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1.
Zhong Nan Da Xue Xue Bao Yi Xue Ban ; 45(1): 61-67, 2020 Jan 28.
Artículo en Inglés, Chino | MEDLINE | ID: mdl-32132299

RESUMEN

Protein tyrosine phosphatase H-type receptor (PTPRH) gene encodes a gastric cancer associated protein, which exerts its biological function through tyrosine phosphorylation in the post-translational COOH- terminal region. PTPRH is abnormally expressed in a variety of tumors, and its biological function is closely related to the occurrence, development and prognosis of tumors.


Asunto(s)
Neoplasias Gástricas , Humanos , Fosforilación , Proteínas Tirosina Fosfatasas , Proteínas , Tirosina
2.
Adv Exp Med Biol ; 1202: 203-222, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32034715

RESUMEN

STAT (signal transducers and activators of transcription) are latent cytoplasmic transcription factors that function as downstream effectors of cytokine and growth factor receptor signaling. The canonical JAK/STAT signaling pathway involves the activation of Janus kinases (JAK) or growth factors receptor kinases, phosphorylation of STAT proteins, their dimerization and translocation into the nucleus where STATs act as transcription factors with pleiotropic downstream effects. STAT signaling is tightly controlled with restricted kinetics due to action of its negative regulators. While STAT1 is believed to play an important role in growth arrest and apoptosis, and to act as a tumor suppressor, STAT3 and 5 are involved in promoting cell cycle progression, cellular transformation, and preventing apoptosis. Aberrant activation of STATs, in particular STAT3 and STAT5, have been found in a large number of human tumors, including gliomas and may contribute to oncogenesis. In this chapter, we have (1) summarized the mechanisms of STAT activation in normal and malignant signaling; (2) discussed evidence for the critical role of constitutively activated STAT3 and STAT5 in glioma pathobiology; (3) disclosed molecular and pharmacological strategies to interfere with STAT signaling for potential therapeutic intervention in gliomas.


Asunto(s)
Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patología , Glioma/metabolismo , Glioma/patología , Factores de Transcripción STAT/metabolismo , Transducción de Señal , Transformación Celular Neoplásica , Humanos , Quinasas Janus/metabolismo , Fosforilación
3.
Cell Physiol Biochem ; 54(2): 195-210, 2020 Feb 22.
Artículo en Inglés | MEDLINE | ID: mdl-32083406

RESUMEN

BACKGROUND/AIMS: Idiopathic pulmonary fibrosis (IPF) is a specific form of progressive and chronic interstitial lung disease of unknown cause. IPF is characterized by excessive deposition of extracellular matrix (ECM) and destructive pathological remodeling due to epithelial-to-mesenchymal transition (EMT). Eventually, lung interstitium thickens and stiffens and breathing becomes difficult. It has been well established that the transforming growth factor-ß1 (TGF-ß1)/Smad signaling pathway plays a critical role in the pathogenesis of pulmonary fibrosis. TGF-ß1-mediated activation of mitogen activated protein kinase (MAPK) family affects Smad signaling. p90RSK is a serine/threonine kinase and is activated by the extracellular signal-regulated kinase (ERK) signaling pathway. However, the roles played by p90RSK in TGF-ß1 signaling and the pathogenesis of pulmonary fibrosis remain unknown. METHODS: We investigated whether p90RSK regulates the pathogenesis of pulmonary fibrosis using in vitro and in vivo systems and Western blotting, real-time quantitative PCR, transcriptional activity assays and immunofluorescence studies. RESULTS: Pharmacological inhibition of p90RSK by FMK or inhibition of p90RSK with adenoviral vector encoding a dominant negative form of p90RSK suppressed TGF-ß1-induced ECM accumulation and EMT in lung epithelial cells and fibroblasts. Interestingly, FMK significantly inhibited TGF-ß1-induced Smad3 nuclear translocation and smad binding element-dependent transcriptional activity, but not Smad3 phosphorylation. Furthermore, in a mouse model of bleomycin-induced lung fibrosis, FMK ameliorated pulmonary fibrosis. CONCLUSION: These findings indicate that p90RSK plays critical roles in pulmonary fibrosis, which suggests it be viewed as a novel therapeutic target for the treatment of lung fibrosis.


Asunto(s)
Proteínas Quinasas S6 Ribosómicas 90-kDa/metabolismo , Proteína smad3/metabolismo , Animales , Cadherinas/genética , Cadherinas/metabolismo , Línea Celular , Transición Epitelial-Mesenquimal/efectos de los fármacos , Matriz Extracelular/metabolismo , Humanos , Isoquinolinas/farmacología , Cetonas/farmacología , Ratones , Ratones Endogámicos C57BL , Fosforilación/efectos de los fármacos , Inhibidor 1 de Activador Plasminogénico/genética , Inhibidor 1 de Activador Plasminogénico/metabolismo , Regiones Promotoras Genéticas/efectos de los fármacos , Fibrosis Pulmonar/inducido químicamente , Fibrosis Pulmonar/metabolismo , Fibrosis Pulmonar/patología , Piridinas/farmacología , Pirroles/farmacología , Proteínas Quinasas S6 Ribosómicas 90-kDa/antagonistas & inhibidores , Transducción de Señal/efectos de los fármacos , Proteína smad3/antagonistas & inhibidores , Proteína smad3/genética , Activación Transcripcional/efectos de los fármacos , Factor de Crecimiento Transformador beta1/metabolismo , Factor de Crecimiento Transformador beta1/farmacología , Tubulina (Proteína)/genética , Tubulina (Proteína)/metabolismo
4.
Nature ; 579(7798): 291-296, 2020 03.
Artículo en Inglés | MEDLINE | ID: mdl-32103174

RESUMEN

The DNA-dependent protein kinase (DNA-PK), which comprises the KU heterodimer and a catalytic subunit (DNA-PKcs), is a classical non-homologous end-joining (cNHEJ) factor1. KU binds to DNA ends, initiates cNHEJ, and recruits and activates DNA-PKcs. KU also binds to RNA, but the relevance of this interaction in mammals is unclear. Here we use mouse models to show that DNA-PK has an unexpected role in the biogenesis of ribosomal RNA (rRNA) and in haematopoiesis. The expression of kinase-dead DNA-PKcs abrogates cNHEJ2. However, most mice that both expressed kinase-dead DNA-PKcs and lacked the tumour suppressor TP53 developed myeloid disease, whereas all other previously characterized mice deficient in both cNHEJ and TP53 expression succumbed to pro-B cell lymphoma3. DNA-PK autophosphorylates DNA-PKcs, which is its best characterized substrate. Blocking the phosphorylation of DNA-PKcs at the T2609 cluster, but not the S2056 cluster, led to KU-dependent defects in 18S rRNA processing, compromised global protein synthesis in haematopoietic cells and caused bone marrow failure in mice. KU drives the assembly of DNA-PKcs on a wide range of cellular RNAs, including the U3 small nucleolar RNA, which is essential for processing of 18S rRNA4. U3 activates purified DNA-PK and triggers phosphorylation of DNA-PKcs at T2609. DNA-PK, but not other cNHEJ factors, resides in nucleoli in an rRNA-dependent manner and is co-purified with the small subunit processome. Together our data show that DNA-PK has RNA-dependent, cNHEJ-independent functions during ribosome biogenesis that require the kinase activity of DNA-PKcs and its phosphorylation at the T2609 cluster.


Asunto(s)
Proteínas de Unión al Calcio/metabolismo , Hematopoyesis/genética , Autoantígeno Ku/metabolismo , Linfoma/enzimología , Linfoma/fisiopatología , ARN Ribosómico 18S/metabolismo , Proteínas de Unión al Calcio/genética , Dominio Catalítico/fisiología , Reparación del ADN/genética , Activación Enzimática/genética , Células HeLa , Humanos , Linfoma/genética , Modelos Animales , Mutación , Fosforilación , Unión Proteica , Biosíntesis de Proteínas/genética , ARN Ribosómico 18S/genética , ARN Nucleolar Pequeño/metabolismo
5.
Anticancer Res ; 40(2): 767-777, 2020 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-32014919

RESUMEN

BACKGROUND/AIM: Chicken ovalbumin upstream promoter-transcription factor II (COUP-TFII) plays an important role in cancer. We examined the effect of COUP-TFII overexpression on the proliferation and invasion of the human colorectal cancer SNU-C4 cells. MATERIALS AND METHODS: SNU-C4 cells were stably transfected with COUP-TFII expression plasmid to overexpress COUP-TFII (COUP-TFII-SNU-C4 cells). Cell proliferation, colony-forming ability and transwell invasion assays were performed. To elucidate the underlying molecular mechanism of COUP-TFII action, western blot analysis, p53 shRNA transfection, and Myr-Akt transfection were performed. RESULTS: Cell proliferation and colony-forming ability were significantly inhibited in COUP-TFII-SNU-C4 cells. Western blot analyses demonstrated that while the expression of p53 and PTEN was increased, the p-Akt levels were decreased in COUP-TFII-SNU-C4 cells. Knockdown of p53 partially restored the cell proliferation, but did not reverse the inhibition of invasion. Constitutive activation of Akt via Myr-Akt transfection reversed the inhibited cell proliferation and invasion by COUP-TFII. CONCLUSION: p53 is required for the inhibition of cell proliferation, and decreased phosphorylation of Akt may mediate the inhibition of cell proliferation and invasion by COUP-TFII.


Asunto(s)
Factor de Transcripción COUP II/biosíntesis , Neoplasias Colorrectales/metabolismo , Fosfohidrolasa PTEN/biosíntesis , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteína p53 Supresora de Tumor/biosíntesis , Factor de Transcripción COUP II/metabolismo , Proliferación Celular/fisiología , Neoplasias Colorrectales/patología , Humanos , Fosfohidrolasa PTEN/metabolismo , Fosforilación , Transfección , Proteína p53 Supresora de Tumor/metabolismo
6.
Braz Oral Res ; 34: e006, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32022225

RESUMEN

Induced pluripotent stem (iPS) cells could be induced into ameloblast-like cells by ameloblasts serum-free conditioned medium (ASF-CM), and bone morphogenetic proteins (BMPs) might be essential during the regulation of this process. The present study investigates the signal transduction that regulates the ameloblastic differentiation of iPS cells induced by ASF-CM. Mouse iPS cells were characterized and then cultured for 14 days in epithelial cell medium (control) or ASF-CM. Bone morphogenetic protein receptor II (BMPR-II) siRNA, inhibitor of Smad1/5 phosphorylation activated by activin receptor-like kinase (ALK) receptors, and inhibitors of mitogen-activated protein kinases (MAPKs) phosphorylation were used to treat the iPS cells in combination with ASF-CM. Real-time PCR, western blotting, and immunofluorescent staining were used to evaluate the expressions of ameloblast markers ameloblastin, enamelin, and cytokeratin-14. BMPR-II gene and protein levels increased markedly in ASF-CM-treated iPS cells compared with the controls, while the mRNA levels of Bmpr-Ia and Bmpr-Ib were similar between the ASF-CM and control groups. ASF-CM stimulation significantly increased the gene and protein expression of ameloblastin, enamelin and cytokeratin-14, and phosphorylated SMAD1/5, p38 MAPK, and ERK1/2 MAPK compared with the controls. Knockdown of BMPR-II and inhibition of Smad1/5 phosphorylation both could significantly reverse the increased expression of ameloblastin, enamelin, and cytokeratin-14 induced by ASF-CM, while neither inhibition of p38 nor ERK1/2 phosphorylation had significant reversing effects. We conclude that smad1/5 signaling transduction, activated by ALK receptors, regulates the ameloblastic differentiation of iPS cells induced by ameloblast-conditioned medium.


Asunto(s)
Ameloblastos/citología , Células Madre Pluripotentes Inducidas/citología , Transducción de Señal/fisiología , Proteína Smad1/fisiología , Receptores de Activinas/análisis , Receptores de Activinas/fisiología , Western Blotting , Receptores de Proteínas Morfogenéticas Óseas de Tipo II/análisis , Receptores de Proteínas Morfogenéticas Óseas de Tipo II/fisiología , Diferenciación Celular/genética , Diferenciación Celular/fisiología , Células Cultivadas , Medio de Cultivo Libre de Suero , Técnica del Anticuerpo Fluorescente , Expresión Génica , Sistema de Señalización de MAP Quinasas/fisiología , Fosforilación , Interferencia de ARN , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Proteína Smad1/análisis , Factores de Tiempo , Proteínas Quinasas p38 Activadas por Mitógenos/análisis , Proteínas Quinasas p38 Activadas por Mitógenos/fisiología
7.
Adv Exp Med Biol ; 1202: 109-128, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32034711

RESUMEN

This chapter describes signaling pathways, stimulated by the P2Y2 nucleotide receptor (P2Y2R), that regulate cellular processes dependent on actin cytoskeleton dynamics in glioma C6 cells. P2Y2R coupled with G-proteins, in response to ATP or UTP, regulates the level of iphosphatidylinositol-4,5-bisphosphate (PIP2) which modulates a variety of actin binding proteins and is involved in calcium response and activates Rac1 and RhoA proteins. The RhoA/ROCK signaling pathway plays an important role in contractile force generation needed for the assembly of stress fibers, focal adhesions and for tail retraction during cell migration. Blocking of this pathway by a specific Rho-kinase inhibitor induces changes in F-actin organization and cell shape and decreases the level of phosphorylated myosin II and cofilin. In glioma C6 cells these changes are reversed after UTP stimulation of P2Y2R. Signaling pathways responsible for this compensation are calcium signaling which regulates MLC kinase activation via calmodulin, and the Rac1/PAK/LIMK cascade. Stimulation of the Rac1 mediated pathway via Go proteins needs additional interaction between αvß5 integrins and P2Y2Rs. Calcium free medium, or growing of the cells in suspension, prevents Gαo activation by P2Y2 receptors. Rac1 activation is necessary for cofilin phosphorylation as well as integrin activation needed for focal complexes formation and stabilization of lamellipodium. Inhibition of positive Rac1 regulation prevents glioma C6 cells from recovery of control cell like morphology.


Asunto(s)
Citoesqueleto/metabolismo , Glioma/metabolismo , Receptores Purinérgicos P2Y2/metabolismo , Transducción de Señal , Actinas/metabolismo , Animales , Línea Celular Tumoral , Glioma/patología , Humanos , Nucleótidos/metabolismo , Fosforilación
8.
Adv Exp Med Biol ; 1202: 151-178, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32034713

RESUMEN

Protein tyrosine kinases are enzymes that are capable of adding a phosphate group to specific tyrosines on target proteins. A receptor tyrosine kinase (RTK) is a tyrosine kinase located at the cellular membrane and is activated by binding of a ligand via its extracellular domain. Protein phosphorylation by kinases is an important mechanism for communicating signals within a cell and regulating cellular activity; furthermore, this mechanism functions as an "on" or "off" switch in many cellular functions. Ninety unique tyrosine kinase genes, including 58 RTKs, were identified in the human genome; the products of these genes regulate cellular proliferation, survival, differentiation, function, and motility. Tyrosine kinases play a critical role in the development and progression of many types of cancer, in addition to their roles as key regulators of normal cellular processes. Recent studies have revealed that RTKs such as epidermal growth factor receptor (EGFR), platelet-derived growth factor receptor (PDGFR), c-Met, Tie, Axl, discoidin domain receptor 1 (DDR1), and erythropoietin-producing human hepatocellular carcinoma (Eph) play a major role in glioma invasion. Herein, we summarize recent advances in understanding the role of RTKs in glioma pathobiology, especially the invasive phenotype, and present the perspective that RTKs are a potential target of glioma therapy.


Asunto(s)
Neoplasias Encefálicas/enzimología , Neoplasias Encefálicas/patología , Glioma/enzimología , Glioma/patología , Proteínas Tirosina Quinasas Receptoras/metabolismo , Animales , Neoplasias Encefálicas/tratamiento farmacológico , Movimiento Celular , Proliferación Celular , Glioma/tratamiento farmacológico , Humanos , Fosforilación , Fosfotirosina/metabolismo , Proteínas Tirosina Quinasas Receptoras/antagonistas & inhibidores
9.
Adv Exp Med Biol ; 1202: 179-201, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32034714

RESUMEN

Transforming growth factor beta (TGF-ß) signaling is involved in the regulation of proliferation, differentiation and survival/or apoptosis of many cells, including glioma cells. TGF-ß acts via specific receptors activating multiple intracellular pathways resulting in phosphorylation of receptor-regulated Smad2/3 proteins that associate with the common mediator, Smad4. Such complex translocates to the nucleus, binds to DNA and regulates transcription of many genes. Furthermore, TGF-ß-activated kinase-1 (TAK1) is a component of TGF-ß signaling and activates mitogen-activated protein kinase (MAPK) cascades. Negative regulation of TGF-ß/Smad signaling may occur through the inhibitory Smad6/7. While genetic alterations in genes related to TGF-ß signaling are relatively rare in gliomas, the altered expression of those genes is a frequent event. The increased expression of TGF-ß1-3 correlates with a degree of malignancy of human gliomas. TGF-ß may contribute to tumor pathogenesis in many ways: by direct support of tumor growth, by maintaining self-renewal of glioma initiating stem cells and inhibiting anti-tumor immunity. Glioma initiating cells are dedifferentiated cells that retain many stem cell-like properties, play a role in tumor initiation and contribute to its recurrence. TGF-ß1,2 stimulate expression of the vascular endothelial growth factor as well as the plasminogen activator inhibitor and some metalloproteinases that are involved in vascular remodeling, angiogenesis and degradation of the extracellular matrix. Inhibitors of TGF-ß signaling reduce viability and invasion of gliomas in animal models and show a great promise as novel, potential anti-tumor therapeutics.


Asunto(s)
Glioma/metabolismo , Glioma/patología , Transducción de Señal , Factor de Crecimiento Transformador beta/metabolismo , Animales , Carcinogénesis , Glioma/tratamiento farmacológico , Humanos , Fosforilación , Receptores de Factores de Crecimiento Transformadores beta/metabolismo
10.
Cell Host Microbe ; 27(3): 476-485.e7, 2020 03 11.
Artículo en Inglés | MEDLINE | ID: mdl-32101706

RESUMEN

Chronically infecting pathogens avoid clearance by the innate immune system by promoting premature transition from an initial pro-inflammatory response toward an anti-inflammatory tissue-repair response. STAT3, a central regulator of inflammation, controls this transition and thus is targeted by numerous chronic pathogens. Here, we show that BepD, an effector of the chronic bacterial pathogen Bartonella henselae targeted to infected host cells, establishes an exceptional pathway for canonical STAT3 activation, thereby impairing secretion of pro-inflammatory TNF-α and stimulating secretion of anti-inflammatory IL-10. Tyrosine phosphorylation of EPIYA-related motifs in BepD facilitates STAT3 binding and activation via c-Abl-dependent phosphorylation of Y705. The tyrosine-phosphorylated scaffold of BepD thus represents a signaling hub for intrinsic STAT3 activation that is independent from canonical STAT3 activation via transmembrane receptor-associated Janus kinases. We anticipate that our findings on a molecular shortcut to STAT3 activation will inspire new treatment options for chronic infections and inflammatory diseases.


Asunto(s)
Proteínas Bacterianas/metabolismo , Bartonella henselae/inmunología , Interleucina-10/metabolismo , Factor de Transcripción STAT3/metabolismo , Transducción de Señal , Secuencia de Aminoácidos , Animales , Línea Celular , Citocinas/inmunología , Femenino , Quinasas Janus/metabolismo , Ratones , Ratones Endogámicos C57BL , Fosforilación , Células RAW 264.7 , Factor de Necrosis Tumoral alfa/metabolismo
12.
Adv Exp Med Biol ; 1227: 51-68, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32072498

RESUMEN

The Notch signaling pathway seems deceptively simple, with its key feature being a direct connection between extracellular signal and transcriptional output without the need for an extended chain of protein intermediaries as required by so many other signaling paradigms. However, this apparent simplicity hides considerable complexity. Consistent with its central role in many aspects of development, Notch signaling has an extensive collection of mechanisms that it employs alongside of its core transcriptional machinery. These so-called noncanonical Notch pathways diversify the potential outputs of Notch, and allow it to coordinate regulation of many aspects of the biology of cells. Here we will review noncanonical Notch signaling with special attention to the role of posttranslational modifications of Notch. We will also consider the importance of coordinating the activity of gene expression with regulation of cell morphology in biological processes, including axon guidance and other morphological events during embryogenesis.


Asunto(s)
Proteolisis , Receptores Notch/metabolismo , Transducción de Señal , Animales , Desarrollo Embrionario , Humanos , Fosforilación
13.
Nat Commun ; 11(1): 539, 2020 Jan 27.
Artículo en Inglés | MEDLINE | ID: mdl-31988277

RESUMEN

In the Caenorhabditis elegans zygote, PAR protein patterns, driven by mutual anatagonism, determine the anterior-posterior axis and facilitate the redistribution of proteins for the first cell division. Yet, the factors that determine the selection of the polarity axis remain unclear. We present a reaction-diffusion model in realistic cell geometry, based on biomolecular reactions and accounting for the coupling between membrane and cytosolic dynamics. We find that the kinetics of the phosphorylation-dephosphorylation cycle of PARs and the diffusive protein fluxes from the cytosol towards the membrane are crucial for the robust selection of the anterior-posterior axis for polarisation. The local ratio of membrane surface to cytosolic volume is the main geometric cue that initiates pattern formation, while the choice of the long-axis for polarisation is largely determined by the length of the aPAR-pPAR interface, and mediated by processes that minimise the diffusive fluxes of PAR proteins between cytosol and membrane.


Asunto(s)
Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/metabolismo , Polaridad Celular , Animales , División Celular Asimétrica , Caenorhabditis elegans/citología , Caenorhabditis elegans/embriología , Biología Computacional , Citosol/metabolismo , Embrión no Mamífero/citología , Embrión no Mamífero/metabolismo , Desarrollo Embrionario , Cinética , Modelos Biológicos , Fosforilación , Transducción de Señal , Termodinámica
15.
Invest Ophthalmol Vis Sci ; 61(1): 1, 2020 Jan 23.
Artículo en Inglés | MEDLINE | ID: mdl-31995153

RESUMEN

Purpose: Vacuolar protein sorting 35 (Vps35) mutations and protein dysfunction have been linked to the hyperphosphorylation and accumulation of tau protein in a number of central neurodegenerative disorders. The aims of the present study were to investigate the mechanism underlying the tau hyperphosphorylation caused by Vps35 deficiency. Methods: The cells used in this study were primary retinal ganglion cells (RGCs). The rat retinal glutamate excitotoxicity model was used in vivo. Fresh retinal tissues or eyeballs were collected at different time points. The expression and interactions of Vps35, Cdk5/p35, tau hyperphosphorylation, LAMP1, EEA1 and UBE1 in RGCs were studied by immunofluorescence staining, Western blotting, and immunoprecipitation. Results: The downregulation and overexpression of Vps35 increased and decreased the expression of p35 and tau hyperphosphorylation, respectively. More important, roscovitine, a Cdk5 inhibitor, could effectively decrease the hyperphosphorylated tau level induced by Vps35 deficiency. Furthermore, this study confirmed that the inhibition of Vps35 could increase the activity of Cdk5/p35 by affecting the lysosomal degradation of p35 and lead to the degeneration of RGCs. Conclusions: These findings demonstrate the possibility that Cdk5/p35 acts as a "cargo" of Vps35 and provide new insights into the pathogenesis of RGC degeneration caused by hyperphosphorylated tau protein. Vps35 is a potential target for basic research and clinical treatment of RGC degeneration in many ocular diseases such as glaucoma.


Asunto(s)
Quinasa 5 Dependiente de la Ciclina/metabolismo , Fosfotransferasas/metabolismo , Células Ganglionares de la Retina/metabolismo , Proteínas de Transporte Vesicular/deficiencia , Proteínas tau/metabolismo , Animales , Western Blotting , Células Cultivadas , Quinasa 5 Dependiente de la Ciclina/antagonistas & inhibidores , Regulación hacia Abajo , Técnica del Anticuerpo Fluorescente Indirecta , Ácido Glutámico/toxicidad , Glicoproteínas de la Membrana Asociadas a los Lisosomas/metabolismo , Masculino , Ratones , Fosforilación , Inhibidores de Proteínas Quinasas/farmacología , Ratas , Ratas Sprague-Dawley , Reacción en Cadena en Tiempo Real de la Polimerasa , Degeneración Retiniana/inducido químicamente , Degeneración Retiniana/metabolismo , Roscovitina/farmacología , Transfección , Enzimas Activadoras de Ubiquitina/metabolismo , Proteínas de Transporte Vesicular/metabolismo
16.
Chem Biol Interact ; 317: 108944, 2020 Feb 01.
Artículo en Inglés | MEDLINE | ID: mdl-31935364

RESUMEN

Bone mesenchymal stem cells (BMSCs) are a well-known donor graft source due to their potential for self-renewal and differentiation into multi-lineage cell types, including osteoblasts that are critical for fracture healing. Fasudil (FAS), a Rho kinase inhibitor, has been proven to induce the differentiation of bone marrow stem cells (BMSCs) into neuron-like cells. However, its role in the osteogenesis of BMSCs remain uncertain. Herein, we for the first time studied the effects of FAS on osteogenic differentiation in a mouse fracture model and further explored the involved mechanisms in mouse BMSCs. The results showed that FAS stimulated bone formation in the fracture mouse model. Additionally, at 30 µM, FAS significantly promotes alkaline phosphatase activity, mineralization, and the expression of osteogenic markers COL-1, RUNX2 and OCN in murine BMSCs. Blocking of P38 by SB202190 significantly reversed the effects of FAS, in vitro, suggesting that P38, but not ERK or JNK activation is required for FAS-induced osteogenesis. Collectively, our results indicate that FAS may be a promising agent for promoting fracture healing.


Asunto(s)
1-(5-Isoquinolinesulfonil)-2-Metilpiperazina/análogos & derivados , Células de la Médula Ósea/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , 1-(5-Isoquinolinesulfonil)-2-Metilpiperazina/farmacología , Animales , Biomarcadores , Proliferación Celular/efectos de los fármacos , Femenino , Fracturas Óseas , Regulación de la Expresión Génica/efectos de los fármacos , Imidazoles/farmacología , Células Madre Mesenquimatosas , Ratones , Osteogénesis/efectos de los fármacos , Fosforilación , Piridinas/farmacología
17.
Nat Commun ; 11(1): 575, 2020 Jan 29.
Artículo en Inglés | MEDLINE | ID: mdl-31996678

RESUMEN

mTORC2 phosphorylates AKT in a hydrophobic motif site that is a biomarker of insulin sensitivity. In brown adipocytes, mTORC2 regulates glucose and lipid metabolism, however the mechanism has been unclear because downstream AKT signaling appears unaffected by mTORC2 loss. Here, by applying immunoblotting, targeted phosphoproteomics and metabolite profiling, we identify ATP-citrate lyase (ACLY) as a distinctly mTORC2-sensitive AKT substrate in brown preadipocytes. mTORC2 appears dispensable for most other AKT actions examined, indicating a previously unappreciated selectivity in mTORC2-AKT signaling. Rescue experiments suggest brown preadipocytes require the mTORC2/AKT/ACLY pathway to induce PPAR-gamma and establish the epigenetic landscape during differentiation. Evidence in mature brown adipocytes also suggests mTORC2 acts through ACLY to increase carbohydrate response element binding protein (ChREBP) activity, histone acetylation, and gluco-lipogenic gene expression. Substrate utilization studies additionally implicate mTORC2 in promoting acetyl-CoA synthesis from acetate through acetyl-CoA synthetase 2 (ACSS2). These data suggest that a principal mTORC2 action is controlling nuclear-cytoplasmic acetyl-CoA synthesis.


Asunto(s)
ATP Citrato (pro-S)-Liasa/metabolismo , Adipocitos Marrones/metabolismo , Lipogénesis/fisiología , Diana Mecanicista del Complejo 2 de la Rapamicina/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Acetato CoA Ligasa/metabolismo , Animales , Proteínas Portadoras , Epigénesis Genética , Ácido Graso Sintasas , Edición Génica , Expresión Génica , Transportador de Glucosa de Tipo 4/genética , Transportador de Glucosa de Tipo 4/metabolismo , Células HEK293 , Histonas/metabolismo , Humanos , Lipogénesis/genética , Ratones , Ratones Endogámicos C57BL , PPAR gamma/metabolismo , Fosforilación , Proteómica , Elementos de Respuesta
18.
Cell Prolif ; 53(2): e12743, 2020 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-31943455

RESUMEN

OBJECTIVES: Alveolar bone osteoporosis has attracted more and more attention because of its profound impact on stomatognathic function and treatment, but current treatments have not been targeted to alveolar bone and might even cause severe side effects. Thus, identifying the effects of anti-osteoporosis agents on alveolar bone is essential. Icariin ameliorates metabolic dysfunction of long bones, but its effects on alveolar bone remain unclarified. MATERIALS AND METHODS: BMSCs were isolated from rat mandibles (mBMSCs). The osteogenic potential of mBMSCs and the signalling pathway involved under icariin treatment were measured by ALP and alizarin red staining, reverse transcription-polymerase chain reaction (RT-PCR), Western blotting and immunofluorescence. Dual-luciferase assay, chromatin immunoprecipitation (ChIP) and co-immunoprecipitation were used to investigate the molecular mechanism. Ovariectomized and sham-operated rats treated with or without icariin were analysed by micro-CT, TRAP staining and calcein double labelling. RESULTS: We found that icariin promoted osteoblast differentiation of mBMSCs. Furthermore, STAT3 was critical for icariin-promoted osteoblast differentiation, as indicated by increased phosphorylation levels in icariin-treated mBMSCs, while preventing STAT3 activation blocked icariin-induced osteoblast differentiation. Mechanistically, icariin-promoted transcription of the downstream osteogenic gene osteocalcin (Ocn) through STAT3 and STAT3 bound to the promoter of Ocn. Notably, icariin prevented the alveolar bone osteoporosis induced by oestrogen deficiency through promoting bone formation. CONCLUSIONS: For the first time, our work provides evidence supporting the potential application of icariin in promoting osteogenesis and treating alveolar bone osteoporosis.


Asunto(s)
Pérdida de Hueso Alveolar/tratamiento farmacológico , Estrógenos/metabolismo , Flavonoides/farmacología , Osteogénesis/efectos de los fármacos , Factor de Transcripción STAT3/metabolismo , Pérdida de Hueso Alveolar/metabolismo , Animales , Diferenciación Celular/efectos de los fármacos , Células Cultivadas , Células Madre Mesenquimatosas/efectos de los fármacos , Células Madre Mesenquimatosas/metabolismo , Osteoblastos/efectos de los fármacos , Osteoblastos/metabolismo , Osteocalcina/efectos de los fármacos , Osteocalcina/metabolismo , Osteoporosis/tratamiento farmacológico , Osteoporosis/metabolismo , Fosforilación/efectos de los fármacos , Ratas , Transducción de Señal/efectos de los fármacos , Transcripción Genética/efectos de los fármacos
19.
Plant Mol Biol ; 102(4-5): 463-475, 2020 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-31916083

RESUMEN

The mechanism by which endogenous salicylic acid (SA) regulates leaf senescence remains elusive. Here we provide direct evidence that an enhancement of endogenous SA level, via chemical-induced upregulation of ISOCHORISMATE SYNTHASE 1 (ICS1), could significantly accelerate the senescence process of old leaves through mediation of the key SA signaling component NON EXPRESSOR OF PATHOGENESIS RELATED GENES 1 (NPR1) in Arabidopsis. Importantly, by taking advantage of this chemically induced leaf senescence system, we identified a mitogen-activated protein kinase (MAPK) cascade MKK4/5-MPK1/2 that is required for the SA/NPR1-mediated leaf senescence. Both MKK4/5 and MPK1/2 exhibited SA-induced kinase activities, with MPK1/2 being the immediate targets of MKK4/5. Double mutants of mkk4 mkk5 and mpk1 mpk2 displayed delayed leaf senescence, while constitutive overexpression of the kinase genes led to premature leaf senescence. Such premature leaf senescence was suppressed when they were overexpressed in an SA synthesis defective mutant (sid2) or signaling detective mutant (npr1). We further showed that MPK1, but not MPK2, could directly phosphorylate NPR1. Meanwhile, MPK1 also mediated NPR1 monomerization. Notably, induction of disease resistance was significantly compromised in the single and double mutants of the kinase genes. Taken together, our data demonstrate that the MKK4/5-MPK1/2 cascade plays a critical role in modulating SA signaling through a complex regulatory network in Arabidopsis.


Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/enzimología , Regulación de la Expresión Génica de las Plantas , Sistema de Señalización de MAP Quinasas , Hojas de la Planta/enzimología , Ácido Salicílico/farmacología , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Fosforilación , Hojas de la Planta/genética , Plantas Modificadas Genéticamente/genética , Transducción de Señal
20.
DNA Cell Biol ; 39(3): 343-348, 2020 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-31999481

RESUMEN

RNA editing is a process by which nascent RNA transcripts are covalently modified, thus enhancing the complexity of the transcriptome. The most common modifications are deaminations of adenosine to inosine at sites of complex RNA secondary structure, a process that is carried out by the adenosine deaminase acting on double-strand RNA (ADAR) family of RNA editases. Although much has been learned about the ADAR family members since their discovery, very little information on their post-transcriptional regulation has been reported. Similar to most proteins, the ADAR family members are post-translationally modified at multiple sites. We recently reported that members of the AKT kinase family directly phosphorylate ADAR1p110 and ADAR2 on a conserved threonine within the catalytic domain of the protein. Phosphorylation was observed to differentially inhibit the enzymatic activity of the ADAR proteins toward known RNA substrates. The direct downstream involvement of the AKT kinases in multiple major signaling pathways associated with cell survival, growth, glucose metabolism (insulin signaling), and differentiation is well established; thus, the AKT kinases represent a link between ADAR-dependent A-to-I editing and major signal transduction pathways that are necessary for cell maintenance and development.


Asunto(s)
Adenosina Desaminasa/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Edición de ARN , Proteínas de Unión al ARN/metabolismo , Transducción de Señal , Animales , Humanos , Fosforilación
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