrating regulatory T (TI-Treg) cells is a potential strategy for cancer therapy. The ATPase p97 in complex with cofactors (such as Npl4) has been investigated as an antitumor drug target; however, it is unclear whether p97 has a function in immune cells or immunotherapy. Here we show that thonzonium bromide is an inhibitor of the interaction of p97 and Npl4 and that this p97-Npl4 complex has a critical function in TI-Treg cells. Thonzonium bromide boosts antitumor immunity without affecting peripheral Treg cell homeostasis. The p97-Npl4 complex bridges Stat3 with E3 ligases PDLIM2 and PDLIM5, thereby promoting Stat3 degradation and enabling TI-Treg cell development. Collectively, this work shows an important role for the p97-Npl4 complex in controlling Treg-TH17 cell balance in tumors and identifies possible targets for immunotherapy.
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KRAS is one of the most commonly mutated proteins in cancer, and efforts to directly inhibit its function have been continuing for decades. The most successful of these has been the development of covalent allele-specific inhibitors that trap KRAS G12C in its inactive conformation and suppress tumour growth in patients1-7. Whether inactive-state selective inhibition can be used to therapeutically target non-G12C KRAS mutants remains under investigation. Here we report the discovery and characterization of a non-covalent inhibitor that binds preferentially and with high affinity to the inactive state of KRAS while sparing NRAS and HRAS. Although limited to only a few amino acids, the evolutionary divergence in the GTPase domain of RAS isoforms was sufficient to impart orthosteric and allosteric constraints for KRAS selectivity. The inhibitor blocked nucleotide exchange to prevent the activation of wild-type KRAS and a broad range of KRAS mutants, including G12A/C/D/F/V/S, G13C/D, V14I, L19F, Q22K, D33E, Q61H, K117N and A146V/T. Inhibition of downstream signalling and proliferation was restricted to cancer cells harbouring mutant KRAS, and drug treatment suppressed KRAS mutant tumour growth in mice, without having a detrimental effect on animal weight. Our study suggests that most KRAS oncoproteins cycle between an active state and an inactive state in cancer cells and are dependent on nucleotide exchange for activation. Pan-KRAS inhibitors, such as the one described here, have broad therapeutic implications and merit clinical investigation in patients with KRAS-driven cancers.
Assuntos
Neoplasias , Proteínas Proto-Oncogênicas p21(ras) , Transdução de Sinais , Animais , Camundongos , Peso Corporal , Ativação Enzimática , Mutação , Neoplasias/tratamento farmacológico , Neoplasias/genética , Neoplasias/patologia , Nucleotídeos/metabolismo , Proteínas Proto-Oncogênicas p21(ras)/antagonistas & inibidores , Proteínas Proto-Oncogênicas p21(ras)/genética , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Transdução de Sinais/efeitos dos fármacos , Divisão Celular/efeitos dos fármacos , Especificidade por SubstratoRESUMO
Immune responses need to be tightly controlled to avoid excessive inflammation and prevent unwanted host damage. Here we report that germinal center kinase MST4 responded dynamically to bacterial infection and acted as a negative regulator of inflammation. We found that MST4 directly interacted with and phosphorylated the adaptor TRAF6 to prevent its oligomerization and autoubiquitination. Accordingly, MST4 did not inhibit lipopolysaccharide-induced cytokine production in Traf6(-/-) embryonic fibroblasts transfected to express a mutant form of TRAF6 that cannot be phosphorylated at positions 463 and 486 (with substitution of alanine for threonine at those positions). Upon developing septic shock, mice in which MST4 was knocked down showed exacerbated inflammation and reduced survival, whereas heterozygous deletion of Traf6 (Traf6(+/-)) alleviated such deleterious effects. Our findings reveal a mechanism by which TRAF6 is regulated and highlight a role for MST4 in limiting inflammatory responses.
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Inflamação/metabolismo , Fosforilação/fisiologia , Proteínas Serina-Treonina Quinases/metabolismo , Fator 6 Associado a Receptor de TNF/metabolismo , Animais , Células Cultivadas , Citocinas/metabolismo , Feminino , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Técnicas de Silenciamento de Genes , Células HEK293 , Humanos , Inflamação/induzido quimicamente , Lipopolissacarídeos/farmacologia , Masculino , Camundongos , Pessoa de Meia-Idade , Sepse/sangue , Choque Séptico/induzido quimicamente , Choque Séptico/metabolismoRESUMO
Inactive state-selective KRAS(G12C) inhibitors1-8 demonstrate a 30-40% response rate and result in approximately 6-month median progression-free survival in patients with lung cancer9. The genetic basis for resistance to these first-in-class mutant GTPase inhibitors remains under investigation. Here we evaluated matched pre-treatment and post-treatment specimens from 43 patients treated with the KRAS(G12C) inhibitor sotorasib. Multiple treatment-emergent alterations were observed across 27 patients, including alterations in KRAS, NRAS, BRAF, EGFR, FGFR2, MYC and other genes. In preclinical patient-derived xenograft and cell line models, resistance to KRAS(G12C) inhibition was associated with low allele frequency hotspot mutations in KRAS(G12V or G13D), NRAS(Q61K or G13R), MRAS(Q71R) and/or BRAF(G596R), mirroring observations in patients. Single-cell sequencing in an isogenic lineage identified secondary RAS and/or BRAF mutations in the same cells as KRAS(G12C), where they bypassed inhibition without affecting target inactivation. Genetic or pharmacological targeting of ERK signalling intermediates enhanced the antiproliferative effect of G12C inhibitor treatment in models with acquired RAS or BRAF mutations. Our study thus suggests a heterogenous pattern of resistance with multiple subclonal events emerging during G12C inhibitor treatment. A subset of patients in our cohort acquired oncogenic KRAS, NRAS or BRAF mutations, and resistance in this setting may be delayed by co-targeting of ERK signalling intermediates. These findings merit broader evaluation in prospective clinical trials.
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Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Resistencia a Medicamentos Antineoplásicos/genética , Mutação , Neoplasias/tratamento farmacológico , Neoplasias/genética , Proteínas Proto-Oncogênicas p21(ras)/antagonistas & inibidores , Proteínas Proto-Oncogênicas p21(ras)/genética , Acetonitrilas/farmacologia , Animais , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/genética , Linhagem Celular , Estudos de Coortes , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Feminino , GTP Fosfo-Hidrolases/genética , GTP Fosfo-Hidrolases/metabolismo , Humanos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Piperazinas/farmacologia , Piperazinas/uso terapêutico , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas B-raf/metabolismo , Piridinas/farmacologia , Piridinas/uso terapêutico , Pirimidinas/farmacologia , Pirimidinas/uso terapêutico , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
KRAS GTPases are activated in one-third of cancers, and KRAS(G12C) is one of the most common activating alterations in lung adenocarcinoma1,2. KRAS(G12C) inhibitors3,4 are in phase-I clinical trials and early data show partial responses in nearly half of patients with lung cancer. How cancer cells bypass inhibition to prevent maximal response to therapy is not understood. Because KRAS(G12C) cycles between an active and inactive conformation4-6, and the inhibitors bind only to the latter, we tested whether isogenic cell populations respond in a non-uniform manner by studying the effect of treatment at a single-cell resolution. Here we report that, shortly after treatment, some cancer cells are sequestered in a quiescent state with low KRAS activity, whereas others bypass this effect to resume proliferation. This rapid divergent response occurs because some quiescent cells produce new KRAS(G12C) in response to suppressed mitogen-activated protein kinase output. New KRAS(G12C) is maintained in its active, drug-insensitive state by epidermal growth factor receptor and aurora kinase signalling. Cells without these adaptive changes-or cells in which these changes are pharmacologically inhibited-remain sensitive to drug treatment, because new KRAS(G12C) is either not available or exists in its inactive, drug-sensitive state. The direct targeting of KRAS oncoproteins has been a longstanding objective in precision oncology. Our study uncovers a flexible non-uniform fitness mechanism that enables groups of cells within a population to rapidly bypass the effect of treatment. This adaptive process must be overcome if we are to achieve complete and durable responses in the clinic.
Assuntos
Mutação , Proteínas Proto-Oncogênicas p21(ras)/antagonistas & inibidores , Adaptação Biológica , Linhagem Celular Tumoral , Inibidores Enzimáticos/farmacologia , Receptores ErbB/genética , Receptores ErbB/metabolismo , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Proteínas Proto-Oncogênicas p21(ras)/genética , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Transdução de Sinais/efeitos dos fármacosRESUMO
In this paper, a highly sensitive sensor consisting of a silicon nanorod and symmetric rings (SNSR) is presented. Theoretically, three Fano resonances with high Q-factors are excited in the near-infrared range by breaking the symmetry structure based on quasi-bound states in the continuum (Q-BICs). The electromagnetic near-field analysis confirms that the resonances are mainly controlled by toroidal dipole (TD) resonance. The structure is optimized by adjusting different geometrical parameters, and the maximum Q-factor of the Fano resonances can reach 7427. To evaluate the sensing performance of the structure, the sensitivity and the figure of merit (FOM) are calculated by adjusting the environmental refractive index: the maximum sensitivity of 474â nm/RIU and the maximum FOM of 3306 RIU-1. The SNSR can be fabricated by semiconductor-compatible processes, which is experimentally evaluated for changes in transmission spectra at different solution concentrations. The results show that the sensitivity and the Q-factor of the designed metasurface can reach 295â nm/RIU and 850, while the FOM can reach 235 RIU-1. Therefore, the metasurface of SNSR is characterized by high sensitivity and multi-wavelength sensing, which are current research hotspots in the field of optics and can be applied to biomedical sensing and multi-target detection.
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This article shows an all-dielectric metasurface consisting of "H"-shaped silicon disks with tilted splitting gaps, which can detect the temperature and refractive index (RI). By introducing asymmetry parameters that excite the quasi-BIC, there are three distinct Fano resonances with nearly 100% modulation depth, and the maximal quality factor (Q-factor) is over 104. The predominant roles of different electromagnetic excitations in three distinct modes are demonstrated through near-field analysis and multipole decomposition. A numerical analysis of resonance response based on different refractive indices reveals a RI sensitivity of 262 nm/RIU and figure of merit (FOM) of 2183 RIU-1. This sensor can detect temperature fluctuations with a temperature sensitivity of 59.5 pm/k. The proposed metasurface provides a novel method to induce powerful TD resonances and offers possibilities for the design of high-performance sensors.
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InAs/GaAs quantum dot (QD) laser monolithically grown on silicon is one of the potential approaches to realizing silicon-based light sources. However, the mismatch between GaAs and Si generates a high density of threading dislocations (TDs) and antiphase boundaries (APBs), which trap carriers and adversely affect device performance. In this paper, we present a simple method to reduce the threading dislocation density (TDD) merely through GaAs buffer, eliminating the intricate dislocation filter layers (DFLs) as well as any intermediate buffer layers whose compositions are different from the target GaAs. An APB-free epitaxial 2.5â µm GaAs film was grown on exact Si (001) by metalorganic chemical vapor deposition (MOCVD) with a TDD of 9.4 × 106â cm-2. InAs/GaAs QDs with a density of 5.2 × 1010â cm-2 were grown on this GaAs/Si (001) virtual substrate by molecular beam epitaxy (MBE) system. The fabricated QD laser has achieved a single facet room temperature continuous-wave output power of 138â mW with a threshold current density of 397 A/cm2 and a lasing wavelength of 1306â nm. In this work, we propose a simplified method to fabricate high-power QD lasers, which is expected to promote the application of photonic integrated circuits.
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In this paper, an all-dielectric metasurface consisting of a unit cell containing a nanocube array and organized periodically on a silicon dioxide substrate is designed and analyzed. By introducing asymmetric parameters that can excite the quasi-bound states in the continuum, three Fano resonances with high Q-factor and high modulation depth may be produced in the near-infrared range. Three Fano resonance peaks are excited by magnetic dipole and toroidal dipole, respectively, in conjunction with the distributive features of electromagnetism. The simulation results indicate that the discussed structure can be utilized as a refractive index sensor with a sensitivity of around 434â nm/RIU, a maximum Q factor of 3327, and a modulation depth equal to 100%. The proposed structure has been designed and experimentally investigated, and its maximum sensitivity is 227â nm/RIU. At the same time, the modulation depth of the resonance peak at λ = 1185.81â nm is nearly 100% when the polarization angle of the incident light is 0 °. Therefore, the suggested metasurface has applications in optical switches, nonlinear optics, and biological sensors.
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A large bandwidth and high-efficiency subwavelength quarter-wave plate (QWP) is an indispensable component of an integrated miniaturized optical system. The bandwidth of existing plasmonic quarter-wave plates with a transmission efficiency of more than 50% is less than 320 nm in the near-infrared band. In this paper, a metallic quarter-wave plate with a bandwidth of 600 nm (0.95-1.55 µm) and an average transmittance of more than 70% has been designed and shows excellent potential to be used in miniaturized optical polarization detection systems and as an optical data storage device. For TE mode incident waves, this miniaturized optical element can be equivalent to a Fabry-Pérot (FP) resonator. Meanwhile, for the TM mode incident wave, the transmission characteristics of this structure are controlled by gap surface plasmon polaritons (G-SPPs) existing in the symmetric metal/insulator/metal (MIM) configuration.
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Long noncoding RNAs (lncRNAs) play critical roles in the pathogenesis of cardiovascular diseases, especially in myocardial infarction (MI). However, the underlying molecular mechanism of how lncRNA involves and affect MI still remains unclear. This study aimed to investigate the expression of lncRNA growth arrest-specific transcript 5 (GAS5) and its effects on myocardial cells' proliferation, cell cycle, and apoptosis. The possible mechanisms involved in GAS5, calmodulin 2 (CALM2), and microRNA (miR)-525-5p were also explored. The messenger RNA (mRNA) level of CALM2, GAS5, and miR-525-5p in postmyocardial infarction (MI) and normal cells were examined by quantitative real-time polymerase chain reaction (RT-qPCR). Western blot analysis assay was conducted to detect the protein levels of CALM2. The changes of cell cycle/apoptosis and cell viability of post-MI myocardial cells (PMMC) were determined by flow cytometry analysis and MTT (3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide) assay after knockdown of GAS5 or CALM2, respectively. Dual luciferase reporter assay and RNA-binding protein immunoprecipitation (RIP) assay were performed to verify the targeting relationship between miR-525-5p and GAS5, CALM2 in myocardial. Hypoxic preconditioning was performed in normal cells, which constructed a simulated MI environment, and the effect of GAS5 on cardiomyocyte apoptosis was detected. Our data showed that the expression of GAS5 and CALM2 in PMMC was significantly upregulated, while the expression of miR-525-5p was downregulated. Overexpression of GAS5 and CALM2 profoundly promoted the apoptosis of myocardial cell. However, the proliferation of myocardial cell was inhibited by the upregulation of GAS5 and CALM2. Moreover, GAS5 was proved to be the target of miR-525-5p and GAS5 downregulated the expression of miR-525-5p and CALM2. In addition, lncRNA GAS5 promotes MI, while CALM2 induced MI can be reversed by miR-525-5p. These data suggested that lncRNA GAS5 promoted the development and progression of MI via targeting of the miR-525-5p/CALM2 axis and it has the potential to be explored as a therapeutic target for the treatment of MI in the future.
Assuntos
Calmodulina/genética , Regulação da Expressão Gênica , MicroRNAs/genética , Infarto do Miocárdio/genética , RNA Longo não Codificante/genética , Animais , Apoptose/genética , Ciclo Celular/genética , Linhagem Celular , Sobrevivência Celular/genética , Modelos Animais de Doenças , Humanos , Miócitos Cardíacos/citologia , Miócitos Cardíacos/metabolismo , RatosRESUMO
RIG-I is a well-studied sensor of viral RNA that plays a key role in innate immunity. p97 regulates a variety of cellular events such as protein quality control, membrane reassembly, DNA repair, and the cell cycle. Here, we report a new role for p97 with Npl4-Ufd1 as its cofactor in reducing antiviral innate immune responses by facilitating proteasomal degradation of RIG-I. The p97 complex is able to directly bind both non-ubiquitinated RIG-I and the E3 ligase RNF125, promoting K48-linked ubiquitination of RIG-I at residue K181. Viral infection significantly strengthens the interaction between RIG-I and the p97 complex by a conformational change of RIG-I that exposes the CARDs and through K63-linked ubiquitination of these CARDs. Disruption of the p97 complex enhances RIG-I antiviral signaling. Consistently, administration of compounds targeting p97 ATPase activity was shown to inhibit viral replication and protect mice from vesicular stomatitis virus (VSV) infection. Overall, our study uncovered a previously unrecognized role for the p97 complex in protein ubiquitination and revealed the p97 complex as a potential drug target in antiviral therapy.
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Adenosina Trifosfatases/metabolismo , Proteínas Nucleares/metabolismo , Receptores do Ácido Retinoico/metabolismo , Transdução de Sinais , Adenosina Trifosfatases/genética , Animais , Linhagem Celular , Células HeLa , Humanos , Camundongos , Proteínas Nucleares/genética , Complexo de Endopeptidases do Proteassoma/metabolismo , Ligação Proteica/genética , Ligação Proteica/fisiologia , Receptores do Ácido Retinoico/genética , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitinação/fisiologia , Estomatite Vesicular/metabolismo , Estomatite Vesicular/prevenção & controle , Replicação Viral/fisiologiaRESUMO
The large volume variations of tin-based oxides hinder their extensive application in the field of lithium-ion batteries (LIBs). In this study, structure design, hybrid fabrication, and carbon-coating approaches have been simultaneously adopted to address these shortcomings. To this end, uniform mesoporous NiO/SnO2 @rGO, Ni-Sn oxide@rGO, and SnO2 @rGO nanosphere composites have been selectively fabricated. Among them, the obtained NiO/SnO2 @rGO composite exhibited a high capacity of 800â mAh g-1 at 1000â mA g-1 after 400 cycles. The electrochemical mechanism of NiO/SnO2 as an anode for LIBs has been preliminarily investigated by ex situ XRD pattern analysis. Furthermore, an NiO/SnO2 @rGO-LiCoO2 lithium-ion full cell showed a high capacity of 467.8â mAh g-1 at 500â mA g-1 after 100 cycles. Notably, the NiO/SnO2 @rGO composite also showed good performance when investigated as an anode for sodium-ion batteries (SIBs). It is believed that the unique mesoporous nanospherical framework, synergistic effects between the various components, and uniform rGO wrapping of NiO/SnO2 shorten the Li+ ion diffusion pathways, maintain sufficient contact between the active material and the electrolyte, mitigate volume changes, and finally improve the electrical conductivity of the electrode.
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Partial degradation of the p100 subunit to generate p52 subunit is a hallmark of the alternative NF-κB pathway, which has been implicated in cancer. Here, we uncovered a role of the p97-Npl4-Ufd1 complex in mediating p100-to-p52 processing and therefore positively regulating the alternative NF-κB pathway. We observed an elevation of p97 mRNA levels in lymphoma patients, which positively correlates with NFKB2 expression, a downstream target gene of the alternative NF-κB pathway. Moreover, NFKB2 mRNA levels were aberrantly down-regulated in patients with inclusion body myopathy associated with Paget's disease of the bone and frontotemporal dementia (IBMPFD), a disease caused by mutation of p97. Inactivation of p97 or depletion of the p97-Npl4-Ufd1 complex inhibits the processing of p100 into p52, decreasing transcription of the downstream target genes. Further analyses reveal that the p97-Npl4-Ufd1 complex interacts with F-box and WD repeats protein SCF(ßTrCP) complex to regulate the partial degradation of p100, a process involving K48- and K11-linked ubiquitination. In line with this, in LPS-induced lung damage mice model, generation of p52 is significantly decreased in p97-KD mice compared with mock mice. Finally, abrogation of p97 ATPase activity by its specific inhibitor DBeQ, efficiently decreased proliferation of lymphoma cells. Collectively, our study revealed a regulatory role of the p97-Npl4-Ufd1 complex in regulating p100 partial degradation, highlighting the potential of p97 as a drug target for cancers with aberrant activation of the alternative NF-κB pathway.
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Linfócitos/metabolismo , Subunidade p52 de NF-kappa B/metabolismo , Proteínas Nucleares/metabolismo , Pneumonia/metabolismo , Proteínas/metabolismo , beta Carioferinas/metabolismo , Proteínas Adaptadoras de Transporte Vesicular , Animais , Linhagem Celular Tumoral , Retículo Endoplasmático/efeitos dos fármacos , Retículo Endoplasmático/metabolismo , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Regulação da Expressão Gênica , Células HEK293 , Humanos , Peptídeos e Proteínas de Sinalização Intracelular , Lipopolissacarídeos , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Pulmão/patologia , Linfócitos/efeitos dos fármacos , Linfócitos/patologia , Masculino , Camundongos , Camundongos Knockout , Subunidade p52 de NF-kappa B/genética , Proteínas Nucleares/genética , Pneumonia/induzido quimicamente , Pneumonia/genética , Pneumonia/patologia , Complexo de Endopeptidases do Proteassoma/efeitos dos fármacos , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteínas/genética , Proteólise/efeitos dos fármacos , Quinazolinas/farmacologia , Proteínas Ligases SKP Culina F-Box/genética , Proteínas Ligases SKP Culina F-Box/metabolismo , Transdução de Sinais , Transcrição Gênica , Ubiquitinação , beta Carioferinas/antagonistas & inibidores , beta Carioferinas/genéticaRESUMO
In response to viral infection, cytosolic retinoic acid-inducible gene I-like receptors sense viral RNA and promote oligomerization of mitochondrial antiviral signaling protein (MAVS), which then recruits tumor necrosis factor receptor-associated factor (TRAF) family proteins, including TRAF6, to activate an antiviral response. Currently, the interaction between MAVS and TRAF6 is only partially understood, and atomic details are lacking. Here, we demonstrated that MAVS directly interacts with TRAF6 through its potential TRAF6-binding motif 2 (T6BM2; amino acids 455-460). Further, we solved the crystal structure of MAVS T6BM2 in complex with the TRAF6 TRAF_C domain at 2.95 Å resolution. T6BM2 of MAVS binds to the canonical adaptor-binding groove of the TRAF_C domain. Structure-directed mutational analyses in vitro and in cells revealed that MAVS binding to TRAF6 via T6BM2 instead of T6BM1 is essential but not sufficient for an optimal antiviral response. Particularly, a MAVS mutant Y460E retained its TRAF6-binding ability as predicted but showed significantly impaired signaling activity, highlighting the functional importance of this tyrosine. Moreover, these observations were further confirmed in MAVS(-/-) mouse embryonic fibroblast cells. Collectively, our work provides a structural basis for understanding the MAVS-TRAF6 antiviral response.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/química , Fibroblastos/metabolismo , Interações Hospedeiro-Patógeno/genética , Mitocôndrias/metabolismo , Proteínas Recombinantes de Fusão/química , Fator 6 Associado a Receptor de TNF/química , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Sequência de Aminoácidos , Animais , Cristalografia por Raios X , Escherichia coli/genética , Escherichia coli/metabolismo , Fibroblastos/virologia , Expressão Gênica , Genes Reporter , Células HEK293 , Humanos , Luciferases/genética , Luciferases/metabolismo , Camundongos , Camundongos Knockout , Mitocôndrias/virologia , Dados de Sequência Molecular , Mutação , Domínios e Motivos de Interação entre Proteínas , Estrutura Secundária de Proteína , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Vírus Sendai/fisiologia , Alinhamento de Sequência , Transdução de Sinais , Fator 6 Associado a Receptor de TNF/genética , Fator 6 Associado a Receptor de TNF/metabolismoRESUMO
The oxidative stress-responsive 1 (OSR1) kinase belongs to the mammalian STE20-like kinase family. OSR1 is activated by with no lysine [K] (WNKs) kinases, and then it phosphorylates cation-coupled Cl-cotransporters, regulating ion homeostasis and cell volume in mammalian cells. However, the specific mechanisms of OSR1 activation remains poorly defined, largely due to its extremely low basal activity. Here, we dissect in detail the regulatory mechanisms of OSR1 activation from the aspects of autoinhibition, upstream kinase WNK, and the newly identified master regulator mouse protein-25 (MO25). Based on our structural and biochemical studies, we propose a "double lock" model, accounting for the tight autoinhibition of OSR1, an effect that has to be removed by WNK before MO25 further activates OSR1. Particularly, the conserved C-terminal (CCT) domain and αAL helix act together to strongly suppress OSR1 basal activity. WNKs bind to the CCT and trigger its conformational rearrangement to release the kinase domain of OSR1, allowing for MO25 binding and full activation. Finally, the regulatory mechanisms of OSR1 activation were further corroborated by cellular studies of OSR1-regulated cell volume control through WNK-OSR1 signaling pathway. Collectively, these results provide insights into the OSR1 kinase activation to facilitate further functional study.
Assuntos
Proteínas Serina-Treonina Quinases/química , Sequência de Aminoácidos , Proteínas de Ligação ao Cálcio/química , Domínio Catalítico , Tamanho Celular , Ativação Enzimática , Células HEK293 , Homeostase , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/química , Antígenos de Histocompatibilidade Menor , Dados de Sequência Molecular , Ligação Proteica , Multimerização Proteica , Proteínas Serina-Treonina Quinases/fisiologia , Proteína Quinase 1 Deficiente de Lisina WNKRESUMO
The tumor suppressor kinase LKB1 and germinal center kinases (GCKs) are key regulators of various cellular functions. The adaptor molecule MO25 not only recruits and activates LKB1 through the pseudokinase STRAD, but also may directly activate GCKs like MST3, MST4, STK25, OSR1 and SPAK. Targeting MO25 in a pathological setting has been recently studied in mouse. Yet the regulatory mechanism of MO25-mediated kinase activation is not fully understood. Here, our structural studies of MO25-related kinases reveal that MO25 binds to and activates GCK kinases or pseudokinase through a unified structural mechanism, featuring an active conformation of the αC helix and A-loop stabilized by MO25. Compared to GCKs that are directly activated by MO25-binding, activation of LKB1 has evolved additional layer of regulatory machinery, i.e., MO25 "activates" the pseudokinase STRAD, which in turn activates LKB1. Importantly, the structures of MO25α-STK25 and MO25α-MST3 determined in this work represent a transition/intermediate state and a fully activated state, respectively during the MO25-mediated kinase activating process.
Assuntos
Proteínas de Ligação ao Cálcio/genética , Peptídeos e Proteínas de Sinalização Intracelular/genética , Modelos Moleculares , Complexos Multiproteicos/química , Proteínas Serina-Treonina Quinases/genética , Proteínas Recombinantes/genética , Proteínas de Ligação ao Cálcio/química , Proteínas de Ligação ao Cálcio/metabolismo , Clonagem Molecular , Cristalização , Escherichia coli , Vetores Genéticos/genética , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Complexos Multiproteicos/metabolismo , Conformação Proteica , Proteínas Serina-Treonina Quinases/química , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Recombinantes/metabolismoRESUMO
BACKGROUND: Advanced heart failure (AHF) secondary to myxomatous mitral valve disease (MMVD) in dogs has unclear predictive variables and survival time. METHODS: This retrospective study included 38 dogs with AHF and 38 with stable congestive heart failure (CHF), both due to MMVD. Predictive variables for AHF were analysed, and survival times were calculated using logistic regression and the Kaplan-Meier method. RESULTS: Left atrium to aortic root ratio, normalised left ventricular dimension at the end-diastole and end-systole, isovolumic relaxation time (IVRT) and early transmitral inflow velocity to IVRT ratio were associated with AHF progression. The median survival times were significantly longer in the stable group than in the AHF group. After AHF diagnosis, the median survival times for all-cause and cardiogenic mortality were 194 and 354 days, respectively. LIMITATIONS: This was a single-centre retrospective observational study. The study population was small, with breed bias (overrepresentation of Maltese dogs). Additionally, the treatment plans depended on clinical experience. CONCLUSIONS: AHF in dogs with CHF secondary to MMVD is linked to left heart chamber enlargement and increased left ventricular dimensions, significantly reducing survival time to around six months post-diagnosis. Early recognition and appropriate management may improve outcomes, highlighting the importance of advanced treatment strategies.
Assuntos
Doenças do Cão , Insuficiência Cardíaca , Cães , Animais , Doenças do Cão/mortalidade , Insuficiência Cardíaca/veterinária , Insuficiência Cardíaca/mortalidade , Insuficiência Cardíaca/etiologia , Estudos Retrospectivos , Masculino , Feminino , Prognóstico , Insuficiência da Valva Mitral/veterinária , Insuficiência da Valva Mitral/mortalidade , Doenças das Valvas Cardíacas/veterinária , Doenças das Valvas Cardíacas/complicações , Doenças das Valvas Cardíacas/mortalidadeRESUMO
Fano resonance with high Q-factor is considered to play an important role in the field of refractive index sensing. In this paper, we theoretically and experimentally investigate a refractive index sensor with high performance, realizing a new approach to excite multiple Fano resonances of high Q-factor by introducing an asymmetric parameter to generate a quasi-bound state in the continuum (BIC). Combined with the electromagnetic properties, the formation mechanism of Fano resonances in multiple different excitation modes is analyzed and the resonant modes of the three resonant peaks are analyzed as toroidal dipole (TD), magnetic quadrupole (MQ), and magnetic dipole (MD), respectively. The simulation results show that the proposed metastructure has excellent sensing properties with a Q-factor of 3668, sensitivity of 350â nm/RIU, and figure of merit (FOM) of 1000. Furthermore, the metastructure has been fabricated and investigated experimentally, and the result shows that its maximum Q-factor, sensitivity and FOM can reach 634, 233â nm/RIU and 115, respectively. The proposed metastructure is believed to further contribute to the development of biosensors, nonlinear optics, and lasers.
RESUMO
A multi-function sensor based on an all-dielectric metastructure for temperature and refractive index sensing simultaneously is designed and analyzed in this paper. The structure is composed of a periodic array of silicon dimers placed on the silicon dioxide substrate. By breaking the symmetry of the structure, the ideal bound states in the continuum can be converted to the quasi-bound states in the continuum, and three Fano resonances are excited in the near-infrared wavelength. Combining with the electromagnetic field distributions, the resonant modes of three Fano resonances are analyzed as magnetic dipole, magnetic toroidal dipole, and electric toroidal dipole, respectively. The proposed sensor exhibits an impressive maximal Q-factor of 9352, with a modulation depth approaching 100%. Our investigation into temperature and refractive index sensing properties reveals a maximum temperature sensitivity of 60 pm/K. Regarding refractive index sensing, the sensitivity and figure of merit are determined to be 279.5â nm/RIU and 2055.1 RIU-1, respectively. These findings underscore the potential of the all-dielectric metastructure for simultaneous multi-parameter measurements. The sensor's versatility suggests promising applications in biological and chemical sensing.