RESUMO
BACKGROUND: Targeting long non-coding RNAs (LncRNAs) is a novel and promising approach in cancer therapy. In our previous study, we investigated the effects of ailanthone (aila), the main active compound derived from the stem barks of Ailanthus altissima (Mill.) Swingle, on the growth of non-small cell lung cancer (NSCLC) cells. Although we observed significant inhibition of NSCLC cell growth of aila, the underlying mechanisms involving LncRNAs, specifically LncRNA growth arrest specific 5 (GAS5), remain largely unknown. METHODS: To further explore the impact of aila on NSCLC, we performed a series of experiments. Firstly, we confirmed the inhibitory effect of aila on NSCLC cell growth using multiple assays, including MTT, wound healing, transwell assay, as well as subcutaneous and metastasis tumor mice models in vivo. Next, we utilized cDNA microarray and RT-QPCR to identify GAS5 as the primary target of aila. To verify the importance of GAS5 in aila-induced tumor inhibition, we manipulated GAS5 expression levels by constructing GAS5 over-expression and knockdown NSCLC cell lines. Furthermore, we investigated the upstream and downstream signaling pathways of GAS5 through western blot and RT-QPCR analysis. RESULTS: Our results showed that aila effectively increased GAS5 expression, as determined by microarray analysis. We also observed that aila significantly enhanced GAS5 expression in a dose- and time-dependent manner across various NSCLC cell lines. Notably, over-expression of GAS5 led to a significant suppression of NSCLC cell tumor growth; while aila had minimal inhibitory effect on GAS5-knockdown NSCLC cells. Additionally, we discovered that aila inhibited ULK1 and autophagy, and this inhibition was reversed by GAS5 knockdown. Moreover, we found that aila up-regulated GAS5 expression by suppressing UPF1-mediated nonsense-mediated mRNA decay (NMD). CONCLUSION: In summary, our findings suggest that aila promotes GAS5 expression by inhibiting UPF1-mediated NMD, leading to the repression of ULK1-mediated autophagy and subsequent inhibitory effects on NSCLC cells. These results indicate that aila is a potent enhancer of GAS5 and holds promising potential for application in NSCLC therapy. However, our research is currently focused only on NSCLC. It remains to be determined whether aila can also inhibit the growth of other types of tumors through the UPF1/GAS5/ULK1 signaling pathway. In future studies, we can further investigate the mechanisms by which aila suppresses other types of tumors and potentially broaden the scope of its application in cancer therapy.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , RNA Longo não Codificante , Transdução de Sinais , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , RNA Longo não Codificante/genética , Humanos , Animais , Neoplasias Pulmonares/tratamento farmacológico , Transdução de Sinais/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Camundongos , Camundongos Nus , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/genética , Transativadores/genética , Transativadores/metabolismo , Ailanthus/química , Antineoplásicos Fitogênicos/farmacologia , Camundongos Endogâmicos BALB C , Quassinas/farmacologia , RNA Helicases/metabolismoRESUMO
Breast cancer is a serious threat to human health. The transforming growth factor-ß signaling pathway is an important pathway involved in the occurrence and development of cancer. The SMAD family genes are responsible for the TGF-ß signaling pathway. However, the mechanism by which genes of the SMAD family are involved in breast cancer is still unclear. Therefore, it is necessary to investigate the biological roles of the SMAD family genes in breast cancer. We downloaded the gene expression data, gene mutation data, and clinical pathological data of breast cancer patients from the UCSC Xena database. We used the Wilcox test to estimate the expression of genes of the SMAD family in cancers. And the biological functions of SMAD family genes using the DAVID website. The Pearson correlation method was used to explore the immune cell infiltration and drug response of SMAD family genes. We conducted in biological experiments vitro and vivo. In this study, we integrated the multi-omics data from TCGA breast cancer patients for analysis. The expression of genes of SMAD family was significantly dysregulated in patients with breast cancer. Except for SMAD6, the expression of other SMAD family genes was positively correlated. We also found that genes of the SMAD family were significantly enriched in the TGF-ß signaling pathway, Hippo signaling pathway, cell cycle, and cancer-related pathways. In addition, SMAD3, SMAD6, and SMAD7 were lowly expressed in stage II breast cancer, while SMAD4 and SMAD2 were lowly expressed in stage III cancer. Furthermore, the expression of genes of the SMAD family was significantly correlated with immune cell infiltration scores. Constructing a xenograft tumor mouse model, we found that SMAD3 knockdown significantly inhibited tumorigenesis. Finally, we analyzed the association between these genes and the IC50 value of drugs. Interestingly, patients with high expression of SMAD3 exhibited significant resistance to dasatinib and staurosporine, while high sensitivity to tamoxifen and auranofin. In addition, SMAD3 knockdown promoted the apoptosis of BT-549 cells and decreased cell activity, and BAY-1161909 and XK-469 increased drug efficacy. In conclusion, genes of the SMAD family play a crucial role in the development of breast cancer.
Assuntos
Neoplasias da Mama , Transativadores , Humanos , Animais , Camundongos , Feminino , Transativadores/metabolismo , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Transdução de Sinais , Proteína Smad4/genética , Proteína Smad4/metabolismo , Proteína Smad2/genética , Proteína Smad2/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Proteína Smad3/genética , Proteína Smad3/metabolismo , Proteínas Smad/genética , Proteínas Smad/metabolismoRESUMO
BACKGROUND: Recent studies have demonstrated that copper (Cu) plays an important role in the progression of tumor diseases. Metastasis associated with colon cancer protein 1 (MACC1) promotes the transcription and expression of various tumor-related genes. Cytochrome c oxidase (COX) 19, present in the cytoplasm and intermembrane space of mitochondria, may transport Cu within the mitochondria. However, the mechanism through which MACC1 regulates the Cu homeostasis mediated by COX19 remains unclear. OBJECTIVES: The aim of this study was to elucidate the mechanism through which MACC1 initiates the transcription and expression of COX19, and promotes malignant behavior in tumor cells. METHODS: Immunohistochemistry, western blotting, and real-time polymerase chain reaction (PCR) analyses were conducted to analyze the expression of MACC1 and COX19 proteins and genes in tumor and normal tissues. RNA-chromatin immunoprecipitation was used to detect the transcriptional initiation of COX19 by MACC1. The effects of MACC1 and COX19 on mitochondrial activity were determined using an ATP assay kit and Cytochrome c Oxidase Assay Kit. A Cell Counting Kit-8 kit was used to detect the effect of high-dose Cu or overexpression of MACC1 and COX19 on tumor cell proliferation. A xenograft mouse model was used to analyze the effect of the COX19 overexpression on the malignant behavior of the tumors. RESULTS: Cu enhanced the proliferation, invasion, and migration and inhibited apoptosis of SW480 cells. MACC1 was highly expressed in colorectal cancer tissues and activated the expression of COX19 by binding to its promoter region of COX19. The overexpression of COX19 increased mitochondrial Cu content and enhanced the activity of mitochondrial COX and ATP content, and inhibited apoptosis, promoted tumor growth of mice. CONCLUSIONS: Our results indicate that COX19 functions as a target gene of MACC1 and regulates mitochondrial activity and promotes the progression of colorectal cancer. MACC1/COX19 may provide a novel therapeutic target for colorectal cancer.
Assuntos
Neoplasias do Colo , Neoplasias Colorretais , Animais , Humanos , Camundongos , Trifosfato de Adenosina/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Neoplasias do Colo/metabolismo , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Cobre/metabolismo , Complexo IV da Cadeia de Transporte de Elétrons/genética , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Regulação Neoplásica da Expressão Gênica , Mitocôndrias/metabolismo , Transativadores/genética , Transativadores/metabolismoRESUMO
Retinoic acid-induced 1 (RAI1) haploinsufficiency causes Smith-Magenis syndrome (SMS), a genetic disorder with symptoms including hyperphagia, hyperlipidemia, severe obesity, and autism phenotypes. RAI1 is a transcriptional regulator with a pan-neural expression pattern and hundreds of downstream targets. The mechanisms linking neural Rai1 to body weight regulation remain unclear. Here we find that hypothalamic brain-derived neurotrophic factor (BDNF) and its downstream signalling are disrupted in SMS (Rai1+/-) mice. Selective Rai1 loss from all BDNF-producing cells or from BDNF-producing neurons in the paraventricular nucleus of the hypothalamus (PVH) induced obesity in mice. Electrophysiological recordings revealed that Rai1 ablation decreased the intrinsic excitability of PVHBDNF neurons. Chronic treatment of SMS mice with LM22A-4 engages neurotrophin downstream signalling and delayed obesity onset. This treatment also partially rescued disrupted lipid profiles, insulin intolerance, and stereotypical repetitive behaviour in SMS mice. These data argue that RAI1 regulates body weight and metabolic function through hypothalamic BDNF-producing neurons and that targeting neurotrophin downstream signalling might improve associated SMS phenotypes.
Assuntos
Fator Neurotrófico Derivado do Encéfalo , Síndrome de Smith-Magenis , Transativadores , Fatores de Transcrição , Animais , Camundongos , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Homeostase , Hipotálamo/metabolismo , Neurônios/metabolismo , Obesidade/genética , Síndrome de Smith-Magenis/genética , Síndrome de Smith-Magenis/metabolismo , Transativadores/metabolismo , Fatores de Transcrição/metabolismo , Fatores de Crescimento Neural/metabolismo , Peso CorporalRESUMO
The transcriptional co-activators Yes-associated protein (YAP) and PDZ-binding domain (TAZ) are the known downstream effectors of the Hippo kinase cascade. YAP/TAZ have been shown to play important roles in cellular growth and differentiation, tissue development and carcinogenesis. Recent studies have found that, in addition to the Hippo kinase cascade, multiple non-Hippo kinases also regulate the YAP/TAZ cellular signaling and produce important effects on cellular functions, particularly on tumorigenesis and progression. In this article, we will review the multifaceted regulation of the YAP/TAZ signaling by the non-Hippo kinases and discuss the potential application of the non-Hippo kinase-regulated YAP/TAZ signaling for cancer therapy.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal , Proteínas Serina-Treonina Quinases , Humanos , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Transativadores/metabolismo , Proteínas de Sinalização YAP , Proteínas com Motivo de Ligação a PDZ com Coativador Transcricional , Via de Sinalização Hippo , Fosfoproteínas/metabolismo , Transdução de Sinais , Fatores de Transcrição/metabolismo , CarcinogêneseRESUMO
Actin has important functions in both cytoplasm and nucleus of the cell, with active nuclear transport mechanisms maintaining the cellular actin balance. Nuclear actin levels are subject to regulation during many cellular processes from cell differentiation to cancer. Here we show that nuclear actin levels increase upon differentiation of PC6.3 cells towards neuron-like cells. Photobleaching experiments demonstrate that this increase is due to decreased nuclear export of actin during cell differentiation. Increased nuclear actin levels lead to decreased nuclear localization of MRTF-A, a well-established transcription cofactor of SRF. In line with MRTF-A localization, transcriptomics analysis reveals that MRTF/SRF target gene expression is first transiently activated, but then substantially downregulated during PC6.3 cell differentiation. This study therefore describes a novel cellular context, where regulation of nuclear actin is utilized to tune MRTF/SRF target gene expression during cell differentiation.
Assuntos
Actinas , Transativadores , Actinas/genética , Actinas/metabolismo , Diferenciação Celular/genética , Expressão Gênica , Regulação da Expressão Gênica , Extratos Vegetais , Fator de Resposta Sérica/genética , Fator de Resposta Sérica/metabolismo , Transativadores/genética , Transativadores/metabolismoRESUMO
We recently described a new member of the CRP (cyclic AMP receptor protein)/FNR (fumarate and nitrate reductase regulatory protein) family called RedB, an acronym for redox brake, that functions to limit the production of ATP and NADH. This study shows that the RedB regulon significantly overlaps the FnrL regulon, with 199 genes being either directly or indirectly regulated by both of these global regulatory proteins. Among these 199 coregulated genes, 192 are divergently regulated, indicating that RedB functions as an antagonist of FnrL. Chromatin immunoprecipitation sequencing (ChIP-seq) analysis indicates that RedB and Fnr directly coregulate only 4 out of 199 genes. The primary mechanism for the divergent regulation of target genes thus involves indirect regulation by both RedB and FnrL (156 cases). Additional regulation involves direct binding by RedB and indirect regulation by FnrL (36 cases) or direct binding by FnrL and indirect regulation by RedB (3 cases). Analysis of physiological pathways under direct and indirect control by these global regulators demonstrates that RedB functions primarily to limit energy production, while FnrL functions to enhance energy production. This regulation includes glycolysis, gluconeogenesis, photosynthesis, hydrogen oxidation, electron transport, carbon fixation, lipid biosynthesis, and protein synthesis. Finally, we show that 75% of genomes from diverse species that code for RedB proteins also harbor genes coding for FNR homologs. This cooccurrence indicates that RedB likely has an important role in buffering FNR-mediated energy production in a broad range of species. IMPORTANCE The CRP/FNR family of regulatory proteins constitutes a large collection of related transcription factors, several of which globally regulate cellular energy production. A well-characterized example is FNR (called FnrL in Rhodobacter capsulatus), which is responsible for regulating the expression of numerous genes that promote maximal energy production and growth under anaerobic conditions. In a companion article (N. Ke, J. E. Kumka, M. Fang, B. Weaver, et al., Microbiol Spectr 10:e02353-22, 2022, https://doi.org/10.1128/Spectrum02353-22), we identified a new subgroup of the CRP/FNR family and demonstrated that a member of this new subgroup, called RedB, has a role in limiting cellular energy production. In this study, we show that numerous genes encompassing the RedB regulon significantly overlap genes that are members of the FnrL regulon. Furthermore, 97% of the genes that are members of both the RedB and FnrL regulons are divergently regulated by these two transcription factors. RedB thus functions as a buffer limiting the amount of energy production that is promoted by FnrL.
Assuntos
Rhodobacter capsulatus , Rhodobacter sphaeroides , Trifosfato de Adenosina/metabolismo , Anaerobiose , Proteínas de Bactérias/metabolismo , Proteína Receptora de AMP Cíclico/metabolismo , Fumaratos/metabolismo , Regulação Bacteriana da Expressão Gênica , Hidrogênio/metabolismo , Lipídeos , NAD/genética , NAD/metabolismo , Oxirredução , Rhodobacter capsulatus/genética , Rhodobacter capsulatus/metabolismo , Rhodobacter sphaeroides/genética , Rhodobacter sphaeroides/metabolismo , Transativadores/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
Plant-associated bacteria, including pathogens, recognise host-derived signals to activate specific responses. The genome of Pseudomonas syringae pv. actinidiae (Psa), the aetiological agent of bacterial canker of kiwifruit, encodes for three putative LuxR-like receptors. Proteins of this family are usually involved in the quorum sensing system, through the perception of autoinducers (AHLs) produced by a cognate LuxI. However, Psa does not produce AHLs according to the lack of LuxI-encoding gene. It has been proposed that the so-called LuxR solos may be involved in the perception of environmental stimuli. We thus hypothesised that Psa LuxR-like receptors could be involved in host-derived signal sensing. Psa virulence traits, i.e., biofilm formation, motility and endophytic colonisation, were stimulated by growing the pathogen in host plant extracts, but not in non-host plant extracts or rich medium. Moreover, the phenotypic analyses of Psa mutant strains lacking the LuxR solo-encoding genes, demonstrated that PsaR2 plays a major role in host recognition and induction of virulence responses. The heterologous expression of PsaR2, followed by affinity chromatography and fraction activity assessment, confirmed the specific recognition of plant-derived components by this sensor. Overall, these data provide a deeper understanding of the regulation of Psa virulence through interkingdom communication, which represents a interesting target for the development of tolerant/resistant genotypes or innovative control strategies.
Assuntos
Pseudomonas syringae , Doenças das Plantas/microbiologia , Extratos Vegetais , Pseudomonas syringae/genética , Transativadores/genética , Transativadores/metabolismo , Virulência/genéticaRESUMO
Dickeya solani is a pathogen most frequently responsible for infecting potato plants in Europe. As in the case of most plant pathogens, its ability to colonize and invade the host depends on chemotaxis and motility. The coordinated movement of Dickeya over solid surfaces is governed by a quorum sensing mechanism. In D. solani motility is regulated by ExpI-ExpR proteins, homologous to luxI-luxR system from Vibrio fisheri, in which N-acyl-homoserine lactones (AHLs) serve as signaling molecules. Moreover, in many Gram-negative bacteria motility is coupled with central metabolism via carbon catabolite repression. This enables them to reach more nutrient-efficient niches. The aim of this study was to analyze the swarming motility of D. solani depending on the volume of the medium in the cultivation plate and glucose content. We show that the ability of this bacterium to move is strictly dependent on both these factors. Moreover, we analyze the production of AHLs and show that the quorum sensing mechanism in D. solani is also influenced by the availability of glucose in the medium and that the distribution of these signaling molecules are different depending on the volume of the medium in the plate.
Assuntos
Acil-Butirolactonas/farmacologia , Proteínas de Bactérias/genética , Dickeya/efeitos dos fármacos , Glucose/farmacologia , Solanum tuberosum/microbiologia , Fatores de Virulência/genética , Acil-Butirolactonas/metabolismo , Proteínas de Bactérias/metabolismo , Quimiotaxia/efeitos dos fármacos , Quimiotaxia/genética , Meios de Cultura/química , Meios de Cultura/farmacologia , Dickeya/genética , Dickeya/metabolismo , Dickeya/patogenicidade , Regulação Bacteriana da Expressão Gênica , Glucose/metabolismo , Doenças das Plantas/microbiologia , Percepção de Quorum/efeitos dos fármacos , Percepção de Quorum/genética , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Transativadores/genética , Transativadores/metabolismo , Fatores de Virulência/metabolismoRESUMO
Saponin gypenoside A (GP) has shown its potential to handle diabetes mellitus. MicroRNA-150-3p (miR-150-3p) is closely related to the dysfunction of pancreatic ß cells by targeting PDX1. Given the function of GP is related to its regulation on different miRs, the current study assessed the role of miR-150-3p as a therapeutic target for the hypoglycemic effects of GP. Pancreatic ß cell dysfunction was induced in mice using the high-fatty diet (HFD) method and then handled with GP. Changes in insulin release and resistance and the activity of the miR-150-3p/PDX1 axis were detected. The expression of miR-150-3p was induced to confirm its central in the effects of GP. The results of in vivo tests were then validated with in vitro assays. HFD administration suppressed glucose tolerance, delayed insulin release, and induced insulin resistance and pancreas apoptosis in mice, which was indicative of the dysfunction of ß pancreatic cells. Changes in pancreatic ß function were associated with the increased expression of miR-150-3p and suppressed expression of PDX1. After the administration of GP, the impairments of the pancreas were alleviated and the expression of miR-150-3p was inhibited, contributing to the restored level of PDX1. The injection of miR-150-3p agomir counteracted the protective effects of GP. In in vitro assays, the pretransfection of miR-150-3p mimetics also counteracted the protective effects of GP on pancreatic ß cells against palmitic acid. Collectively, miR-150-3p played a key role in the protective effects of GP against pancreatic ß cell dysfunction by inhibiting PDX1 expression.
Assuntos
Células Secretoras de Insulina , MicroRNAs , Animais , Gynostemma , Proteínas de Homeodomínio/metabolismo , Insulina/metabolismo , Células Secretoras de Insulina/metabolismo , Camundongos , MicroRNAs/genética , MicroRNAs/metabolismo , Extratos Vegetais , Transdução de Sinais , Transativadores/metabolismoRESUMO
Phyllanthus emblica is a traditional medicinal plant that is endowed with curative properties including anti-bacterial, anti-fungal, anti-viral, and analgesic properties. Bacteria make use of cell-cell signaling system known as quorum sensing (QS) and respond to their own population. In most gram-negative bacteria, the transcriptional regulators belonging to the Lux R protein play a crucial role in the QS mechanism by detecting the presence of signaling molecules known as N-acyl homoserine lactones (AHLs). In this present work, the anti-quorum sensing activity of Phyllanthus emblica was evaluated against Pseudomonas aeruginosa. Anti-quorum sensing efficacy of Phyllanthus emblica was estimated with reference to QS bio-monitoring strain Chromobacterium violaceum. The binding efficacy of the phytochemicals of Phyllanthus emblica against CviR protein from Chromobacterium violaceum and LasR protein from Phyllanthus emblica were studied.
Assuntos
Acil-Butirolactonas , Antibacterianos , Proteínas de Bactérias , Simulação de Acoplamento Molecular , Phyllanthus emblica/química , Compostos Fitoquímicos , Pseudomonas aeruginosa , Percepção de Quorum/efeitos dos fármacos , Transativadores , Acil-Butirolactonas/química , Acil-Butirolactonas/farmacologia , Antibacterianos/química , Antibacterianos/farmacologia , Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Compostos Fitoquímicos/química , Compostos Fitoquímicos/farmacologia , Pseudomonas aeruginosa/química , Pseudomonas aeruginosa/metabolismo , Transativadores/química , Transativadores/metabolismoRESUMO
Cancer is a global health problem. The delineation of molecular mechanisms pertinent to cancer initiation and development has spurred cancer therapy in the form of precision medicine. The Hippo signalling pathway is a tumour suppressor pathway implicated in a multitude of cancers. Elucidation of the Hippo pathway has revealed an increasing number of regulators that are implicated, some being potential therapeutic targets for cancer interventions. WW domain-binding protein 2 (WBP2) is an oncogenic transcriptional co-factor that interacts, amongst others, with two other transcriptional co-activators, YAP and TAZ, in the Hippo pathway. WBP2 was recently discovered to modulate the upstream Hippo signalling components by associating with LATS2 and WWC3. Exacerbating the complexity of the WBP2/Hippo network, WBP2 itself is reciprocally regulated by Hippo-mediated microRNA biogenesis, contributing to a positive feedback loop that further drives carcinogenesis. Here, we summarise the biological mechanisms of WBP2/Hippo reciprocal regulation and propose therapeutic strategies to overcome Hippo defects in cancers through targeting WBP2.
Assuntos
Via de Sinalização Hippo , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Transativadores/metabolismo , Variações do Número de Cópias de DNA/genética , Humanos , Modelos Biológicos , Medicina de PrecisãoRESUMO
The ubiquitous Ca2+ sensor calmodulin (CaM) binds and regulates many proteins, including ion channels, CaM kinases, and calcineurin, according to Ca2+-CaM levels. What regulates neuronal CaM levels, is, however, unclear. CaM-binding transcription activators (CAMTAs) are ancient proteins expressed broadly in nervous systems and whose loss confers pleiotropic behavioral defects in flies, mice, and humans. Using Caenorhabditis elegans and Drosophila, we show that CAMTAs control neuronal CaM levels. The behavioral and neuronal Ca2+ signaling defects in mutants lacking camt-1, the sole C. elegans CAMTA, can be rescued by supplementing neuronal CaM. CAMT-1 binds multiple sites in the CaM promoter and deleting these sites phenocopies camt-1. Our data suggest CAMTAs mediate a conserved and general mechanism that controls neuronal CaM levels, thereby regulating Ca2+ signaling, physiology, and behavior.
Assuntos
Proteínas de Ligação a Calmodulina/metabolismo , Calmodulina/metabolismo , Proteínas de Drosophila/metabolismo , Neurônios/metabolismo , Transativadores/metabolismo , Fatores de Transcrição/metabolismo , Animais , Caenorhabditis elegans/metabolismo , Calcineurina/metabolismo , Cálcio/metabolismo , Drosophila/metabolismo , Proteínas de Drosophila/genética , Drosophila melanogaster , Feminino , Edição de Genes , Regulação da Expressão Gênica , Humanos , Masculino , Camundongos , Ligação Proteica , Transdução de Sinais , Transativadores/genética , TranscriptomaRESUMO
BACKGROUND: Bu Shen Yi Sui capsule (BSYS) is a traditional Chinese medicine prescription that has shown antineuroinflammatory and neuroprotective effects in treating multiple sclerosis (MS) and its animal model of experimental autoimmune encephalomyelitis (EAE). Microglia play an important role in neuroinflammation. The M1 phenotype of microglia is involved in the proinflammatory process of the disease, while the M2 phenotype plays an anti-inflammatory role. Promoting the polarization of microglia to M2 in MS/EAE is a promising therapeutic strategy. This study is aimed at exploring the effects of BSYS on microglial polarization in mice with EAE. METHODS: The EAE model was established by the intraperitoneal injection of pertussis toxin and subcutaneous injection of myelin oligodendrocyte glycoprotein (MOG)35-55 in C57BL/6J mice. The mice were treated with BSYS (3.02 g/kg), FTY720 (0.3 mg/kg), or distilled water by intragastric administration. H&E and LFB staining, transmission electron microscopy, qRT-PCR, immunofluorescence, ELISA, fluorescence in situ hybridization, and western blotting were used to detect the histological changes in myelin, microglial M1/M2 polarization markers, and the expression of key genes involved in EAE. Results and Conclusions. BSYS treatment of EAE mice increased the body weight, decreased the clinical score, and reduced demyelination induced by inflammatory infiltration. BSYS also inhibited the mRNA expression of M1 microglial markers while increasing the mRNA level of M2 markers. Additionally, BSYS led to a marked decrease in the ratio of M1 microglia (iNOS+/Iba1+) and an obvious increase in the number of M2 microglia (Arg1+/Iba1+). In the EAE mouse model, miR-124 expression was decreased, and miR-155 expression was increased, while BSYS treatment significantly reversed this effect and modulated the levels of C/EBP α, PU.1, and SOCS1 (target genes of miR-124 and miR-155). Therefore, the neuroprotective effect of BSYS against MS/EAE was related to promoting microglia toward M2 polarization, which may be correlated with changes in miR-124 and miR-155 in vivo.
Assuntos
Encéfalo/patologia , Doenças Desmielinizantes/genética , Medicamentos de Ervas Chinesas/farmacologia , Encefalomielite Autoimune Experimental/genética , Inflamação/patologia , MicroRNAs/metabolismo , Microglia/patologia , Animais , Peso Corporal/efeitos dos fármacos , Proteínas Estimuladoras de Ligação a CCAAT/metabolismo , Cápsulas , Diferenciação Celular/efeitos dos fármacos , Polaridade Celular/efeitos dos fármacos , Citocinas/metabolismo , Encefalomielite Autoimune Experimental/sangue , Encefalomielite Autoimune Experimental/patologia , Exossomos/metabolismo , Feminino , Inflamação/genética , Camundongos Endogâmicos C57BL , MicroRNAs/sangue , MicroRNAs/genética , Oligodendroglia/efeitos dos fármacos , Oligodendroglia/patologia , Fenótipo , Proteínas Proto-Oncogênicas/metabolismo , Medula Espinal/patologia , Transativadores/metabolismo , Regulação para Cima/genéticaRESUMO
Atopic dermatitis (AD) is a severe inflammatory skin disease. Langerhans cells and inflammatory dendritic epidermal cells (IDEC) are located in the epidermis of AD patients and contribute to the inflammatory processes. Both express robustly the high-affinity receptor for IgE, FcεRI, and thereby sense allergens. A beneficial role of vitamin D3 in AD is discussed to be important especially in patients with allergic sensitization. We hypothesized that vitamin D3 impacts FcεRI expression and addressed this in human ex vivo skin, in vitro Langerhans cells, and IDEC models generated from primary human precursor cells. We show in this article that biologically active vitamin D3 [1,25(OH)2-D3] significantly downregulated FcεRI at the protein and mRNA levels of the receptor's α-chain, analyzed by flow cytometry and quantitative RT-PCR. We also describe the expression of a functional vitamin D receptor in IDEC. 1,25(OH)2-D3-mediated FcεRI reduction was direct and resulted in impaired activation of IDEC upon FcεRI engagement as monitored by CD83 expression. FcεRI regulation by 1,25(OH)2-D3 was independent of maturation and expression levels of microRNA-155 and PU.1 (as upstream regulatory axis of FcεRI) and transcription factors Elf-1 and YY1. However, 1,25(OH)2-D3 induced dissociation of PU.1 and YY1 from the FCER1A promotor, evaluated by chromatin immunoprecipitation. We show that vitamin D3 directly reduces FcεRI expression on dendritic cells by inhibiting transcription factor binding to its promotor and subsequently impairs IgE-mediated signaling. Thus, vitamin D3 as an individualized therapeutic supplement for those AD patients with allergic sensitization interferes with IgE-mediated inflammatory processes in AD patients.
Assuntos
Colecalciferol/metabolismo , Células Dendríticas/imunologia , Dermatite Atópica/imunologia , Proteínas Proto-Oncogênicas/metabolismo , Receptores de IgE/metabolismo , Transativadores/metabolismo , Fator de Transcrição YY1/metabolismo , Adulto , Idoso , Células Cultivadas , Regulação para Baixo , Feminino , Humanos , Imunoglobulina E/metabolismo , Masculino , Pessoa de Meia-Idade , Regiões Promotoras Genéticas , Ligação Proteica , Proteínas Proto-Oncogênicas/genética , Receptores de IgE/genética , Transdução de Sinais , Transativadores/genética , Fator de Transcrição YY1/genética , Adulto JovemRESUMO
Yin Yang 1 (YY1) regulates gene transcription in a variety of biological processes. In this study, we aim to determine the role of YY1 in vascular smooth muscle cell (VSMC) phenotypic modulation both in vivo and in vitro. Here we show that vascular injury in rodent carotid arteries induces YY1 expression along with reduced expression of smooth muscle differentiation markers in the carotids. Consistent with this finding, YY1 expression is induced in differentiated VSMCs in response to serum stimulation. To determine the underlying molecular mechanisms, we found that YY1 suppresses the transcription of CArG box-dependent SMC-specific genes including SM22α, SMα-actin and SMMHC. Interestingly, YY1 suppresses the transcriptional activity of the SM22α promoter by hindering the binding of serum response factor (SRF) to the proximal CArG box. YY1 also suppresses the transcription and the transactivation of myocardin (MYOCD), a master regulator for SMC-specific gene transcription by binding to SRF to form the MYOCD/SRF/CArG box triad (known as the ternary complex). Mechanistically, YY1 directly interacts with MYOCD to competitively displace MYOCD from SRF. This is the first evidence showing that YY1 inhibits SMC differentiation by directly targeting MYOCD. These findings provide new mechanistic insights into the regulatory mechanisms that govern SMC phenotypic modulation in the pathogenesis of vascular diseases.
Assuntos
Ribonucleoproteínas Nucleares Heterogêneas Grupo A-B/metabolismo , Miócitos de Músculo Liso/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Repressoras/metabolismo , Fator de Resposta Sérica/metabolismo , Transativadores/metabolismo , Fatores de Transcrição/metabolismo , Transcrição Gênica , Fator de Transcrição YY1/metabolismo , Animais , Masculino , Camundongos , Ratos , Ratos Sprague-DawleyRESUMO
HBV cccDNA stably exists in the nuclei of infected cells as an episomal munichromosome which is responsible for viral persistence and failure of current antiviral treatments. However, the regulatory mechanism of cccDNA transcription by viral and host cellular factors is not well understood. In this study, we investigated whether cccDNA could be recruited into a specific region of the nucleus via specific interaction with a cellular chromatin to regulate its transcription activity. To investigate this hypothesis, we used chromosome conformation capture (3C) technology to search for the potential interaction of cccDNA and cellular chromatin through rcccDNA transfection in hepatoma cells and found that cccDNA is specifically associated with human chromosome 19p13.11 region, which contains a highly active enhancer element. We also confirmed that cellular transcription factor Yin-Yang 1 (YY1) and viral protein HBx mediated the spatial regulation of HBV cccDNA transcription by 19p13.11 enhancer. Thus, These findings indicate that YY1 and HBx mediate the recruitment of HBV cccDNA minichromosomes to 19p13.11 region for transcription activation, and YY1 may present as a novel therapeutic target against HBV infection.
Assuntos
Carcinoma Hepatocelular/virologia , Cromossomos Humanos Par 19/metabolismo , DNA Viral/metabolismo , Vírus da Hepatite B/genética , Neoplasias Hepáticas/virologia , Transativadores/metabolismo , Proteínas Virais Reguladoras e Acessórias/metabolismo , Fator de Transcrição YY1/metabolismo , Carcinoma Hepatocelular/metabolismo , Linhagem Celular , Núcleo Celular/metabolismo , Núcleo Celular/virologia , Elementos Facilitadores Genéticos , Genoma Viral , Células Hep G2 , Vírus da Hepatite B/metabolismo , Humanos , Neoplasias Hepáticas/metabolismo , Transcrição Gênica , Replicação ViralRESUMO
The high mortality rate of colorectal cancer (CRC) patients is directly associated with metastatic dissemination. However, therapeutic options specifically for metastasis are still limited. We previously identified Metastasis-Associated in Colon Cancer 1 (MACC1) as a major causal metastasis-inducing gene. Numerous studies confirmed its value as a biomarker for metastasis risk. We investigated the inhibitory impact of saffron on MACC1-induced cancer cell growth and motility. Saffron crudes restricted the proliferation and migration of MACC1-expressing CRC cells in a concentration- and MACC1-dependent manner. Saffron delays cell cycle progression at G2/M-phase and does not induce apoptosis. Rescue experiments showed that these effects are reversible. Analysis of active saffron compounds elucidated that crocin was the main compound that reproduced total saffron crudes effects. We showed the interaction of MACC1 with the cancer stem cell (CSC) marker DCLK1, which contributes to metastasis formation in different tumor entities. Saffron extracts reduced DCLK1 with crocin being responsible for this reduction. Saffron's anti-proliferative and anti-migratory effects in MACC1-expressing cells are mediated by crocin through DCLK1 down-regulation. This research is the first identification of saffron-based compounds restricting cancer cell proliferation and motility progression via the novel target MACC1.
Assuntos
Antineoplásicos/farmacologia , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Neoplasias Colorretais/patologia , Crocus/química , Transativadores/metabolismo , Antineoplásicos/química , Carotenoides/análise , Carotenoides/farmacologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Quinases Semelhantes a Duplacortina , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Extratos Vegetais/química , Extratos Vegetais/farmacologia , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Transativadores/genéticaRESUMO
l-Carnitine is essential in the intermediary metabolism of eukaryotes and is involved in the ß-oxidation of medium- and long-chain fatty acids; thus, it has applications for medicinal purposes and as a dietary supplement. In addition, l-carnitine plays roles in bacterial physiology and metabolism, which have been exploited by the industry to develop biotechnological carnitine production processes. Here, on the basis of studies of l-carnitine metabolism in Escherichia coli and its activation by the transcriptional activator CaiF, a biosensor was developed. It expresses a fluorescent reporter gene that responds in a dose-dependent manner to crotonobetainyl-CoA, which is an intermediate of l-carnitine metabolism in E. coli and is proposed to be a coactivator of CaiF. Moreover, a dual-input biosensor for l-carnitine and crotonobetaine was developed. As an application of the biosensor, potential homologues of the betaine:CoA ligase CaiC from Citrobacter freundii, Proteus mirabilis, and Arcobacter marinus were screened and shown to be functionally active CaiC variants. These variants and the developed biosensor may be valuable for improving l-carnitine production processes.