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1.
Nat Commun ; 12(1): 1678, 2021 03 15.
Artículo en Inglés | MEDLINE | ID: mdl-33723245

RESUMEN

Protein homeostasis is modulated by stress response pathways and its deficiency is a hallmark of aging. The integrated stress response (ISR) is a conserved stress-signaling pathway that tunes mRNA translation via phosphorylation of the translation initiation factor eIF2. ISR activation and translation initiation are finely balanced by eIF2 kinases and by the eIF2 guanine nucleotide exchange factor eIF2B. However, the role of the ISR during aging remains poorly understood. Using a genomic mutagenesis screen for longevity in Caenorhabditis elegans, we define a role of eIF2 modulation in aging. By inhibiting the ISR, dominant mutations in eIF2B enhance protein homeostasis and increase lifespan. Consistently, full ISR inhibition using phosphorylation-defective eIF2α or pharmacological ISR inhibition prolong lifespan. Lifespan extension through impeding the ISR occurs without a reduction in overall protein synthesis. Instead, we observe changes in the translational efficiency of a subset of mRNAs, of which the putative kinase kin-35 is required for lifespan extension. Evidently, lifespan is limited by the ISR and its inhibition may provide an intervention in aging.


Asunto(s)
Longevidad , Mutagénesis , Mutación , Biosíntesis de Proteínas/genética , Animales , Caenorhabditis elegans , Proteínas de Caenorhabditis elegans/genética , Factor 2 Eucariótico de Iniciación/genética , Factor 2 Eucariótico de Iniciación/metabolismo , Factor 2B Eucariótico de Iniciación/metabolismo , Factores de Intercambio de Guanina Nucleótido/metabolismo , Fosforilación , ARN Mensajero , Receptor de Insulina/genética , eIF-2 Quinasa/metabolismo
2.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi ; 37(3): 199-204, 2021 Mar.
Artículo en Chino | MEDLINE | ID: mdl-33766226

RESUMEN

Objective To investigate the effect of insulin-like growth factor 1 (IGF-1) on the phagocytic activity of mouse BV-2 microglial cells. Methods Western blotting was performed to detect the protein levels of IGF-1 and IGF-1 receptor (IGF-1R) in the murine brain after the establishment of acute central nervous system inflammation models by intraperitoneal lipopolysaccharide (LPS) injection (10 mg/kg). The protein level of IGF-1R on BV-2 microglial cells that had been stimulated by 500 ng/mL LPS for 4, 12 and 24 hours was measured by Western blotting. To assess the phagocytic activity of microglial cells in response to IGF-1, BV-2 microglial cells were stimulated by IGF-1 at different concentrations for 24 hours after pretreated with or without wortmannin (PI3K/AKT signaling pathway blocker), and then incubated with fluorescent microbeads for 2 hours followed by measurement of phagocytosis of the fluorescent microbeads by flow cytometry. After treatment of IGF-1 (50 ng/mL), p-AKT and AKT signaling pathways in the BV-2 microglial cells were detected by Western blotting. Results Intraperitoneal LPS injection caused increased levels of IGF-1 and IGF-1R in the mouse brain. LPS upregulated the protein expression of IGF-1R on BV-2 microglial cells. The activity of BV-2 microglial cells to phagocytose fluorescent microbeads gradually increased with IGF-1 concentration rising and peaked in the IGF-1 treatment at 50 ng/mL, and gradually decreased thereafter. And IGF-1 induced the phosphorylation of AKT in BV-2 microglial cells. However, after the PI3K/AKT signaling pathway was blocked via wortmannin, the effect of IGF-1 on the activity of BV-2 microglial cells to phagocytose fluorescent microbeads was significantly alleviated. Conclusion IGF-1 can promote phagocytic activity of BV-2 cells via activating PI3K/AKT signaling pathway, which suggests a potential role of IGF-1 in regulating the cerebral inflammation.


Asunto(s)
Factor I del Crecimiento Similar a la Insulina , Fosfatidilinositol 3-Quinasas , Animales , Factor I del Crecimiento Similar a la Insulina/metabolismo , Ratones , Microglía/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Fosforilación , Proteínas Proto-Oncogénicas c-akt/metabolismo , Receptor IGF Tipo 1 , Transducción de Señal
3.
Nat Commun ; 12(1): 1677, 2021 03 15.
Artículo en Inglés | MEDLINE | ID: mdl-33723253

RESUMEN

The seven 14-3-3 isoforms are highly abundant human proteins encoded by similar yet distinct genes. 14-3-3 proteins recognize phosphorylated motifs within numerous human and viral proteins. Here, we analyze by X-ray crystallography, fluorescence polarization, mutagenesis and fusicoccin-mediated modulation the structural basis and druggability of 14-3-3 binding to four E6 oncoproteins of tumorigenic human papillomaviruses. 14-3-3 isoforms bind variant and mutated phospho-motifs of E6 and unrelated protein RSK1 with different affinities, albeit following an ordered affinity ranking with conserved relative KD ratios. Remarkably, 14-3-3 isoforms obey the same hierarchy when binding to most of their established targets, as supported by literature and a recent human complexome map. This knowledge allows predicting proportions of 14-3-3 isoforms engaged with phosphoproteins in various tissues. Notwithstanding their individual functions, cellular concentrations of 14-3-3 may be collectively adjusted to buffer the strongest phosphorylation outbursts, explaining their expression variations in different tissues and tumors.


Asunto(s)
Proteínas 14-3-3/química , Isoformas de Proteínas , Proteínas 14-3-3/metabolismo , Secuencia de Aminoácidos , Cristalografía por Rayos X , Humanos , Papillomaviridae , Fosfoproteínas , Fosforilación , Unión Proteica , Isoformas de Proteínas/metabolismo
4.
Medicine (Baltimore) ; 100(10): e25124, 2021 Mar 12.
Artículo en Inglés | MEDLINE | ID: mdl-33725911

RESUMEN

ABSTRACT: Although some studies have reported the expression and clinical significance of phosphorylated signal transducer and activator of transcription 3 (p-STAT3) in breast cancer tissues, it is still controversial whether p-STAT3 play a role in promoting or suppressing cancer. Here, we used immunohistochemistry analysis to explore expression of p-STAT3 in 407 cases of breast cancer, and analyzed the relationship between p-STAT3 expression and the clinicopathological characteristics and prognosis of breast cancer patients. Positive p-STAT3 expression was seen in 112 cases (27.5%) of breast cancer. p-STAT3 expression was negatively correlated with tumor size, tumor stage and human epidermal growth factor receptor 2 (HER2) status, and the positive rate of p-STAT3 was lowest in HER2-enriched subtype breast cancer (15.3%), while other subtypes were luminal B (23.0%), luminal A (30.2%), and triple-negative breast cancer (TNBC) (37.5%). Logistic regression model multivariate analysis showed that the independent correlation factor of p-STAT3 expression in breast cancer was tumor size (OR = 0.187, 95% CI = 0.042-0.839, P = .029) and HER2 status (OR = 0.392, 95% CI = 0.216-0.710, P = .002). In this study, no clear relationship was observed between patients' prognosis and expression of p-STAT3. Therefore, we suggest that p-STAT3 expression in breast cancer is negatively correlated with tumor size and HER2 status, but appears to have no effect on survival.


Asunto(s)
Biomarcadores de Tumor/metabolismo , Neoplasias de la Mama/patología , Receptor ErbB-2/metabolismo , Factor de Transcripción STAT3/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores de Tumor/análisis , Mama/patología , Mama/cirugía , Neoplasias de la Mama/mortalidad , China/epidemiología , Supervivencia sin Enfermedad , Femenino , Humanos , Inmunohistoquímica , Estimación de Kaplan-Meier , Mastectomía , Persona de Mediana Edad , Fosforilación , Pronóstico , Receptor ErbB-2/análisis , Factor de Transcripción STAT3/análisis , Análisis de Matrices Tisulares , Carga Tumoral
5.
Science ; 371(6535): 1204-1205, 2021 03 19.
Artículo en Inglés | MEDLINE | ID: mdl-33737475
6.
Adv Exp Med Biol ; 1316: 191-211, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33740251

RESUMEN

Immune checkpoints are essential for the regulation of immune cell functions. Although the abrogation of immunosurveillance of tumor cells is known, the regulators of immune checkpoints are not clear. Lipid metabolism is one of the important metabolic activities in organisms. In lipid metabolism, a large number of metabolites produced can regulate the gene expression and activation of immune checkpoints through various pathways. In addition, increasing evidence has shown that lipid metabolism leads to transient generation or accumulation of toxic lipids that result in endoplasmic reticulum (ER) stress and then regulate the transcriptional and posttranscriptional modifications of immune checkpoints, including transcription, protein folding, phosphorylation, palmitoylation, etc. More importantly, the lipid metabolism can also affect exosome transportation of checkpoints and the degradation of checkpoints by affecting ubiquitination and lysosomal trafficking. In this chapter, we mainly empathize on the roles of lipid metabolism in the regulation of immune checkpoints, such as gene expression, activation, and degradation.


Asunto(s)
Metabolismo de los Lípidos , Respuesta de Proteína Desplegada , Estrés del Retículo Endoplásmico , Metabolismo de los Lípidos/genética , Fosforilación , Pliegue de Proteína
7.
Ecotoxicol Environ Saf ; 214: 112058, 2021 May.
Artículo en Inglés | MEDLINE | ID: mdl-33714136

RESUMEN

Nuclear factor erythroid 2-related factor 2 (Nrf2) is a nuclear transcription factor of great concern which is widely involved in physiological and pathological processes of the organism, but the role and regulatory mechanism of Nrf2 in kidney exposed to cadmium (Cd) remain largely unknown. Here we demonstrated that Cd exposure induced injury in primary rat proximal tubular (rPT) cells and NRK-52E cell line, which was accompanied by autophagic flux blockade and subsequent accumulation of p62. Cd-activated nucleus translocation of Nrf2 depended on p62, which promoted antioxidant genes transcription, but it failed to against Cd-induced cell injury and ultimately succumbed to Cd toxicity. CDDO Methyl Ester (CDDO-ME) or ML385 treatment aggravated or alleviated rPT cells injury induced by Cd respectively, indicating that Nrf2 nucleus translocation played a negative role during Cd-induced rPT cells injury. Phosphorylation of 5' AMP-activated protein kinase (AMPK) decreased together with enhanced Nrf2 nucleus translocation in rPT cells exposed to Cd. Dephosphorylation of AMPK induced by Cd were facilitated or restored by CDDO-ME or ML385 treatment, which confirmed AMPK is a downstream factor of Nrf2. Simultaneously, CDDO-ME further enhanced Phosphorylation of mTOR and AKT which increased during Cd exposure. While, Cd-induced phosphorylation of mTOR and AKT were reversed by ML385 treatment. These results illustrated that Cd mediated Nrf2 nucleus translocation depends on p62 accumulation which results from autophagic flux inhibition. The enhanced nucleus translocation of Nrf2 suppresses phosphorylation of AMPK to inactivate AKT/mTOR signaling, and results in rPT cells injury finally.


Asunto(s)
Cadmio/toxicidad , Contaminantes Ambientales/toxicidad , Proteínas Quinasas Activadas por AMP/metabolismo , Animales , Antioxidantes/metabolismo , Autofagia/efectos de los fármacos , Núcleo Celular/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Túbulos Renales Proximales/citología , Túbulos Renales Proximales/metabolismo , Túbulos Renales Proximales/patología , Factor 2 Relacionado con NF-E2/metabolismo , Estrés Oxidativo/efectos de los fármacos , Fosforilación , Proteínas Proto-Oncogénicas c-akt/metabolismo , Ratas , Transducción de Señal/efectos de los fármacos , Serina-Treonina Quinasas TOR/metabolismo
8.
Medicine (Baltimore) ; 100(12): e25093, 2021 Mar 26.
Artículo en Inglés | MEDLINE | ID: mdl-33761671

RESUMEN

ABSTRACT: Based on the Thompson classification of intervertebral discs (IVDs), we systematically analyzed gene expression differences between severely degenerated and mildly degenerated IVDs and explored the underlying molecular mechanisms using bioinformatics methods and multichip integration. We used multiomics analysis, includes mRNA microarray and methylation chips, to explore the genetic network and mechanisms of lumbar disc herniation (LDH). Subsequently, the Combat function of the R language SVA package was applied to eliminate heterogeneity between the gene expression data. And the protein-protein interaction (PPI) network, gene ontology (GO), and molecular pathways were used to constructs the mechanisms network. Consequently, we obtained 149 differentially expressed genes. Related molecular pathways are the following: ribosome activity, oxidative phosphorylation, extracellular matrix response. Besides, through PPI network analysis, genes with higher connectivity such as UBA52, RPLP0, RPL3, RPLP2, and RPL27 were also identified, suggesting that they play important regulatory roles in the complex network associated with LDH. Additionally, cg12556991 (RPL27) and cg06852319 (RPLP0) were found to be LDH-related candidate DNA methylation modification sites in the IVDs tissue of LDH patients. In conclusions, ribosome activity, oxidative phosphorylation, and extracellular matrix response may be potential molecular mechanisms underlying LDH, while hub genes involved in UBA52, RPLP0, RPL3, RPLP2, and RPL27, and candidate DNA methylation modification sites of cg12556991and cg06852319 are likely key regulators in the development of LDH.


Asunto(s)
Metilación de ADN/genética , Matriz Extracelular/genética , Desplazamiento del Disco Intervertebral/genética , Fosforilación/genética , Proteínas Ribosómicas/genética , Biología Computacional , Expresión Génica/genética , Perfilación de la Expresión Génica , Ontología de Genes , Redes Reguladoras de Genes/genética , Humanos , Vértebras Lumbares/metabolismo , Análisis por Micromatrices , Mapas de Interacción de Proteínas/genética , ARN Mensajero/metabolismo
9.
Nat Commun ; 12(1): 1472, 2021 03 05.
Artículo en Inglés | MEDLINE | ID: mdl-33674566

RESUMEN

Modules that switch protein-protein interactions on and off are essential to develop synthetic biology; for example, to construct orthogonal signaling pathways, to control artificial protein structures dynamically, and for protein localization in cells or protocells. In nature, the E. coli MinCDE system couples nucleotide-dependent switching of MinD dimerization to membrane targeting to trigger spatiotemporal pattern formation. Here we present a de novo peptide-based molecular switch that toggles reversibly between monomer and dimer in response to phosphorylation and dephosphorylation. In combination with other modules, we construct fusion proteins that couple switching to lipid-membrane targeting by: (i) tethering a 'cargo' molecule reversibly to a permanent membrane 'anchor'; and (ii) creating a 'membrane-avidity switch' that mimics the MinD system but operates by reversible phosphorylation. These minimal, de novo molecular switches have potential applications for introducing dynamic processes into designed and engineered proteins to augment functions in living cells and add functionality to protocells.


Asunto(s)
Membrana Celular/metabolismo , Escherichia coli/metabolismo , Péptidos/metabolismo , Dimerización , Escherichia coli/genética , Cinética , Fosforilación , Ingeniería de Proteínas , Transducción de Señal , Biología Sintética
10.
Nat Commun ; 12(1): 1756, 2021 03 25.
Artículo en Inglés | MEDLINE | ID: mdl-33767161

RESUMEN

The levels of nuclear protein Lamin A/C are crucial for nuclear mechanotransduction. Lamin A/C levels are known to scale with tissue stiffness and extracellular matrix levels in mesenchymal tissues. But in epithelial tissues, where cells lack a strong interaction with the extracellular matrix, it is unclear how Lamin A/C is regulated. Here, we show in epithelial tissues that Lamin A/C levels scale with apico-basal cell compression, independent of tissue stiffness. Using genetic perturbations in Drosophila epithelial tissues, we show that apico-basal cell compression regulates the levels of Lamin A/C by deforming the nucleus. Further, in mammalian epithelial cells, we show that nuclear deformation regulates Lamin A/C levels by modulating the levels of phosphorylation of Lamin A/C at Serine 22, a target for Lamin A/C degradation. Taken together, our results reveal a mechanism of Lamin A/C regulation which could provide key insights for understanding nuclear mechanotransduction in epithelial tissues.


Asunto(s)
Núcleo Celular/fisiología , Proteínas de Drosophila/metabolismo , Lamina Tipo A/metabolismo , Laminas/metabolismo , Mecanotransducción Celular/fisiología , Estrés Mecánico , Animales , Línea Celular , Perros , Drosophila , Proteínas de Drosophila/genética , Epitelio/metabolismo , Lamina Tipo A/genética , Laminas/genética , Células de Riñón Canino Madin Darby , Fosforilación
11.
Nat Commun ; 12(1): 1863, 2021 03 25.
Artículo en Inglés | MEDLINE | ID: mdl-33767186

RESUMEN

Embryonic stem cells (ESCs) can be maintained in the naïve state through inhibition of Mek1/2 and Gsk3 (2i). A relevant effect of 2i is the inhibition of Cdk8/19, which are negative regulators of the Mediator complex, responsible for the activity of enhancers. Inhibition of Cdk8/19 (Cdk8/19i) stimulates enhancers and, similar to 2i, stabilizes ESCs in the naïve state. Here, we use mass spectrometry to describe the molecular events (phosphoproteome, proteome, and metabolome) triggered by 2i and Cdk8/19i on ESCs. Our data reveal widespread commonalities between these two treatments, suggesting overlapping processes. We find that post-transcriptional de-repression by both 2i and Cdk8/19i might support the mitochondrial capacity of naive cells. However, proteome reprogramming in each treatment is achieved by different mechanisms. Cdk8/19i acts directly on the transcriptional machinery, activating key identity genes to promote the naïve program. In contrast, 2i stabilizes the naïve circuitry through, in part, de-phosphorylation of downstream transcriptional effectors.


Asunto(s)
Quinasa 8 Dependiente de Ciclina/antagonistas & inhibidores , Quinasas Ciclina-Dependientes/antagonistas & inhibidores , Glucógeno Sintasa Quinasa 3/antagonistas & inhibidores , MAP Quinasa Quinasa 2/antagonistas & inhibidores , Células Madre Embrionarias de Ratones/citología , Células Madre Pluripotentes/citología , Animales , Benzamidas/farmacología , Línea Celular , Difenilamina/análogos & derivados , Difenilamina/farmacología , MAP Quinasa Quinasa 1/antagonistas & inhibidores , Espectrometría de Masas , Ratones , Ratones Endogámicos C57BL , Mitocondrias/metabolismo , Fosforilación/fisiología , Transcripción Genética/efectos de los fármacos , Transcripción Genética/genética
12.
Chin J Physiol ; 64(1): 43-50, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33642343

RESUMEN

Focal adhesion (FA) turnover has been demonstrated to play an important role in cell migration; however, the mechanism of FA turnover is complicated and requires further investigation. In this study, Rab11, which is involved in endosome recycling, was examined in terms of a direct regulatory function in FA formation during cell migration. Wild-type and dominant negative (DN) Rab11 or shRab11 were transfected into human HT1080 fibrosarcoma cells; the cell motility and migration abilities were determined, and localization of Rab11 and FA molecules was monitored by confocal microscopy. The results showed that Rab11 deficiency or the DN form inhibited sarcoma cell migration. Rab11 was also found to be co-localized with recycled ß1 integrin and affected FA formation. We further employed immunofluorescence and immunoprecipitation to examine the physical interaction between Rab11 and focal adhesion kinase (FAK), and the results suggested that Rab11 affected cell migration by regulating FAK recycling to aid formation of an FA complex on the cell membrane.


Asunto(s)
Adhesiones Focales , Sarcoma , Adhesión Celular , Movimiento Celular , Proteína-Tirosina Quinasas de Adhesión Focal/metabolismo , Adhesiones Focales/metabolismo , Humanos , Fosforilación , Sarcoma/metabolismo
13.
Hua Xi Kou Qiang Yi Xue Za Zhi ; 39(1): 88-93, 2021 Feb 01.
Artículo en Inglés, Chino | MEDLINE | ID: mdl-33723942

RESUMEN

Porphyromonas gingivalis (P. gingivalis), a Gram-negative oral anaerobe, is considered to be a major pathogenic agent involved in the onset and progression of chronic periodontitis. P. gingivalis must be able to perceive and respond to the complicated changes in host to survive the environmental challenges, in which the two-component signal transduction systems (TCSs) play critical roles by connecting input signals to cellular physiological output. Canonical TCS consists of a sensor histidine kinase and a cognate response regulator that functions via a phosphorylation cascade. In this review, the roles of TCSs in P. gingivalis were demonstrated by illustrating the target genes and modulation modes, which may help elucidate the underlying mechanisms in future studies.


Asunto(s)
Porphyromonas gingivalis , Transducción de Señal , Fosforilación
14.
Sci Total Environ ; 770: 144727, 2021 May 20.
Artículo en Inglés | MEDLINE | ID: mdl-33736362

RESUMEN

Melamine poisoning incidents and potential health risks raise global attention. Recent studies imply that melamine exposure is related to male reproductive dysfunction, however, the underlying mechanisms are unclear. In this study, 32 male Kunming mice were administered with 0, 12.5, 25, and 50 mg/L melamine via drinking water for 13 weeks, respectively. Sperm quality, testicular morphology, and the mRNA expression levels of MAPK family members p38, ERK5, ERK1/2, JNK1/2/3 and their downstream transcription factors GADD153, MAX, MEF2C, CREB, c-Myc, JunD, c-JUN, Sap1a, p53, ATF-2, Elk1, and Nur77 in testes were investigated. The results revealed that low-dose melamine exposure reduced sperm quality, altered the testicular histological structure, and reduced the mRNA expression levels of p38, ERK1/2, MAX and Sap1a in the testes. The p38 and phosphorylated-p38 expressions analysis further suggested that the down-regulated phosphorylation of p38 and downstream transcription factors MAX and Sap1a play key roles in male reproductive dysfunction caused by melamine. Altogether, our study provides a new insight to elucidate the underlying mechanisms by which melamine induces male reproductive toxicity, and to evaluate the health risks of melamine.


Asunto(s)
Testículo , Factores de Transcripción , Animales , Masculino , Ratones , Fosforilación , Testículo/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Triazinas/metabolismo , Triazinas/toxicidad
15.
Sci Signal ; 14(673)2021 03 09.
Artículo en Inglés | MEDLINE | ID: mdl-33688079

RESUMEN

IL-1ß is a key mediator of the cytokine storm linked to high morbidity and mortality from COVID-19, and IL-1ß blockade with anakinra and canakinumab during COVID-19 infection has entered clinical trials. Using mass cytometry of human peripheral blood mononuclear cells, we identified effector memory CD4+ T cells and CD4-CD8low/-CD161+ T cells, specifically those positive for the chemokine receptor CCR6, as the circulating immune subtypes with the greatest response to IL-1ß. This response manifested as increased phosphorylation and, thus, activation of the proinflammatory transcription factor NF-κB and was also seen in other subsets, including CD11c+ myeloid dendritic cells, classical monocytes, two subsets of natural killer cells (CD16-CD56brightCD161- and CD16-CD56dimCD161+), and lineage- (Lin-) cells expressing CD161 and CD25. IL-1ß also induced a rapid but less robust increase in the phosphorylation of the kinase p38 as compared to that of NF-κB in most of these immune cell subsets. Prolonged IL-1ß stimulation increased the phosphorylation of the transcription factor STAT3 and to a lesser extent that of STAT1 and STAT5 across various immune cell types. IL-1ß-induced production of IL-6 likely led to the activation of STAT1 and STAT3 at later time points. Interindividual heterogeneity and inhibition of STAT activation by anakinra raise the possibility that assays measuring NF-κB phosphorylation in response to IL-1ß in CCR6+ T cell subtypes could identify those patients at higher risk of cytokine storm and most likely to benefit from IL-1ß-neutralizing therapies.


Asunto(s)
/inmunología , Interleucina-1beta/sangre , Subgrupos de Linfocitos T/inmunología , /sangre , Síndrome de Liberación de Citoquinas/sangre , Síndrome de Liberación de Citoquinas/etiología , Síndrome de Liberación de Citoquinas/inmunología , Células Dendríticas/inmunología , Células Dendríticas/metabolismo , Citometría de Flujo , Humanos , Interleucina-1beta/farmacología , Células Asesinas Naturales/inmunología , Células Asesinas Naturales/metabolismo , Monocitos/clasificación , Monocitos/inmunología , Monocitos/metabolismo , FN-kappa B/sangre , Pandemias , Fosforilación , Receptores CCR6/sangre , Factores de Transcripción STAT/sangre , Factores de Transcripción STAT/inmunología , Transducción de Señal/inmunología , Subgrupos de Linfocitos T/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/sangre
16.
Int J Mol Med ; 47(4)2021 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-33649779

RESUMEN

Oxidative stress serves a key role in doxorubicin (DOX)­induced cardiotoxicity. The peptide Szeto­Schiller (SS)31 is an efficacious antioxidant with the capacity to reduce mitochondrial reactive oxygen species (ROS) levels and scavenge free radicals. Although SS31 is involved in the pathophysiological process of various cardiovascular diseases, the role of SS31 in DOX­induced cardiotoxicity remains unclear. To explore the effects of SS31 in DOX­induced cardiotoxicity, the present study first constructed DOX­induced cardiotoxicity models, in which H9c2 cells were incubated with 1 µM DOX for 24 h and C57BL/6 mice were administered DOX (20 mg/kg cumulative dose). The results of various assays in these models demonstrated that SS31 exhibited a cardioprotective effect in vitro and in vivo by attenuating the level of ROS, stabilizing the mitochondrial membrane potential and ameliorating myocardial apoptosis as well as fibrosis following treatment with DOX. Mechanistically, the results of the present study revealed that the p38 MAPK signaling pathway was inhibited by SS31 in DOX­treated H9c2 cells, which was associated with the cardioprotective function of SS31. In addition, P79350, a selective agonist of p38 MAPK, reversed the protective effects of SS31. Taken together, these results demonstrated the effects of SS31 on ameliorating DOX­induced cardiotoxicity and indicated its potential as a drug for the treatment of DOX­induced cardiotoxicity.


Asunto(s)
Cardiotónicos/uso terapéutico , Doxorrubicina/toxicidad , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Miocardio/patología , Oligopéptidos/uso terapéutico , Proteínas Quinasas p38 Activadas por Mitógenos/antagonistas & inhibidores , Animales , Antioxidantes/uso terapéutico , Apoptosis/efectos de los fármacos , Cardiotoxicidad/tratamiento farmacológico , Cardiotoxicidad/prevención & control , Fibrosis Endomiocárdica/tratamiento farmacológico , Fibrosis Endomiocárdica/prevención & control , Masculino , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Ratones , Ratones Endogámicos C57BL , Mitocondrias/metabolismo , Estrés Oxidativo/efectos de los fármacos , Fosforilación/efectos de los fármacos , Ratas , Especies Reactivas de Oxígeno/metabolismo
18.
Molecules ; 26(5)2021 Feb 26.
Artículo en Inglés | MEDLINE | ID: mdl-33652602

RESUMEN

Hepatitis B virus (HBV) is a circular, and partially double-stranded DNA virus. Upon infection, the viral genome is translocated into the cell nucleus, generating the covalently closed circular DNA (cccDNA) intermediate, and forming a mini chromosome. HBV HBx is a small protein displaying multiple roles in HBV-infected cells, and in different subcellular locations. In the nucleus, the HBx protein is required to initiate and maintain viral transcription from the viral mini chromosome. In contrast, HBx also functions in the cytoplasm, where it is able to alter multiple cellular functions such as mitochondria metabolism, apoptosis and signal transduction pathways. It has been reported that in cultured cells, at low expression levels, the HBx protein is localized in the nucleus, whereas at high expression levels, it accumulates in the cytoplasm. This dynamic subcellular distribution of HBx might be essential to exert its multiple roles during viral infection. However, the mechanism that regulates different subcellular localizations of the HBx protein is unknown. We have previously taken a bioinformatics approach to investigate whether HBx might be regulated via post-translational modification, and we have proposed that the multiple nucleocytoplasmic functions of HBx might be regulated by an evolutionarily conserved mechanism via phosphorylation. In the current study, phylogenetically conserved amino acids of HBx with a high potential of phosphorylation were targeted for site-directed mutagenesis. Two conserved serine (Ser25 and Ser41), and one conserved threonine (Thr81) amino acids were replaced by either alanine or aspartic acid residues to simulate an unphosphorylated or phosphorylated state, respectively. Human hepatoma cells were transfected with increasing amounts of the HBx DNA constructs, and the cells were analyzed by fluorescence microscopy. Together, our results show that the nucleocytoplasmic distribution of the HBx protein could be regulated by phosphorylation since some of the modified proteins were mainly confined to distinct subcellular compartments. Remarkably, both HBx Ser41A, and HBx Thr81D proteins were predominantly localized within the nuclear compartment throughout the different expression levels of HBx mutants.


Asunto(s)
Carcinoma Hepatocelular/genética , Hepatitis B/genética , Neoplasias Hepáticas/genética , Transactivadores/genética , Proteínas Reguladoras y Accesorias Virales/genética , Secuencia de Aminoácidos/genética , Carcinoma Hepatocelular/patología , Carcinoma Hepatocelular/virología , Secuencia Conservada/genética , Regulación Viral de la Expresión Génica/genética , Genoma Viral/genética , Células Hep G2 , Hepatitis B/patología , Hepatitis B/virología , Virus de la Hepatitis B/genética , Virus de la Hepatitis B/patogenicidad , Humanos , Neoplasias Hepáticas/patología , Neoplasias Hepáticas/virología , Fosforilación/genética , Filogenia
19.
Nat Commun ; 12(1): 1379, 2021 03 02.
Artículo en Inglés | MEDLINE | ID: mdl-33654074

RESUMEN

Many immune responses depend upon activation of NF-κB, an important transcription factor in the elicitation of a cytokine response. Here we show that N4BP1 inhibits TLR-dependent activation of NF-κB by interacting with the NF-κB signaling essential modulator (NEMO, also known as IκB kinase γ) to attenuate NEMO-NEMO dimerization or oligomerization. The UBA-like (ubiquitin associated-like) and CUE-like (ubiquitin conjugation to ER degradation-like) domains in N4BP1 mediate interaction with the NEMO COZI domain. Both in vitro and in mice, N4bp1 deficiency specifically enhances TRIF-independent (TLR2, TLR7, or TLR9-mediated) but not TRIF-dependent (TLR3 or TLR4-mediated) NF-κB activation, leading to increased production of proinflammatory cytokines. In response to TLR4 or TLR3 activation, TRIF causes activation of caspase-8, which cleaves N4BP1 distal to residues D424 and D490 and abolishes its inhibitory effect. N4bp1-/- mice also have diminished numbers of T cells in the peripheral blood. Our work identifies N4BP1 as an inhibitory checkpoint protein that must be overcome to activate NF-κB, and a TRIF-initiated caspase-8-dependent mechanism by which this is accomplished.


Asunto(s)
Proteínas Portadoras/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , FN-kappa B/metabolismo , Multimerización de Proteína , Proteínas Adaptadoras del Transporte Vesicular/metabolismo , Animales , Proteínas Portadoras/química , Proteínas Portadoras/genética , Caspasa 8/metabolismo , Femenino , Regulación de la Expresión Génica/efectos de los fármacos , Células HEK293 , Herpesvirus Humano 1/fisiología , Humanos , Interleucina-6/sangre , Macrófagos Peritoneales/efectos de los fármacos , Macrófagos Peritoneales/metabolismo , Ratones Endogámicos C57BL , Mutación/genética , Inhibidor NF-kappaB alfa/metabolismo , Oligodesoxirribonucleótidos/farmacología , Fosforilación/efectos de los fármacos , Unión Proteica/efectos de los fármacos , Dominios Proteicos , Multimerización de Proteína/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Ubiquitina/metabolismo
20.
Nat Commun ; 12(1): 1382, 2021 03 02.
Artículo en Inglés | MEDLINE | ID: mdl-33654076

RESUMEN

Mechanistic understanding of oncogenic variants facilitates the development and optimization of treatment strategies. We recently identified in-frame, tandem duplication of EGFR exons 18 - 25, which causes EGFR Kinase Domain Duplication (EGFR-KDD). Here, we characterize the prevalence of ERBB family KDDs across multiple human cancers and evaluate the functional biochemistry of EGFR-KDD as it relates to pathogenesis and potential therapeutic intervention. We provide computational and experimental evidence that EGFR-KDD functions by forming asymmetric EGF-independent intra-molecular and EGF-dependent inter-molecular dimers. Time-resolved fluorescence microscopy and co-immunoprecipitation reveals EGFR-KDD can form ligand-dependent inter-molecular homo- and hetero-dimers/multimers. Furthermore, we show that inhibition of EGFR-KDD activity is maximally achieved by blocking both intra- and inter-molecular dimerization. Collectively, our findings define a previously unrecognized model of EGFR dimerization, providing important insights for the understanding of EGFR activation mechanisms and informing personalized treatment of patients with tumors harboring EGFR-KDD. Finally, we establish ERBB KDDs as recurrent oncogenic events in multiple cancers.


Asunto(s)
Receptores ErbB/química , Receptores ErbB/metabolismo , Duplicación de Gen , Terapia Molecular Dirigida , Oncogenes , Secuencia de Aminoácidos , Animales , Línea Celular , Proliferación Celular , Epítopos/metabolismo , Receptores ErbB/genética , Ligandos , Ratones , Neoplasias/metabolismo , Fosforilación , Unión Proteica , Dominios Proteicos , Multimerización de Proteína , Relación Estructura-Actividad
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