RESUMEN
MicroRNA-mediated gene silencing is a fundamental mechanism in the regulation of gene expression. It remains unclear how the efficiency of RNA silencing could be influenced by RNA-binding proteins associated with the microRNA-induced silencing complex (miRISC). Here we report that fused in sarcoma (FUS), an RNA-binding protein linked to neurodegenerative diseases including amyotrophic lateral sclerosis (ALS), interacts with the core miRISC component AGO2 and is required for optimal microRNA-mediated gene silencing. FUS promotes gene silencing by binding to microRNA and mRNA targets, as illustrated by its action on miR-200c and its target ZEB1. A truncated mutant form of FUS that leads its carriers to an aggressive form of ALS, R495X, impairs microRNA-mediated gene silencing. The C. elegans homolog fust-1 also shares a conserved role in regulating the microRNA pathway. Collectively, our results suggest a role for FUS in regulating the activity of microRNA-mediated silencing.
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Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/metabolismo , Silenciador del Gen , MicroARNs/metabolismo , ARN de Helminto/metabolismo , Proteína FUS de Unión a ARN/metabolismo , Esclerosis Amiotrófica Lateral/genética , Esclerosis Amiotrófica Lateral/metabolismo , Esclerosis Amiotrófica Lateral/patología , Animales , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/genética , Células HEK293 , Humanos , Ratones , MicroARNs/genética , ARN de Helminto/genética , Proteína FUS de Unión a ARN/genéticaRESUMEN
Cells undergo metabolic adaptation during environmental changes by using evolutionarily conserved stress response programs. This metabolic homeostasis is exquisitely regulated, and its imbalance could underlie human pathological conditions. We report here that C9orf72, which is linked to the most common forms of the neurodegenerative diseases amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), is a key regulator of lipid metabolism under stress. Loss of C9orf72 leads to an overactivation of starvation-induced lipid metabolism that is mediated by dysregulated autophagic digestion of lipids and increased de novo fatty acid synthesis. C9orf72 acts by promoting the lysosomal degradation of coactivator-associated arginine methyltransferase 1 (CARM1), which in turn regulates autophagy-lysosomal functions and lipid metabolism. In ALS/FTD patient-derived neurons or tissues, a reduction in C9orf72 function is associated with dysregulation in the levels of CARM1, fatty acids, and NADPH oxidase NOX2. These results reveal a C9orf72-CARM1 axis in the control of stress-induced lipid metabolism and implicates epigenetic dysregulation in relevant human diseases.
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Proteína C9orf72/fisiología , Glucosa/fisiología , Metabolismo de los Lípidos , Proteína-Arginina N-Metiltransferasas/metabolismo , Estrés Fisiológico , Esclerosis Amiotrófica Lateral/genética , Esclerosis Amiotrófica Lateral/metabolismo , Animales , Proteína C9orf72/genética , Proteína C9orf72/metabolismo , Células Cultivadas , Ácidos Grasos/metabolismo , Demencia Frontotemporal/genética , Demencia Frontotemporal/metabolismo , Células HEK293 , Humanos , Lisosomas/metabolismo , Ratones , Proteína-Arginina N-Metiltransferasas/fisiologíaRESUMEN
Nutrient utilization and energy metabolism are critical for the maintenance of cellular homeostasis. A mutation in the C9orf72 gene has been linked to the most common forms of neurodegenerative diseases that include amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Here we have identified an evolutionarily conserved function of C9orf72 in the regulation of the transcription factor EB (TFEB), a master regulator of autophagic and lysosomal genes that is negatively modulated by mTORC1. Loss of the C. elegans orthologue of C9orf72, ALFA-1, causes the nuclear translocation of HLH-30/TFEB, leading to activation of lipolysis and premature lethality during starvation-induced developmental arrest in C. elegans. A similar conserved pathway exists in human cells, in which C9orf72 regulates mTOR and TFEB signaling. C9orf72 interacts with and dynamically regulates the level of Rag GTPases, which are responsible for the recruitment of mTOR and TFEB on the lysosome upon amino acid signals. These results have revealed previously unknown functions of C9orf72 in nutrient sensing and metabolic pathways and suggest that dysregulation of C9orf72 functions could compromise cellular fitness under conditions of nutrient stress.
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Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Proteína C9orf72/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Lipólisis , Transporte Activo de Núcleo Celular , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Proteína C9orf72/genética , Caenorhabditis elegans , Proteínas de Caenorhabditis elegans/genética , Núcleo Celular/metabolismo , Lisosomas/metabolismo , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Proteínas de Unión al GTP Monoméricas/genética , Proteínas de Unión al GTP Monoméricas/metabolismoRESUMEN
Human leukocyte antigen (HLA) class I has more than 18,000 alleles, each of which binds to a set of unique peptides from the cellular degradome. Deciphering the interaction between antigenic peptides and HLA proteins is crucial for understanding immune responses in autoimmune diseases and cancer. In this study, we aimed to characterize the peptidome that binds to HLA-A*33:03, which is one of the most prevalent HLA-A alleles in the Northeast Asian population, but poorly studied. For this purpose, we analyzed the HLA-A*33:03 monoallelic B cell line using immunoprecipitation of HLA-A and peptide complexes, followed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). In this study, we identified 5731 unique peptides that were associated with HLA A*33:03, and experimentally validated the affinity of 40 peptides for HLA-A*33:03 and their stability in HLA A*33:03-peptides complexes. To our knowledge, this study represents the largest dataset of peptides associated with HLA-A*33:03. Also, this is the first study in which HLA A*33:03-associated peptides were experimentally validated.
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Antígenos HLA-A , Espectrometría de Masas en Tándem , Cromatografía Liquida , Epítopos , Humanos , InmunoprecipitaciónRESUMEN
BACKGROUND: Hepatocytes usually express fibroblast growth factor receptor 4 (FGFR4), but not its ligand, fibroblast growth factor 19 (FGF19). A subtype of hepatocellular carcinoma (HCC) expresses FGF19, which activates the FGFR4 signaling pathway that induces cell proliferation. FGFR4 inhibitors that target this mechanism are under clinical development for the treatment of HCCs with FGF19 amplification or FGFR4 overexpression. Src plays an essential role in the FGFR1 and FGFR2 signaling pathways. However, it is yet to be understood whether Src has any role in the FGF19-FGFR4 pathway in HCCs. In this study, we aimed to elucidate the role of Src in the FGF19-FGFR4 axis in HCC. METHODS: 3 HCC cell lines expressing both FGF19 and FGFR4 were selected. The expression of each protein was suppressed by siRNA treatment, and the activity-regulating relationship between FGFR4 and Src was investigated by westernblot. Co-immunoprecipitation was performed using the FGFR4 antibody to identify the endosomal complex formation and receptor endocytosis. The intracellular migration pathways of the endosomal complex were observed by immuno-fluorescence and nuclear co-immunoprecipitation. Dasatinib and BLU9931 were used for cytotoxicity comparison. RESULTS: FGFR4 modulates the activity of Src and Src modulates the expression of FGFR4, showing a mutual regulatory relationship. FGFR4 activated by FGF19 formed an endosomal complex with Src and STAT3 and moved to the nucleus. However, when Src was suppressed, the formation of the endosomal complex was not observed. FGFR4 was released from the complex transferred into the nucleus and the binding of Src and STAT3 was maintained. Dasatinib showed cytotoxic results comparable to BLU9931. The results of our study demonstrated that Src is essential for the nuclear transport of STAT3, as it induces the endosomal delivery of FGFR4 in FGF19-expressing HCC cell lines. CONCLUSIONS: We found that Src is essential for the endosomal delivery of the FGFR4 signaling complex in HCC. Our findings provide a scientific rationale for repurposing Src inhibitors for the treatment of HCCs in which the FGFR4 pathway is activated.
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Carcinoma Hepatocelular , Neoplasias Hepáticas , Carcinoma Hepatocelular/tratamiento farmacológico , Proliferación Celular , Factores de Crecimiento de Fibroblastos , Humanos , Neoplasias Hepáticas/tratamiento farmacológico , Receptor Tipo 4 de Factor de Crecimiento de Fibroblastos/metabolismo , Transducción de SeñalRESUMEN
The formation of tissue boundaries is dependent on the cell-cell adhesion/repulsion system that is required for normal morphogenetic processes during development. The Smad ubiquitin regulatory factors (Smurfs) are E3 ubiquitin ligases with established roles in cell growth and differentiation, but whose roles in regulating cell adhesion and migration are just beginning to emerge. Here, we demonstrate that the Smurfs regulate tissue separation at mesoderm/ectoderm boundaries through antagonistic interactions with ephrinB1, an Eph receptor ligand that has a key role in regulating the separation of embryonic germ layers. EphrinB1 is targeted by Smurf2 for degradation; however, a Smurf1 interaction with ephrinB1 prevents the association with Smurf2 and precludes ephrinB1 from ubiquitination and degradation, since it is a substantially weaker substrate for Smurf1. Inhibition of Smurf1 expression in embryonic mesoderm results in loss of ephrinB1-mediated separation of this tissue from the ectoderm, which can be rescued by the coincident inhibition of Smurf2 expression. This system of differential interactions between Smurfs and ephrinB1 regulates the maintenance of tissue boundaries through the control of ephrinB protein levels.
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Efrina-B1/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Xenopus/genética , Xenopus/metabolismo , Animales , Embrión no Mamífero/enzimología , Regulación del Desarrollo de la Expresión Génica , Técnicas de Silenciamiento del Gen , Proteínas de Unión al GTP Monoméricas/metabolismo , Ubiquitina-Proteína Ligasas/genética , Proteínas de Pez Cebra/metabolismoRESUMEN
The most common cause of the neurodegenerative diseases amyotrophic lateral sclerosis and frontotemporal dementia is a hexanucleotide repeat expansion in C9orf72. Here we report a study of the C9orf72 protein by examining the consequences of loss of C9orf72 functions. Deletion of one or both alleles of the C9orf72 gene in mice causes age-dependent lethality phenotypes. We demonstrate that C9orf72 regulates nutrient sensing as the loss of C9orf72 decreases phosphorylation of the mTOR substrate S6K1. The transcription factor EB (TFEB), a master regulator of lysosomal and autophagy genes, which is negatively regulated by mTOR, is substantially up-regulated in C9orf72 loss-of-function animal and cellular models. Consistent with reduced mTOR activity and increased TFEB levels, loss of C9orf72 enhances autophagic flux, suggesting that C9orf72 is a negative regulator of autophagy. We identified a protein complex consisting of C9orf72 and SMCR8, both of which are homologous to DENN-like proteins. The depletion of C9orf72 or SMCR8 leads to significant down-regulation of each other's protein level. Loss of SMCR8 alters mTOR signaling and autophagy. These results demonstrate that the C9orf72-SMCR8 protein complex functions in the regulation of metabolism and provide evidence that loss of C9orf72 function may contribute to the pathogenesis of relevant diseases.
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Factores de Transcripción Básicos con Cremalleras de Leucinas y Motivos Hélice-Asa-Hélice/genética , Proteínas Portadoras/genética , Expansión de las Repeticiones de ADN/genética , Demencia Frontotemporal/genética , Factores de Intercambio de Guanina Nucleótido/genética , Serina-Treonina Quinasas TOR/genética , Alelos , Esclerosis Amiotrófica Lateral/genética , Esclerosis Amiotrófica Lateral/metabolismo , Esclerosis Amiotrófica Lateral/patología , Animales , Autofagia/genética , Factores de Transcripción Básicos con Cremalleras de Leucinas y Motivos Hélice-Asa-Hélice/biosíntesis , Proteína C9orf72 , Demencia Frontotemporal/metabolismo , Demencia Frontotemporal/patología , Regulación de la Expresión Génica , Humanos , Ratones , Fenotipo , Proteínas Quinasas S6 Ribosómicas 90-kDa/biosíntesis , Transducción de Señal/genética , Serina-Treonina Quinasas TOR/biosíntesisRESUMEN
BACKGROUND: A target-controlled infusion (TCI) of a propofol system uses a pharmacokinetic model to achieve and maintain a selected target blood propofol concentration. The aim of this study was to assess whether the propofol TCI system could be safely used by gastroenterologists in patients undergoing endoscopic resection including endoscopic submucosal dissection (ESD) and endoscopic mucosal resection (EMR) compared with a manually controlled infusion (MCI) system. METHODS: A total of 431 patients undergoing therapeutic endoscopy (178 ESD and 253 EMR) were consecutively included from November 2011 to August 2014. The patients were divided into the MCI (271) and TCI (160) propofol infusion groups. We compared adverse event rates in MCI and TCI groups and assessed independent risk factors for adverse events. RESULTS: The total sedation-related adverse event rate was 5.8 % (25/431). Most of the events were minor, and the rate of major events was 0.5 % (2/431). There was no significant difference in adverse event rate between the MCI and TCI groups [5.5 % (15/271) vs. 6.3 % (10/160); P = 0.759]. In univariate analysis, the propofol infusion time was significantly associated with adverse events (94.88 vs. 59.45 min, P = 0.017). In the multivariate analysis, there were no significant factors associated with adverse events. TCI was not an independent risk factor for adverse events despite the fact that the TCI had a longer duration of infusion and higher total infusion dose (95 % CI, 0.343-2.216; P = 0.773). CONCLUSIONS: TCI of propofol by gastroenterologists may provide safe sedation in patients undergoing ESD and EMR under careful respiratory monitoring.
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Algoritmos , Anestésicos Intravenosos/administración & dosificación , Sedación Profunda/métodos , Resección Endoscópica de la Mucosa/métodos , Endoscopía del Sistema Digestivo/métodos , Gastroenterólogos , Propofol/administración & dosificación , Adulto , Anciano , Anciano de 80 o más Años , Anestésicos Intravenosos/efectos adversos , Estudios de Casos y Controles , Femenino , Masaje Cardíaco/estadística & datos numéricos , Hipo/inducido químicamente , Humanos , Hipotensión/inducido químicamente , Hipotensión/tratamiento farmacológico , Hipoxia/inducido químicamente , Hipoxia/terapia , Infusiones Intravenosas , Intubación Intratraqueal/estadística & datos numéricos , Masculino , Persona de Mediana Edad , Propofol/efectos adversos , Aspiración Respiratoria/inducido químicamente , Estudios Retrospectivos , Ronquido/inducido químicamente , Vasoconstrictores/uso terapéutico , Adulto JovenRESUMEN
The Eph receptors and their membrane-bound ligands, ephrins, play important roles in various biological processes such as cell adhesion and movement. The transmembrane ephrinBs transduce reverse signaling in a tyrosine phosphorylation-dependent or -independent, as well as PDZ-dependent manner. Here, we show that ephrinB1 interacts with Connector Enhancer of KSR1 (CNK1) in an EphB receptor-independent manner. In cultured cells, cotransfection of ephrinB1 with CNK1 increases JNK phosphorylation. EphrinB1/CNK1-mediated JNK activation is reduced by overexpression of dominant-negative RhoA. Overexpression of CNK1 alone is sufficient for activation of RhoA; however, both ephrinB1 and CNK1 are required for JNK phosphorylation. Co-immunoprecipitation data showed that ephrinB1 and CNK1 act as scaffold proteins that connect RhoA and JNK signaling components, such as p115RhoGEF and MKK4. Furthermore, adhesion to fibronectin or active Src overexpression increases ephrinB1/CNK1 binding, whereas blocking Src activity by a pharmacological inhibitor decreases not only ephrinB1/CNK1 binding, but also JNK activation. EphrinB1 overexpression increases cell motility, however, CNK1 depletion by siRNA abrogates ephrinB1-mediated cell migration and JNK activation. Moreover, Rho kinase inhibitor or JNK inhibitor treatment suppresses ephrinB1-mediated cell migration. Taken together, our findings suggest that CNK1 is required for ephrinB1-induced JNK activation and cell migration.
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Movimiento Celular , Efrina-B1/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Línea Celular , Línea Celular Tumoral , Activación Enzimática , Efrina-B1/genética , Humanos , Péptidos y Proteínas de Señalización Intracelular/genética , Proteínas Quinasas JNK Activadas por Mitógenos/genética , Fosforilación , Unión Proteica , Proteína de Unión al GTP rhoA/genética , Proteína de Unión al GTP rhoA/metabolismoRESUMEN
BACKGROUND: Mutations in heat shock 27 kDa protein 1 (HSP27 or HSPB1) cause distal hereditary motor neuropathy (dHMN) or Charcot-Marie-Tooth disease type 2 F (CMT2F) according to unknown factors. Mutant HSP27 proteins affect axonal transport by reducing acetylated tubulin. RESULTS: We generated a transgenic mouse model overexpressing HSP27-S135F mutant protein driven by Cytomegalovirus (CMV) immediate early promoter. The mouse phenotype was similar to dHMN patients in that they exhibit motor neuropathy. To determine the phenotypic aberration of transgenic mice, behavior test, magnetic resonance imaging (MRI), electrophysiological study, and pathology were performed. Rotarod test showed that founder mice exhibited lowered motor performance. MRI also revealed marked fatty infiltration in the anterior and posterior compartments at calf level. Electrophysiologically, compound muscle action potential (CMAP) but not motor nerve conduction velocity (MNCV) was reduced in the transgenic mice. Toluidine staining with semi-thin section of sciatic nerve showed the ratio of large myelinated axon fiber was reduced, which might cause reduced locomotion in the transgenic mice. Electron microscopy also revealed abundant aberrant myelination. Immunohistochemically, neuronal dysfunctions included elevated level of phosphorylated neurofilament and reduced level of acetylated tubulin in the sural nerve of transgenic mice. There was no additional phenotype besides motor neuronal defects. CONCLUSIONS: Overexpression of HSP27-S135F protein causes peripheral neuropathy. The mouse model can be applied to future development of therapeutic strategies for dHMN or CMT2F.
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Enfermedad de Charcot-Marie-Tooth/genética , Proteínas de Choque Térmico/biosíntesis , Atrofia Muscular Espinal/genética , Proteínas de Neoplasias/biosíntesis , Enfermedades del Sistema Nervioso Periférico/genética , Animales , Enfermedad de Charcot-Marie-Tooth/fisiopatología , Modelos Animales de Enfermedad , Proteínas de Choque Térmico/genética , Humanos , Ratones , Ratones Transgénicos , Chaperonas Moleculares , Neuronas Motoras/metabolismo , Neuronas Motoras/patología , Atrofia Muscular Espinal/patología , Mutación , Proteínas de Neoplasias/genética , Enfermedades del Sistema Nervioso Periférico/fisiopatologíaRESUMEN
Abl interactor 1 (Abi1) is a scaffold protein that plays a central role in the regulation of actin cytoskeleton dynamics as a constituent of several key protein complexes, and homozygous loss of this protein leads to embryonic lethality in mice. Because this scaffold protein has been shown in cultured cells to be a critical component of pathways controlling cell migration and actin regulation at cell-cell contacts, we were interested to investigate the in vivo role of Abi1 in morphogenesis during the development of Xenopus embryos. Using morpholino-mediated translation inhibition, we demonstrate that knockdown of Abi1 in the whole embryo, or specifically in eye field progenitor cells, leads to disruption of eye morphogenesis. Moreover, signaling through the Src homology 3 domain of Abi1 is critical for proper movement of retinal progenitor cells into the eye field and their appropriate differentiation, and this process is dependent upon an interaction with the nucleation-promoting factor Wasp (Wiskott-Aldrich syndrome protein). Collectively, our data demonstrate that the Abi1 scaffold protein is an essential regulator of cell movement processes required for normal eye development in Xenopus embryos and specifically requires an Src homology 3 domain-dependent interaction with Wasp to regulate this complex morphogenetic process.
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Proteínas Adaptadoras Transductoras de Señales/metabolismo , Ojo/embriología , Regulación del Desarrollo de la Expresión Génica , Proteína del Síndrome de Wiskott-Aldrich/metabolismo , Proteínas de Xenopus/fisiología , Xenopus/embriología , Citoesqueleto de Actina/metabolismo , Proteínas Adaptadoras Transductoras de Señales/química , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/fisiología , Animales , Linaje de la Célula , Movimiento Celular , Sistemas de Lectura Abierta , Fosforilación , Unión Proteica , Estructura Terciaria de Proteína , Retina/embriología , Transducción de Señal , Células Madre/citología , Xenopus/genética , Proteínas de Xenopus/química , Dominios Homologos srcRESUMEN
BACKGROUND: The incretin effect is known to be decreased in type 2 diabetes. However, there are limited data on the incretin effect in non-Caucasian subjects. Because Asian patients with type 2 diabetes are characterized by decreased insulin secretion, this study set out to examine the incretin effect in Korean subjects with normal glucose tolerance (NGT) or type 2 diabetes. METHODS: We performed 75-g oral glucose tolerance tests (OGTTs) and corresponding isoglycaemic intravenous glucose infusion (IIGI) studies in Korean subjects with NGT (n = 14) or type 2 diabetes (n = 16). The incretin effect was calculated based on the incremental area under the curves (iAUCs) of the plasma levels of insulin, C-peptide or insulin secretion rate (ISR). The plasma levels of total glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP) were measured by ELISA. RESULTS: The incretin effect was not different between the subjects with NGT and type 2 diabetes (43 ± 6% vs 47 ± 4%, P = 0·575 by insulin; 29 ± 7% vs 38 ± 4%, P = 0·253 by C-peptide; 28 ± 7% vs 35 ± 5%, P = 0·372 by ISR, respectively). However, the gastrointestinally mediated glucose disposal (GIGD) was markedly decreased in type 2 diabetes (28·5 ± 4·2% vs 59·0 ± 4·3%, P < 0·001). The plasma levels of the total GLP-1 and GIP during the OGTTs were comparable between the two groups. CONCLUSION: In Koreans, the secretion of GLP-1 or GIP during OGTTs and the incretin effect were comparable between subjects with NGT and type 2 diabetes, whereas the GIGD was significantly decreased in patients with type 2 diabetes.
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Diabetes Mellitus Tipo 2/sangre , Prueba de Tolerancia a la Glucosa , Incretinas/sangre , Adulto , Pueblo Asiatico , Glucemia/metabolismo , Péptido C/sangre , Diabetes Mellitus Tipo 2/etnología , Polipéptido Inhibidor Gástrico/sangre , Glucagón/sangre , Péptido 1 Similar al Glucagón/sangre , Humanos , Insulina/sangre , Insulina/metabolismo , Secreción de Insulina , Masculino , Persona de Mediana Edad , República de CoreaRESUMEN
BACKGROUND: Pervasive usage of alternative promoters leads to the deregulation of gene expression in carcinogenesis and may drive the emergence of new genes in spermatogenesis. However, little is known regarding the mechanisms underpinning the activation of alternative promoters. RESULTS: Here we describe how alternative cancer-testis-specific transcription is activated. We show that intergenic and intronic CTCF binding sites, which are transcriptionally inert in normal somatic cells, could be epigenetically reprogrammed into active de novo promoters in germ and cancer cells. BORIS/CTCFL, the testis-specific paralog of the ubiquitously expressed CTCF, triggers the epigenetic reprogramming of CTCF sites into units of active transcription. BORIS binding initiates the recruitment of the chromatin remodeling factor, SRCAP, followed by the replacement of H2A histone with H2A.Z, resulting in a more relaxed chromatin state in the nucleosomes flanking the CTCF binding sites. The relaxation of chromatin around CTCF binding sites facilitates the recruitment of multiple additional transcription factors, thereby activating transcription from a given binding site. We demonstrate that the epigenetically reprogrammed CTCF binding sites can drive the expression of cancer-testis genes, long noncoding RNAs, retro-pseudogenes, and dormant transposable elements. CONCLUSIONS: Thus, BORIS functions as a transcription factor that epigenetically reprograms clustered CTCF binding sites into transcriptional start sites, promoting transcription from alternative promoters in both germ cells and cancer cells.
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Proteínas de Unión al ADN , Factores de Transcripción , Masculino , Humanos , Proteínas de Unión al ADN/metabolismo , Factor de Unión a CCCTC/metabolismo , Factores de Transcripción/metabolismo , Histonas/metabolismo , Cromatina , Sitios de UniónRESUMEN
BACKGROUND: Sphingolipids are signaling molecules and structural components of the axolemma and myelin sheath. Plasma sphingolipid levels may reflect disease status of neuromyelitis optica spectrum disorder (NMOSD). We aimed to examine plasma sphingolipids as disease severity biomarkers for NMOSD and compare their characteristics with those of serum neurofilament light chain (sNfL) and glial fibrillary acidic protein (sGFAP). METHODS: We measured plasma sphingolipids, sNfL, and sGFAP levels in NMOSD cases with anti-aquaporin-4-antibody. An unbiased approach, partial least square discriminant analysis (PLS-DA), was utilized to determine whether sphingolipid profiles differ according to the disease state of NMOSD (presence, moderate-to-severe disability [Expanded Disease Severity Scale, (EDSS) > 3.0], and relapses). RESULTS: We investigated 81 patients and 10 controls. PLS-DA models utilizing sphingolipids successfully differentiated patients with EDSS > 3.0, but failed to identify the presence of disease and relapses. Ceramide-C14-a significant contributor to differentiating EDSS > 3.0-positively correlated with EDSS, while its levels were independent of age and the presence of relapses. This characteristic was unique from those of sNfL and sGFAP, which were affected by age and relapses as well as EDSS. CONCLUSION: Plasma sphingolipids may be useful NMOSD biomarkers for disability with distinct characteristics compared to sNfL and sGFAP.
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Biomarcadores , Proteínas de Neurofilamentos , Neuromielitis Óptica , Esfingolípidos , Humanos , Neuromielitis Óptica/sangre , Neuromielitis Óptica/diagnóstico , Biomarcadores/sangre , Femenino , Esfingolípidos/sangre , Adulto , Masculino , Persona de Mediana Edad , Proteínas de Neurofilamentos/sangre , Proteína Ácida Fibrilar de la Glía/sangre , Índice de Severidad de la Enfermedad , Acuaporina 4/sangre , Acuaporina 4/inmunologíaRESUMEN
Introduction: CTCF-related disorder (CRD) is a neurodevelopmental disorder (NDD) caused by monoallelic pathogenic variants in CTCF. The first CTCF variants in CRD cases were documented in 2013. To date, 76 CTCF variants have been further described in the literature. In recent years, due to the increased application of next-generation sequencing (NGS), growing numbers of CTCF variants are being identified, and multiple genotype-phenotype databases cataloging such variants are emerging. Methods: In this study, we aimed to expand the genotypic spectrum of CRD, by cataloging NDD phenotypes associated with reported CTCF variants. Here, we systematically reviewed all known CTCF variants reported in case studies and large-scale exome sequencing cohorts. We also conducted a meta-analysis using public variant data from genotype-phenotype databases to identify additional CTCF variants, which we then curated and annotated. Results: From this combined approach, we report an additional 86 CTCF variants associated with NDD phenotypes that have not yet been described in the literature. Furthermore, we describe and explain inconsistencies in the quality of reported variants, which impairs the reuse of data for research of NDDs and other pathologies. Discussion: From this integrated analysis, we provide a comprehensive and annotated catalog of all currently known CTCF mutations associated with NDD phenotypes, to aid diagnostic applications, as well as translational and basic research.
RESUMEN
Since the emergence of SARS-CoV-2, dozens of variants of interest and half a dozen variants of concern (VOCs) have been documented by the World Health Organization. The emergence of these VOCs due to the continuous evolution of the virus is a major concern for COVID-19 therapeutic antibodies and vaccines because they are designed to target prototype/previous strains and lose effectiveness against new VOCs. Therefore, there is a need for time- and cost-effective strategies to estimate the immune escape and redirect therapeutic antibodies against newly emerging variants. Here, we computationally predicted the neutralization escape of the SARS-CoV-2 Delta and Omicron variants against the mutational space of RBD-mAbs interfaces. Leveraging knowledge of the existing RBD-mAb interfaces and mutational space, we fine-tuned and redirected CT-p59 (Regdanvimab) and Etesevimab against the escaped variants through complementarity-determining regions (CDRs) diversification. We identified antibodies against the Omicron lineage BA.1 and BA.2 and Delta variants with comparable or better binding affinities to that of prototype Spike. This suggests that CDRs diversification by hotspot grafting, given an existing insight into the Ag-Abs interface, is an exquisite strategy to redirect antibodies against preselected epitopes and combat the neutralization escape of emerging SARS-CoV-2 variants.
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COVID-19 , SARS-CoV-2 , Humanos , SARS-CoV-2/genética , Anticuerpos Monoclonales/uso terapéutico , Regiones Determinantes de Complementariedad/genéticaRESUMEN
Tonsils comprise part of the mucosal immune system and contain lymphocytes, macrophages, and follicular dendritic cells (FDCs). FDCs are located in the B cell area of the follicles of secondary lymphoid organs, such as the spleen, tonsils, or lymph nodes, and they trap and retain immune complexes on their surfaces to regulate B cell activation and maturation. Stromal cells from the palatine tonsils are often used for FDC in vitro studies, and it has been reported that human palatine tonsils may be a good source of multipotent mesenchymal cells. Therefore, we assessed whether tonsil-derived mesenchymal stromal cells could differentiate into a FDC-like phenotype. We discovered that stromal cells isolated from human tonsils not only had the potential to differentiate into various cell types of mesenchymal origin, but they also could differentiate into FDC-like cells under cytokine stimulation in vitro.
Asunto(s)
Citocinas/farmacología , Células Dendríticas Foliculares/citología , Células Madre Mesenquimatosas/citología , Tonsila Palatina/citología , Antígenos de Superficie/metabolismo , Diferenciación Celular/efectos de los fármacos , Células Cultivadas , Células Dendríticas Foliculares/efectos de los fármacos , Células Dendríticas Foliculares/metabolismo , Citometría de Flujo , Humanos , Células Madre Mesenquimatosas/efectos de los fármacos , Células Madre Mesenquimatosas/metabolismo , Fenotipo , Células del Estroma/citología , Células del Estroma/efectos de los fármacos , Células del Estroma/metabolismoRESUMEN
BACKGROUND AIMS: Although mesenchymal stromal cells (MSC) from human palatine tonsils (tonsillar MSC, T-MSC) have been isolated, whether T-MSC isolated from multiple donors are feasible for cell banking has not been studied. METHODS: T-MSC before and after a standard protocol of cryopreservation and thawing were assessed regarding several basic characteristics, including colony-forming unit-fibroblast features, MSC-specific surface antigen profiles, and inhibition of alloreactive T-cell proliferation. In vitro mesodermal differentiation potentials to adipocytes, osteocytes and chondrocytes were detected by staining with either cell-specific dyes or antibody after incubation with each appropriate differentiation medium. Expression of mesoderm-specific genes was also quantified by real-time polymerase chain reaction (PCR) assay. Expression profiles of endoderm-specific genes were identified by reverse transcription PCR assay. The feasibility of T-MSC in future engraftment was tested by short tandem repeat (STR) analysis using genomic DNA isolated randomly from three independent subjects. RESULTS: Both fresh and cryopreserved-thawed T-MSC showed a similar high proliferation capacity and expressed primitive cell-surface markers. Hematopoietic cell markers, HLA-DR, co-stimulatory molecules and follicular dendritic cell markers were not detected. In addition to mesodermal differentiation, fresh and cryopreserved-thawed cells also underwent endodermal differentiation, as evidenced by the expression of endoderm-specific genes including forkhead box A2 (FoxA2), SIX homeobox 1 (Six1) and chemokine (C-C motif) ligand 21 (CCL21). Both cells significantly decreased phorbol 12- myristate 13-acetate (PMA)-induced T-cell proliferation. T-MSC from three independent donors formed chimerism in STR analysis. CONCLUSIONS: Our results demonstrate for the first time that T-MSC are a potentially good source for MSC banking.
Asunto(s)
Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/inmunología , Tonsila Palatina/citología , Bancos de Tejidos , Antígenos de Superficie/metabolismo , Diferenciación Celular/genética , Linaje de la Célula/genética , Proliferación Celular , Células Cultivadas , Niño , Quimerismo , Ensayo de Unidades Formadoras de Colonias , Endodermo/citología , Endodermo/metabolismo , Femenino , Fibroblastos/citología , Humanos , Masculino , Células Madre Mesenquimatosas/metabolismo , Mesodermo/citología , Mesodermo/metabolismo , Fenotipo , Linfocitos T/citología , Linfocitos T/inmunología , Donantes de TejidosAsunto(s)
Antígeno B7-H1/metabolismo , Queratinocitos/inmunología , Psoriasis/inmunología , Aminoquinolinas , Animales , Línea Celular Transformada , Dermatitis/inmunología , Dermatitis/metabolismo , Modelos Animales de Enfermedad , Humanos , Imiquimod , Queratinocitos/metabolismo , Ratones , Poli I-C , Psoriasis/metabolismo , Receptor Toll-Like 3/metabolismoRESUMEN
OBJECTIVE: The objective of this review is to characterize the state of literature regarding forcibly displaced persons' sexual and reproductive health in urban areas in low- and middle-income countries. Specific objectives include describing the sexual and reproductive health outcomes among forcibly displaced persons relocating in urban environments. INTRODUCTION: As a result of persecution, conflict, violence, human rights violations, and disruptive events, 89.3 million people worldwide were forcibly displaced as of the end of 2021. Forcibly displaced people face a wide range of sexual and reproductive health challenges in their countries of origin, en route to final destinations, and on arrival in host communities. There is a growing urbanization of forcibly displaced persons, yet there is limited attention on sexual and reproductive health outcomes of this population. INCLUSION CRITERIA: This review will consider studies that include sexual and/or reproductive health outcomes and needs of forcibly displaced persons within urban environments in low- and middle-income countries. Published and unpublished evidence, including quantitative, qualitative, mixed methods research, and gray literature, will be eligible for inclusion. METHODS: MEDLINE, Embase, PsycINFO, CINAHL, IBSS, ASSIA, SSCI, and Global Medicus Index will be searched for English-language articles. Titles and abstracts will be screened against the inclusion criteria, followed by full-text review of potentially eligible studies, which will be independently assessed by 2 reviewers. Eligible articles will be extracted and charted. Results from extracted data will be tabulated and accompanied by a narrative summary to summarize and contextualize the extracted data to describe how the results relate to the review's objectives and question.