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1.
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
2.
Int J Mol Med ; 47(4): 1, 2021 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-33537817

RESUMEN

Inflammation is the most common cause of most acute and chronic debilitating diseases. Towards unveiling novel therapeutic options for patients with such complications, N­bromotaurine (TauNHBr) has emerged as a potential anti­inflammatory agent; however, its therapeutic efficacy is hindered due to its relatively poor stability. To address this challenge, the present study focused on examining the effects of a stable active bromine compound, named bromamine T (BAT). The present study examined the protective properties of BAT against lipopolysaccharide (LPS)­mediated inflammation in vitro, by using LPS­stimulated murine J774.A1 macrophages (Mφs), as well as in vivo, by using a murine LPS­mediated air­pouch model. Additionally, its efficacy was compared with that of taurine, a known potent anti­inflammatory molecule. In LPS­stimulated J774A.1 Mφs, BAT and taurine were very effective in reducing the secretion of pro­inflammatory mediators. The in vitro experiments indicated that LPS­mediated inflammation was attenuated due to the protective properties of BAT and of taurine, probably through the inhibition of phosphorylated p65 NF­κB subunit (Ser 536) nuclear translocation. The in vivo experiments also revealed that BAT and taurine inhibited LPS­mediated inflammation by reducing total cell/polymorphonuclear cell (PMN) infiltration in the air­pouch and by decreasing pouch wall thickness. The analysis of exudates obtained from pouches highlighted that the inhibitory effects of BAT and taurine on the secretion of pro­inflammatory cytokines were similar to those observed in vitro. Notably, the effect of BAT at the highest concentration tested was superior to that of taurine at the highest concentration. Taken together, the findings of the present study indicate that BAT prevents the LPS­induced inflammatory response both in vitro and in vivo.


Asunto(s)
Bromo/uso terapéutico , Inflamación/tratamiento farmacológico , Sulfonamidas/uso terapéutico , Animales , Antiinflamatorios/farmacología , Antiinflamatorios/uso terapéutico , Bromo/farmacología , Línea Celular , Núcleo Celular/efectos de los fármacos , Núcleo Celular/metabolismo , Modelos Animales de Enfermedad , Regulación de la Expresión Génica/efectos de los fármacos , Inflamación/patología , Mediadores de Inflamación/metabolismo , Lipopolisacáridos , Ratones Endogámicos C57BL , Fosforilación/efectos de los fármacos , Transporte de Proteínas/efectos de los fármacos , ARN Mensajero/genética , ARN Mensajero/metabolismo , Sulfonamidas/farmacología , Taurina/farmacología , Factor de Transcripción ReIA/metabolismo , Transcripción Genética/efectos de los fármacos
3.
Int J Mol Sci ; 22(3)2021 Feb 02.
Artículo en Inglés | MEDLINE | ID: mdl-33540814

RESUMEN

Vascular smooth muscle cells (VSMCs) provide vital contractile force within blood vessel walls, yet can also propagate cardiovascular pathologies through proliferative and pro-inflammatory activities. Such phenotypes are driven, in part, by the diverse effects of long non-coding RNAs (lncRNAs) on gene expression. However, lncRNA characterisation in VSMCs in pathological states is hampered by incomplete lncRNA representation in reference annotation. We aimed to improve lncRNA representation in such contexts by assembling non-reference transcripts in RNA sequencing datasets describing VSMCs stimulated in vitro with cytokines, growth factors, or mechanical stress, as well as those isolated from atherosclerotic plaques. All transcripts were then subjected to a rigorous lncRNA prediction pipeline. We substantially improved coverage of lncRNAs responding to pro-mitogenic stimuli, with non-reference lncRNAs contributing 21-32% for each dataset. We also demonstrate non-reference lncRNAs were biased towards enriched expression within VSMCs, and transcription from enhancer sites, suggesting particular relevance to VSMC processes, and the regulation of neighbouring protein-coding genes. Both VSMC-enriched and enhancer-transcribed lncRNAs were large components of lncRNAs responding to pathological stimuli, yet without novel transcript discovery 33-46% of these lncRNAs would remain hidden. Our comprehensive VSMC lncRNA repertoire allows proper prioritisation of candidates for characterisation and exemplifies a strategy to broaden our knowledge of lncRNA across a range of disease states.


Asunto(s)
Músculo Liso Vascular/citología , Miocitos del Músculo Liso/metabolismo , Placa Aterosclerótica/metabolismo , ARN Largo no Codificante/análisis , Aorta/citología , Vasos Coronarios/citología , Citocinas/farmacología , Conjuntos de Datos como Asunto , Elementos de Facilitación Genéticos , Perfilación de la Expresión Génica , Humanos , Péptidos y Proteínas de Señalización Intercelular/farmacología , Músculo Liso Vascular/metabolismo , Miocitos del Músculo Liso/efectos de los fármacos , ARN Largo no Codificante/aislamiento & purificación , RNA-Seq , Estrés Mecánico , Transcripción Genética/efectos de los fármacos , Transcriptoma
4.
Nat Commun ; 12(1): 223, 2021 01 11.
Artículo en Inglés | MEDLINE | ID: mdl-33431820

RESUMEN

Enhancers are DNA sequences that enable complex temporal and tissue-specific regulation of genes in higher eukaryotes. Although it is not entirely clear how enhancer-promoter interactions can increase gene expression, this proximity has been observed in multiple systems at multiple loci and is thought to be essential for the maintenance of gene expression. Bromodomain and Extra-Terminal domain (BET) and Mediator proteins have been shown capable of forming phase condensates and are thought to be essential for super-enhancer function. Here, we show that targeting of cells with inhibitors of BET proteins or pharmacological degradation of BET protein Bromodomain-containing protein 4 (BRD4) has a strong impact on transcription but very little impact on enhancer-promoter interactions. Dissolving phase condensates reduces BRD4 and Mediator binding at enhancers and can also strongly affect gene transcription, without disrupting enhancer-promoter interactions. These results suggest that activation of transcription and maintenance of enhancer-promoter interactions are separable events. Our findings further indicate that enhancer-promoter interactions are not dependent on high levels of BRD4 and Mediator, and are likely maintained by a complex set of factors including additional activator complexes and, at some sites, CTCF and cohesin.


Asunto(s)
Elementos de Facilitación Genéticos , Regiones Promotoras Genéticas , Transcripción Genética , Factor de Unión a CCCTC/metabolismo , Carcinogénesis/efectos de los fármacos , Carcinogénesis/genética , Proteínas de Ciclo Celular/metabolismo , Línea Celular Tumoral , Cromatina/metabolismo , Proteínas Cromosómicas no Histona/metabolismo , Proteínas de Unión al ADN/metabolismo , Glicoles/farmacología , Histonas/metabolismo , Humanos , Leucemia/genética , Leucemia/patología , Modelos Genéticos , Unión Proteica/efectos de los fármacos , Proteínas Proto-Oncogénicas c-myc/genética , Transcripción Genética/efectos de los fármacos
5.
Nat Commun ; 12(1): 301, 2021 01 12.
Artículo en Inglés | MEDLINE | ID: mdl-33436596

RESUMEN

Macrophages are innate immune cells that contribute to fighting infections, tissue repair, and maintaining tissue homeostasis. To enable such functional diversity, macrophages resolve potentially conflicting cues in the microenvironment via mechanisms that are unclear. Here, we use single-cell RNA sequencing to explore how individual macrophages respond when co-stimulated with inflammatory stimuli LPS and IFN-γ and the resolving cytokine IL-4. These co-stimulated macrophages display a distinct global transcriptional program. However, variable negative cross-regulation between some LPS + IFN-γ-specific and IL-4-specific genes results in cell-to-cell heterogeneity in transcription. Interestingly, negative cross-regulation leads to mutually exclusive expression of the T-cell-polarizing cytokine genes Il6 and Il12b versus the IL-4-associated factors Arg1 and Chil3 in single co-stimulated macrophages, and single-cell secretion measurements show that these specialized functions are maintained for at least 48 h. This study suggests that increasing functional diversity in the population is one strategy macrophages use to respond to conflicting environmental cues.


Asunto(s)
Polaridad Celular , Macrófagos/citología , Animales , Arginasa/metabolismo , Polaridad Celular/efectos de los fármacos , Polaridad Celular/genética , Femenino , Regulación de la Expresión Génica/efectos de los fármacos , Interferón gamma/farmacología , Interleucina-12/metabolismo , Interleucina-6/metabolismo , Lipopolisacáridos/farmacología , Aprendizaje Automático , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Masculino , Ratones Endogámicos C57BL , Redes Neurales de la Computación , Oportunidad Relativa , Análisis de la Célula Individual , Factores de Transcripción/metabolismo , Transcripción Genética/efectos de los fármacos
6.
Int J Mol Sci ; 22(2)2021 Jan 19.
Artículo en Inglés | MEDLINE | ID: mdl-33478005

RESUMEN

The objective was to investigate the anti-cancer effects and underlying molecular mechanisms of cytostasis which were activated by an anti-microtubule drug, ABT-751, in two urinary bladder urothelial carcinoma (UBUC)-derived cell lines, BFTC905 and J82, with distinct genetic backgrounds. A series of in vitro assays demonstrated that ABT-751 induced G2/M cell cycle arrest, decreased cell number in the S phase of the cell cycle and suppressed colony formation/independent cell growth, accompanied with alterations of the protein levels of several cell cycle regulators. In addition, ABT-751 treatment significantly hurdled cell migration and invasion along with the regulation of epithelial-mesenchymal transition-related proteins. ABT-751 triggered autophagy and apoptosis, downregulated the mechanistic target of rapamycin kinase (MTOR) and upregulated several pro-apoptotic proteins that are involved in extrinsic and intrinsic apoptotic pathways. Inhibition of autophagosome and autolysosome enhanced apoptosis was also observed. Through the inhibition of the NFκB signaling pathway, ABT-751 suppressed S-phase kinase associated protein 2 (SKP2) transcription and subsequent translation by downregulation of active/phospho-AKT serine/threonine kinase 1 (AKT1), component of inhibitor of nuclear factor kappa B kinase complex (CHUK), NFKB inhibitor alpha (NFKBIA), nuclear RELA proto-oncogene, NFκB subunit (RELA) and maintained a strong interaction between NFKBIA and RELA to prevent RELA nuclear translocation for SKP2 transcription. ABT-751 downregulated stable/phospho-SKP2 including pSKP2(S64) and pSKP2(S72), which targeted cyclin-dependent kinase inhibitors for degradation through the inactivation of AKT. Our results suggested that ABT-751 may act as an anti-cancer drug by inhibiting cell migration, invasion yet inducing cell cycle arrest, autophagy and apoptosis in distinct UBUC-derived cells. Particularly, the upstream molecular mechanism of its anticancer effects was identified as ABT-751-induced cytostasis through the inhibition of SKP2 at both transcriptional and post-translational levels to stabilize cyclin dependent kinase inhibitor 1A (CDKN1A) and CDKN1B proteins.


Asunto(s)
Carcinoma/tratamiento farmacológico , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/genética , Inhibidor p27 de las Quinasas Dependientes de la Ciclina/genética , Proteínas Quinasas Asociadas a Fase-S/genética , Transcripción Genética/efectos de los fármacos , Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Autofagia/efectos de los fármacos , Carcinoma/genética , Carcinoma/patología , Puntos de Control del Ciclo Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Fosfatidilinositol 3-Quinasa/genética , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Quinasas Asociadas a Fase-S/antagonistas & inhibidores , Transducción de Señal/efectos de los fármacos , Sulfonamidas/farmacología , Vejiga Urinaria/efectos de los fármacos , Vejiga Urinaria/patología , Urotelio/efectos de los fármacos , Urotelio/patología
7.
J Med Chem ; 64(1): 42-70, 2021 01 14.
Artículo en Inglés | MEDLINE | ID: mdl-33355454

RESUMEN

Guanine-rich DNA sequences have the propensity to adopt four-stranded tetrahelical G-quadruplex (G4) structures that are overrepresented in gene promoters. The structural polymorphism and physicochemical properties of these non-Watson-Crick G4 structures make them important targets for drug development. The guanine-rich nuclease hypersensitivity element III1 present in the upstream of P1 promoter of c-MYC oncogene has the ability to form an intramolecular parallel G4 structure. The G4 structure that forms transiently in the c-MYC promoter functions as a transcriptional repressor element. The c-MYC oncogene is overexpressed in a wide variety of cancers and plays a key role in cancer progression. Till now, a large number of compounds that are capable of interacting and stabilizing thec-MYC G4 have been reported. In this review, we summarize various c-MYC G4 specific molecules and discuss their effects on c-MYC gene expression in vitro and in vivo.


Asunto(s)
Antineoplásicos/farmacología , G-Cuádruplex , Genes myc , Bibliotecas de Moléculas Pequeñas/farmacología , Sitios de Unión , Sistemas de Liberación de Medicamentos , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Ligandos , Regiones Promotoras Genéticas , Transcripción Genética/efectos de los fármacos
8.
Nature ; 588(7839): 712-716, 2020 12.
Artículo en Inglés | MEDLINE | ID: mdl-33328633

RESUMEN

Altered expression of mitochondrial DNA (mtDNA) occurs in ageing and a range of human pathologies (for example, inborn errors of metabolism, neurodegeneration and cancer). Here we describe first-in-class specific inhibitors of mitochondrial transcription (IMTs) that target the human mitochondrial RNA polymerase (POLRMT), which is essential for biogenesis of the oxidative phosphorylation (OXPHOS) system1-6. The IMTs efficiently impair mtDNA transcription in a reconstituted recombinant system and cause a dose-dependent inhibition of mtDNA expression and OXPHOS in cell lines. To verify the cellular target, we performed exome sequencing of mutagenized cells and identified a cluster of amino acid substitutions in POLRMT that cause resistance to IMTs. We obtained a cryo-electron microscopy (cryo-EM) structure of POLRMT bound to an IMT, which further defined the allosteric binding site near the active centre cleft of POLRMT. The growth of cancer cells and the persistence of therapy-resistant cancer stem cells has previously been reported to depend on OXPHOS7-17, and we therefore investigated whether IMTs have anti-tumour effects. Four weeks of oral treatment with an IMT is well-tolerated in mice and does not cause OXPHOS dysfunction or toxicity in normal tissues, despite inducing a strong anti-tumour response in xenografts of human cancer cells. In summary, IMTs provide a potent and specific chemical biology tool to study the role of mtDNA expression in physiology and disease.


Asunto(s)
Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Bibliotecas de Moléculas Pequeñas/química , Bibliotecas de Moléculas Pequeñas/farmacología , Transcripción Genética/efectos de los fármacos , Animales , Proliferación Celular/efectos de los fármacos , Microscopía por Crioelectrón , ADN Mitocondrial/efectos de los fármacos , ADN Mitocondrial/genética , ARN Polimerasas Dirigidas por ADN/metabolismo , Regulación hacia Abajo/efectos de los fármacos , Estabilidad de Enzimas/efectos de los fármacos , Femenino , Regulación de la Expresión Génica/efectos de los fármacos , Genes Mitocondriales/efectos de los fármacos , Humanos , Masculino , Ratones , Neoplasias/tratamiento farmacológico , Neoplasias/patología , Especificidad por Sustrato/efectos de los fármacos , Ensayos Antitumor por Modelo de Xenoinjerto
9.
Cell Death Dis ; 11(12): 1052, 2020 12 11.
Artículo en Inglés | MEDLINE | ID: mdl-33311440

RESUMEN

The abnormal PI3K/AKT/mTOR pathway is one of the most common genomic abnormalities in breast cancers including triple-negative breast cancer (TNBC), and pharmacologic inhibition of these aberrations has shown activity in TNBC patients. Here, we designed and identified a small-molecule Comp34 that suppresses both AKT and mTOR protein expression and exhibits robust cytotoxicity towards TNBC cells but not nontumorigenic normal breast epithelial cells. Mechanically, long noncoding RNA (lncRNA) AL354740.1-204 (also named as NUDT3-AS4) acts as a microRNA sponge to compete with AKT1/mTOR mRNAs for binding to miR-99s, leading to decrease in degradation of AKT1/mTOR mRNAs and subsequent increase in AKT1/mTOR protein expression. Inhibition of lncRNA-NUDT3-AS4 and suppression of the NUDT3-AS4/miR-99s association contribute to Comp34-affected biologic pathways. In addition, Comp34 alone is effective in cells with secondary resistance to rapamycin, the best-known inhibitor of mTOR, and displays a greater in vivo antitumor efficacy and lower toxicity than rapamycin in TNBC xenografted models. In conclusion, NUDT3-AS4 may play a proproliferative role in TNBC and be considered a relevant therapeutic target, and Comp34 presents promising activity as a single agent to inhibit TNBC through regulation of NUDT3-AS4 and miR-99s.


Asunto(s)
Antineoplásicos/uso terapéutico , Curcumina/uso terapéutico , ARN Largo no Codificante/metabolismo , Neoplasias de la Mama Triple Negativas/tratamiento farmacológico , Neoplasias de la Mama Triple Negativas/genética , Antineoplásicos/farmacología , Secuencia de Bases , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Curcumina/química , Curcumina/farmacología , Progresión de la Enfermedad , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , MicroARNs/genética , MicroARNs/metabolismo , Células Madre Neoplásicas/efectos de los fármacos , Células Madre Neoplásicas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , ARN Largo no Codificante/genética , Transducción de Señal/efectos de los fármacos , Sirolimus/farmacología , Serina-Treonina Quinasas TOR/metabolismo , Transcripción Genética/efectos de los fármacos
10.
Nat Commun ; 11(1): 6430, 2020 12 22.
Artículo en Inglés | MEDLINE | ID: mdl-33353937

RESUMEN

The trp operon of Chlamydia trachomatis is organized differently from other model bacteria. It contains trpR, an intergenic region (IGR), and the biosynthetic trpB and trpA open-reading frames. TrpR is a tryptophan-dependent repressor that regulates the major promoter (PtrpR), while the IGR harbors an alternative promoter (PtrpBA) and an operator sequence for the iron-dependent repressor YtgR to regulate trpBA expression. Here, we report that YtgR repression at PtrpBA is also dependent on tryptophan by regulating YtgR levels through a rare triple-tryptophan motif (WWW) in the YtgCR precursor. Inhibiting translation during tryptophan limitation at the WWW motif subsequently promotes Rho-independent transcription termination of ytgR, thereby de-repressing PtrpBA. Thus, YtgR represents an alternative strategy to attenuate trpBA expression, expanding the repertoire for trp operon attenuation beyond TrpL- and TRAP-mediated mechanisms described in other bacteria. Furthermore, repurposing the iron-dependent repressor YtgR underscores the fundamental importance of maintaining tryptophan-dependent attenuation of the trpRBA operon.


Asunto(s)
Proteínas Bacterianas/metabolismo , Chlamydia trachomatis/genética , Hierro/metabolismo , Operón/genética , Triptófano/metabolismo , Secuencias de Aminoácidos , Proteínas Bacterianas/química , Proteínas Bacterianas/genética , Chlamydia trachomatis/efectos de los fármacos , Regulación Bacteriana de la Expresión Génica/efectos de los fármacos , Células HeLa , Humanos , Indoles/farmacología , Modelos Biológicos , Regiones Promotoras Genéticas , Biosíntesis de Proteínas/efectos de los fármacos , Dominios Proteicos , ARN de Transferencia de Triptófano/metabolismo , Transcripción Genética/efectos de los fármacos , Proteínas de Unión al GTP rho/metabolismo
11.
J Toxicol Sci ; 45(10): 619-624, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-33012730

RESUMEN

Manganese (Mn) poisoning may result in a neurological disorder called manganism. Although the neurotoxic mechanism of Mn is unclear, oxidative stress may be involved based on the interactions between neurotransmitter catecholamines and metals such as iron. Here, we propose a novel mechanism in which Mn oxidizes catecholamines and inhibits cellular transcription. Mn accelerated the oxidation of adrenaline (Ad) and produced adrenochrome (AdC) more effectively than iron. Furthermore, the oxidation of DNA bases increased when Ad, Mn, and iron were present. However, despite the absence of iron, cell viability decreased in the presence of AdC or Ad with Mn, which suggests there is another mechanism independent of oxidative DNA damage. AdC or preincubated Ad with Mn reduced mRNA synthesis in T7 RNA polymerase-driven transcription. RNA synthesis decreased in AdC-treated cells dose-dependently. These results show that Mn disrupts neuronal function via catecholamine oxidation-mediated transcriptional inhibition.


Asunto(s)
Catecolaminas/genética , Catecolaminas/metabolismo , Intoxicación por Manganeso , Manganeso/toxicidad , Transcripción Genética/efectos de los fármacos , Adrenocromo/metabolismo , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Daño del ADN/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Epinefrina/metabolismo , Humanos , Hierro/metabolismo , Oxidación-Reducción/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , ARN Mensajero/metabolismo
12.
Mol Cell ; 80(2): 210-226.e7, 2020 10 15.
Artículo en Inglés | MEDLINE | ID: mdl-33002424

RESUMEN

Many bacterial pathogens regulate their virulence genes via phase variation, whereby length-variable simple sequence repeats control the transcription or coding potential of those genes. Here, we have exploited this relationship between DNA structure and physiological function to discover a globally acting small RNA (sRNA) regulator of virulence in the gastric pathogen Helicobacter pylori. Our study reports the first sRNA whose expression is affected by a variable thymine (T) stretch in its promoter. We show the sRNA post-transcriptionally represses multiple major pathogenicity factors of H. pylori, including CagA and VacA, by base pairing to their mRNAs. We further demonstrate transcription of the sRNA is regulated by the nickel-responsive transcriptional regulator NikR (thus named NikS for nickel-regulated sRNA), thereby linking virulence factor regulation to nickel concentrations. Using in-vitro infection experiments, we demonstrate NikS affects host cell internalization and epithelial barrier disruption. Together, our results show NikS is a phase-variable, post-transcriptional global regulator of virulence properties in H. pylori.


Asunto(s)
Helicobacter pylori/genética , Helicobacter pylori/patogenicidad , ARN Bacteriano/genética , Secuencias Repetitivas de Ácidos Nucleicos/genética , Factores de Virulencia/metabolismo , Proteínas Bacterianas/metabolismo , Secuencia de Bases , Recuento de Colonia Microbiana , Endocitosis/efectos de los fármacos , Eliminación de Gen , Regulación Bacteriana de la Expresión Génica/efectos de los fármacos , Helicobacter pylori/efectos de los fármacos , Interacciones Huésped-Patógeno/efectos de los fármacos , Níquel/farmacología , Fenotipo , Regiones Promotoras Genéticas , ARN Mensajero/genética , ARN Mensajero/metabolismo , Transcripción Genética/efectos de los fármacos
13.
Aquat Toxicol ; 228: 105650, 2020 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-33035767

RESUMEN

Micro(nano)plastics (MPs/NPs) are already present as contaminants in the natural environment globally and have been shown to be difficult to degrade, resulting in the potential for ecological damage and public health concerns. However, the adverse effects of exposure to MPs/NPs by aquatic organisms, especially freshwater microalgae, remains unclear. In the present study, the growth, physiology and transcriptome of the freshwater microalgae Euglena gracilis were comprehensively analyzed following exposure to 1 mg/L of polystyrene (PS) microbeads (5 µm PS-MPs and 100 nm PS-NPs), 0.5 mg/L cadmium (Cd), or a mixture of PS microbeads and Cd for 96 h. Results showed that the toxicity of PS-MPs to microalgae was greater than PS-NPs, inducing increased growth inhibition, oxidative damage and decreased photosynthesis pigment concentrations. PS-MPs alone or in combination with Cd caused cavitation within microalgal cells, as well as increasing the number and volume of vacuoles. The combined exposure toxicity test showed that a combination of Cd + PS-NPs was more toxic than Cd + PS-MPs, which may be explained by the transcriptomic analysis results. Differentially expressed genes (DEGs) in the Cd + PS-NPs group were mainly enriched in metabolism-related pathways, suggesting that algal metabolism was hindered, resulting in aggravation of toxicity. The reduced toxicity induced by Cd + PS-MPs may indicate a response to resist external stress processes. In addition, no adsorption of 0.5 mg/L Cd to 1 mg/L PS microbeads was observed, suggesting that adsorption of MPs/NPs and Cd was not the key factor determining the combined toxicity effects in this study.


Asunto(s)
Cadmio/toxicidad , Exposición a Riesgos Ambientales , Euglena gracilis/genética , Euglena gracilis/fisiología , Microalgas/genética , Microesferas , Poliestirenos/toxicidad , Transcripción Genética/efectos de los fármacos , Organismos Acuáticos/efectos de los fármacos , Organismos Acuáticos/genética , Organismos Acuáticos/crecimiento & desarrollo , Euglena gracilis/efectos de los fármacos , Euglena gracilis/ultraestructura , Perfilación de la Expresión Génica , Ontología de Genes , Microalgas/efectos de los fármacos , Microalgas/fisiología , Microalgas/ultraestructura , Estrés Oxidativo/efectos de los fármacos , Fotosíntesis/efectos de los fármacos , Pigmentos Biológicos/metabolismo , Contaminantes Químicos del Agua/toxicidad
14.
Anticancer Res ; 40(11): 6051-6062, 2020 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-33109543

RESUMEN

BACKGROUND/AIM: Chemoresistance is a major obstacle in the treatment of prostate cancer (PCa). It is imperative to develop novel strategies for overcoming chemoresistance and improving clinical outcomes. We evaluated the in vitro activity and mechanism of action of dihydroergocristine (DHECS), an ergot alkaloid approved for the treatment of dementia, in PCa cells. MATERIALS AND METHODS: The in vitro effects of DHECS on PCa cell cycle and viability were determined by flow cytometry and colorimetric assay. The effects of DHECS on PCa cell signaling were evaluated by quantitative PCR, western blot analysis and reporter assay. RESULTS: DHECS was effective in inducing cell cycle arrest and apoptosis in human PCa cells. Of particular interest, DHECS demonstrated high potency against chemoresistant PCa cells. At the molecular level, DHECS affected multiple factors implicated in the regulation of cancer cell cycle and programmed cell death, including p53, mouse double minute 2 homolog (MDM2), retinoblastoma protein (RB), p21, E2F transcription factor 1 (E2F1), survivin, myeloid cell leukemia 1 (Mcl-1) and poly ADP ribose polymerase (PARP). Furthermore, DHECS may function through dopamine receptor-mediated effects on 5'-AMP-activated protein kinase (AMPK) and nuclear factor kappa B (NF-ĸB). CONCLUSION: DHECS has the potential to be repurposed as a novel anticancer agent for the management of chemoresistant PCa.


Asunto(s)
Dihidroergocristina/farmacología , Resistencia a Antineoplásicos/efectos de los fármacos , Neoplasias de la Próstata/patología , Apoptosis/efectos de los fármacos , Ciclo Celular/efectos de los fármacos , Muerte Celular/efectos de los fármacos , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Humanos , Masculino , Neoplasias de la Próstata/genética , Receptores Dopaminérgicos/metabolismo , Transducción de Señal/efectos de los fármacos , Survivin/genética , Survivin/metabolismo , Transcripción Genética/efectos de los fármacos
15.
Ecotoxicol Environ Saf ; 203: 110961, 2020 Oct 15.
Artículo en Inglés | MEDLINE | ID: mdl-32888621

RESUMEN

Cadmium (Cd), which seriously affects plant growth and crop production, is harmful to humans. Previous studies revealed ryegrass (Lolium multiflorum Lam.) exhibits Cd tolerance, and may be useful as a potential hyperaccumulator because of its wide distribution. In this study, the physiological and transcriptional responses of two ryegrass cultivars [i.e., high (LmHC) and low (LmLC) Cd tolerance] to Cd stress were investigated and compared. The Cd tolerance of LmHC was greater than that of LmLC at various Cd concentrations. The uptake of Evans blue dye revealed that Cd-induced root cell mortality was higher in LmLC than in LmHC after a 12-h Cd treatment. Furthermore, the content and influx rate of Cd in LmLC roots were greater than in LmHC roots under Cd stress conditions. The RNA sequencing and quantitative real-time PCR data indicated that the Cd transport regulatory genes (ABCG37, ABCB4, NRAMP4, and HMA5) were differentially expressed between the LmLC and LmHC roots. This expression-level diversity may contribute to the differences in the Cd accumulation and translocation between LmLC and LmHC. These findings may help clarify the physiological and molecular mechanisms underlying ryegrass responses to Cd toxicity. Additionally, ryegrass may be able to hyperaccumulate toxic heavy metals during the phytoremediation of contaminated soil.


Asunto(s)
Adaptación Biológica , Cadmio/metabolismo , Lolium/efectos de los fármacos , Raíces de Plantas/efectos de los fármacos , Contaminantes del Suelo/metabolismo , Transcripción Genética/efectos de los fármacos , Adaptación Biológica/efectos de los fármacos , Adaptación Biológica/genética , Biodegradación Ambiental , Cadmio/análisis , Cadmio/toxicidad , Genes de Plantas , Lolium/química , Lolium/genética , Raíces de Plantas/química , Raíces de Plantas/genética , Contaminantes del Suelo/análisis , Contaminantes del Suelo/toxicidad
16.
Nat Commun ; 11(1): 4902, 2020 09 29.
Artículo en Inglés | MEDLINE | ID: mdl-32994402

RESUMEN

Living cells and tissues experience various complex modes of forces that are important in physiology and disease. However, how different force modes impact gene expression is elusive. Here we apply local forces of different modes via a magnetic bead bound to the integrins on a cell and quantified cell stiffness, chromatin deformation, and DHFR (dihydrofolate reductase) gene transcription. In-plane stresses result in lower cell stiffness than out-of-plane stresses that lead to bead rolling along the cell long axis (i.e., alignment of actin stress fibers) or at different angles (90° or 45°). However, chromatin stretching and ensuing DHFR gene upregulation by the in-plane mode are similar to those induced by the 45° stress mode. Disrupting stress fibers abolishes differences in cell stiffness, chromatin stretching, and DHFR gene upregulation under different force modes and inhibiting myosin II decreases cell stiffness, chromatin deformation, and gene upregulation. Theoretical modeling using discrete anisotropic stress fibers recapitulates experimental results and reveals underlying mechanisms of force-mode dependence. Our findings suggest that forces impact biological responses of living cells such as gene transcription via previously underappreciated means.


Asunto(s)
Cromatina/química , Fibras de Estrés/química , Tetrahidrofolato Deshidrogenasa/genética , Transcripción Genética/fisiología , Regulación hacia Arriba/fisiología , Animales , Anisotropía , Fenómenos Biomecánicos/genética , Células CHO , Cromatina/metabolismo , Cricetulus , Compuestos Heterocíclicos de 4 o más Anillos/farmacología , Microscopía Intravital , Microscopía Fluorescente , Miosina Tipo II/antagonistas & inhibidores , Miosina Tipo II/metabolismo , Fibras de Estrés/efectos de los fármacos , Fibras de Estrés/metabolismo , Estrés Mecánico , Transcripción Genética/efectos de los fármacos , Regulación hacia Arriba/efectos de los fármacos
17.
Anticancer Res ; 40(10): 5529-5538, 2020 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-32988876

RESUMEN

BACKGROUND/AIM: Triple-negative breast cancer (TNBC) is a unique subtype that lacks expression of several conventional biomarkers and has a higher incidence of lymph node invasion and distal metastasis among all breast cancers. Anoikis resistance is the fundamental reason behind tumor cells' survival without their attachment to the extracellular matrix and metastasis to distal organs. Therefore, finding novel anti-cancer drugs that can suppress anoikis resistance in cancer cells is critical for patients with TNBC. MATERIALS AND METHODS: Curcumol, a natural compound, was used to assess whether it can inhibit the anoikis resistance and affects cell mortality and motility of IV2-1 TNBC cells. RESULTS: Curcumol suppressed anoikis resistance and inhibited TNBC cell survival in suspension. Additionally, these anti-cancer effects induced by curcumol could be related to the YAP1/Skp2 molecular pathway. CONCLUSION: Curcumol is an effective Skp2-targeted therapy that attenuates anoikis resistance and metastasis in TNBC cells.


Asunto(s)
MicroARNs/genética , Proteínas Quinasas Asociadas a Fase-S/genética , Sesquiterpenos/farmacología , Neoplasias de la Mama Triple Negativas/tratamiento farmacológico , Anoicis/efectos de los fármacos , Antineoplásicos/farmacología , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Resistencia a Antineoplásicos/efectos de los fármacos , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Metástasis de la Neoplasia , Transducción de Señal/efectos de los fármacos , Transcripción Genética/efectos de los fármacos , Neoplasias de la Mama Triple Negativas/genética , Neoplasias de la Mama Triple Negativas/patología
18.
Nat Commun ; 11(1): 4634, 2020 09 14.
Artículo en Inglés | MEDLINE | ID: mdl-32929078

RESUMEN

The current opioid epidemic necessitates a better understanding of human addiction neurobiology to develop efficacious treatment approaches. Here, we perform genome-wide assessment of chromatin accessibility of the human striatum in heroin users and matched controls. Our study reveals distinct neuronal and non-neuronal epigenetic signatures, and identifies a locus in the proximity of the gene encoding tyrosine kinase FYN as the most affected region in neurons. FYN expression, kinase activity and the phosphorylation of its target Tau are increased by heroin use in the post-mortem human striatum, as well as in rats trained to self-administer heroin and primary striatal neurons treated with chronic morphine in vitro. Pharmacological or genetic manipulation of FYN activity significantly attenuates heroin self-administration and responding for drug-paired cues in rodents. Our findings suggest that striatal FYN is an important driver of heroin-related neurodegenerative-like pathology and drug-taking behavior, making FYN a promising therapeutic target for heroin use disorder.


Asunto(s)
Cromatina/metabolismo , Cuerpo Estriado/enzimología , Dependencia de Heroína/enzimología , Terapia Molecular Dirigida , Proteínas Proto-Oncogénicas c-fyn/metabolismo , Animales , Secuencia de Bases , Conducta Animal/efectos de los fármacos , Señales (Psicología) , Genoma , Células HEK293 , Heroína/efectos adversos , Humanos , Masculino , Neuronas/metabolismo , Fosforilación/efectos de los fármacos , Regiones Promotoras Genéticas/genética , Proteínas Proto-Oncogénicas c-fyn/antagonistas & inhibidores , Ratas Long-Evans , Autoadministración , Transcripción Genética/efectos de los fármacos , Proteínas tau/metabolismo
19.
Chemosphere ; 254: 126909, 2020 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-32957299

RESUMEN

Soil contamination by heavy metals (HMs) is an environmental problem, and nanoremediation by using zero-valent iron nanoparticles (nZVI) has attracted increasing interest. We used ecotoxicological test and global transcriptome analysis with DNA microarrays to assess the suitability of C. elegans as a useful bioindicator to evaluate such strategy of nanoremediation in a highly polluted soil with Pb, Cd and Zn. The HMs produced devastating effect on C. elegans. nZVI treatment reversed this deleterious effect up to day 30 after application, but the reduction in the relative toxicity of HMs was lower at day 120. We stablished gene expression profile in C. elegans exposed to the polluted soil, treated and untreated with nZVI. The percentage of differentially expressed genes after treatment decreases with exposure time. After application of nZVI we found decreased toxicity, but increased biosynthesis of defensive enzymes responsive to oxidative stress. At day 14, when a decrease in toxicity has occurred, genes related to specific heavy metal detoxification mechanisms or to response to metal stress, were down regulated: gst-genes, encoding for glutathione-S-transferase, htm-1 (heavy metal tolerance factor), and pgp-5 and pgp-7, related to stress response to metals. At day 120, we found increased HMs toxicity compared to day 14, whereas the transcriptional oxidative and metal-induced responses were attenuated. These findings indicate that the profiled gene expression in C. elegans may be considered as an indicator of stress response that allows a reliable evaluation of the nanoremediation strategy.


Asunto(s)
Caenorhabditis elegans/efectos de los fármacos , Hierro/química , Metales Pesados/toxicidad , Nanopartículas/química , Estrés Oxidativo/efectos de los fármacos , Contaminantes del Suelo/toxicidad , Transcripción Genética/efectos de los fármacos , Animales , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/genética , Ecotoxicología , Nanopartículas del Metal , Metales Pesados/análisis , Estrés Oxidativo/genética , Suelo/química , Microbiología del Suelo , Contaminantes del Suelo/análisis , Toxicogenética
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