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Neurofeedback, a non-invasive intervention, has been increasingly used as a potential treatment for major depressive disorders. However, the effectiveness of neurofeedback in alleviating depressive symptoms remains uncertain. To address this gap, we conducted a comprehensive meta-analysis to evaluate the efficacy of neurofeedback as a treatment for major depressive disorders. We conducted a comprehensive meta-analysis of 22 studies investigating the effects of neurofeedback interventions on depression symptoms, neurophysiological outcomes, and neuropsychological function. Our analysis included the calculation of Hedges' g effect sizes and explored various moderators like intervention settings, study designs, and demographics. Our findings revealed that neurofeedback intervention had a significant impact on depression symptoms (Hedges' g = -0.600) and neurophysiological outcomes (Hedges' g = -0.726). We also observed a moderate effect size for neurofeedback intervention on neuropsychological function (Hedges' g = -0.418). As expected, we observed that longer intervention length was associated with better outcomes for depressive symptoms (ß = -4.36, P < 0.001) and neuropsychological function (ß = -2.89, P = 0.003). Surprisingly, we found that shorter neurofeedback sessions were associated with improvements in neurophysiological outcomes (ß = 3.34, P < 0.001). Our meta-analysis provides compelling evidence that neurofeedback holds promising potential as a non-pharmacological intervention option for effectively improving depressive symptoms, neurophysiological outcomes, and neuropsychological function in individuals with major depressive disorders.
Assuntos
Transtorno Depressivo Maior , Neurorretroalimentação , Neurorretroalimentação/métodos , Humanos , Transtorno Depressivo Maior/terapia , Transtorno Depressivo Maior/fisiopatologia , Resultado do Tratamento , Eletroencefalografia/métodosRESUMO
The emergence of machine learning (ML) techniques has opened up new avenues for identifying biomarkers associated with schizophrenia (SCZ) using task-related fMRI (t-fMRI) designs. To evaluate the effectiveness of this approach, we conducted a comprehensive meta-analysis of 31 t-fMRI studies using a bivariate model. Our findings revealed a high overall sensitivity of 0.83 and specificity of 0.82 for t-fMRI studies. Notably, neuropsychological domains modulated the classification performance, with selective attention demonstrating a significantly higher specificity than working memory (ß = 0.98, z = 2.11, P = 0.04). Studies involving older, chronic patients with SCZ reported higher sensitivity (P <0.015) and specificity (P <0.001) than those involving younger, first-episode patients or high-risk individuals for psychosis. Additionally, we found that the severity of negative symptoms was positively associated with the specificity of the classification model (ß = 7.19, z = 2.20, P = 0.03). Taken together, these results support the potential of using task-based fMRI data in combination with machine learning techniques to identify biomarkers related to symptom outcomes in SCZ, providing a promising avenue for improving diagnostic accuracy and treatment efficacy. Future attempts to deploy ML classification should consider the factors of algorithm choice, data quality and quantity, as well as issues related to generalization.
Assuntos
Esquizofrenia , Humanos , Esquizofrenia/diagnóstico por imagem , Neuroimagem/métodos , Imageamento por Ressonância Magnética/métodos , Aprendizado de Máquina , BiomarcadoresRESUMO
T helper 17 (Th17) cells are a subset of CD4+ T helper cells involved in the inflammatory response in autoimmunity. Th17 cells secrete Th17 specific cytokines, such as IL-17A and IL17-F, which are governed by the master transcription factor RoRγt. However, the epigenetic mechanism regulating Th17 cell function is still not fully understood. Here, we reveal that deletion of RNA 5-methylcytosine (m5C) methyltransferase Nsun2 in mouse CD4+ T cells specifically inhibits Th17 cell differentiation and alleviates Th17 cell-induced colitis pathogenesis. Mechanistically, RoRγt can recruit Nsun2 to chromatin regions of their targets, including Il17a and Il17f, leading to the transcription-coupled m5C formation and consequently enhanced mRNA stability. Our study demonstrates a m5C mediated cell intrinsic function in Th17 cells and suggests Nsun2 as a potential therapeutic target for autoimmune disease.
Assuntos
Colite , Células Th17 , Animais , Camundongos , Diferenciação Celular/genética , Colite/genética , Regulação da Expressão Gênica , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/metabolismo , Fatores de Transcrição/genéticaRESUMO
We report a comprehensive proteomic study of a 90-case cohort of paired samples of triple-negative breast cancer (TNBC) in quantification, phosphorylation, and DNA-binding capacity. Four integrative subtypes (iP-1-4) are stratified on the basis of global proteome and phosphoproteome, each of which exhibits distinct molecular and pathway features. Scaffold and co-expression network analyses of three proteomic datasets, integrated with those from genome and transcriptome of the same cohort, reveal key pathways and master regulators that, characteristic of TNBC subtypes, play important regulatory roles within and between scaffold sub-structures and co-expression communities. We find that NAE1 is a potential drug target for subtype iP-1, and a series of key molecules in fatty acid metabolism, such as AKT1/FASN, are plausible targets for subtype iP-2. Libraries of proteins, pathways and networks of TNBC provide a valuable molecular infrastructure for further clinical exploration and in-depth studies of the molecular mechanisms of the disease.
Assuntos
Neoplasias de Mama Triplo Negativas , Genoma , Humanos , Proteoma/genética , Proteômica , Transcriptoma , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/metabolismoRESUMO
Great advances have been made in mass spectrometric data interpretation for intact glycopeptide analysis. However, accurate identification of intact glycopeptides and modified saccharide units at the site-specific level and with fast speed remains challenging. Here, we present a glycan-first glycopeptide search engine, pGlyco3, to comprehensively analyze intact N- and O-glycopeptides, including glycopeptides with modified saccharide units. A glycan ion-indexing algorithm developed for glycan-first search makes pGlyco3 5-40 times faster than other glycoproteomic search engines without decreasing accuracy or sensitivity. By combining electron-based dissociation spectra, pGlyco3 integrates a dynamic programming-based algorithm termed pGlycoSite for site-specific glycan localization. Our evaluation shows that the site-specific glycan localization probabilities estimated by pGlycoSite are suitable to localize site-specific glycans. With pGlyco3, we confidently identified N-glycopeptides and O-mannose glycopeptides that were extensively modified by ammonia adducts in yeast samples. The freely available pGlyco3 is an accurate and flexible tool that can be used to identify glycopeptides and modified saccharide units.
Assuntos
Biologia Computacional/métodos , Glicopeptídeos/química , Proteoma , Proteômica/métodos , Algoritmos , Animais , Vaga-Lumes , Glicosilação , Células HEK293 , Humanos , Manose/química , Polissacarídeos/química , Probabilidade , Reprodutibilidade dos Testes , Saccharomyces cerevisiae , Schizosaccharomyces , SoftwareRESUMO
The ARID1A gene, which encodes a subunit of the SWI/SNF chromatin remodeling complex, has been found to be frequently mutated in many human cancer types. However, the function and mechanism of ARID1A in cancer metastasis are still unclear. Here, we show that knockdown of ARID1A increases the ability of breast cancer cells to proliferate, migrate, invade, and metastasize in vivo. The ARID1A-related SWI/SNF complex binds to the second exon of CDH1 and negatively modulates the expression of E-cadherin/CDH1 by recruiting the transcriptional repressor ZEB2 to the CDH1 promoter and excluding the presence of RNA polymerase II. The silencing of CDH1 attenuated the migration, invasion, and metastasis of breast cancer cells in which ARID1A was silenced. ARID1A depletion increased the intracellular enzymatic processing of E-cadherin and the production of C-terminal fragment 2 (CTF2) of E-cadherin, which stabilized ß-catenin by competing for binding to the phosphorylation and degradation complex of ß-catenin. The matrix metalloproteinase inhibitor GM6001 inhibited the production of CTF2. In zebrafish and nude mice, ARID1A silencing or CTF2 overexpression activated ß-catenin signaling and promoted migration/invasion and metastasis of cancer cells in vivo. The inhibitors GM6001, BB94, and ICG-001 suppressed the migration and invasion of cancer cells with ARID1A-deficiency. Our findings provide novel insights into the mechanism of ARID1A metastasis and offer a scientific basis for targeted therapy of ARID1A-deficient cancer cells.
Assuntos
Antígenos CD , Caderinas , Animais , Humanos , CamundongosRESUMO
Mammalian cell cycle is a central process for tissue growth and maintenance. Protein O-linked ß-N-acetylglucosamine (O-GlcNAc) modification has been found to occur on several important cell cycle regulators. However, the O-GlcNAcylated proteome has not been extensively profiled during cell cycle progression. Herein, we report a quantitative profiling of protein O-GlcNAcylation sites in cell proliferation, by using an O-GlcNAc chemoproteomic strategy. In HeLa cells, a total of 902, 439, and 872 high-confidence O-GlcNAcylation sites distributed on 414, 265, and 425 proteins are identified in the interphase, early mitosis, and mitotic exit stages, respectively. The identified O-GlcNAcylation events occur on a variety of important regulators, which are involved in the processes of cell division, DNA repair, and cell death. Furthermore, we show that O-GlcNAcylation is dynamically regulated in a cell cycle stage-dependent manner. Our results provide a valuable resource for investigating the functional roles of O-GlcNAc in the mammalian cell cycle.
Assuntos
Acetilglucosamina/análise , Ciclo Celular/fisiologia , Glicoproteínas/análise , Glicoproteínas/metabolismo , Proteoma/análise , Proteoma/metabolismo , Anáfase/fisiologia , Glicoproteínas/química , Glicosilação , Células HeLa , Humanos , Interfase/fisiologia , Processamento de Proteína Pós-Traducional , Proteoma/química , ProteômicaRESUMO
PURPOSE: Colorectal cancer (CRC) is the third most common cancer in males and the second in females worldwide with very poor prognosis. Extracellular matrix proteins like collagens play important roles in cancer progression. Collagen type V α2 (COL5A2) is increased in several cancers but its role in cancer remains unclear. METHODS: COL5A2 expression was evaluated by interrogation of public Oncomine gene microarray datasets and immunohistochemistry (IHC) analyses of two tissue microarrays containing 180 paired CRC cases. Survival analysis was performed using Kaplan-Meier survival curve and Cox proportional hazards regression methods. COL5A2 was ectopically expressed in CRC cells, and the cell proliferation was measured using the methylthiazolyldiphenyl-tetrazolium bromide (MTT) method. RESULTS: COL5A2 gene was significantly upregulated in the most types of CRC comparing with the normal counterparts. The mRNA expression of COL5A2 was associated with cancer stages, gender, recurrence, microsatellite instability and KRAS status of CRC. COL5A2 protein increased in the cancer epithelial cells comparing with the normal counterpart and associated with age and T stage of CRC, whereas stromal expression of COL5A2 has no significant change between cancerous and normal tissues. COL5A2 gene and protein (epithelial expression) are independent risk factors and predict poor prognosis of CRC. Ectopic expression of COL5A2 drives colon cancer cell growth and upregulates WNT/ß-catenin and PI3K/mTOR signaling via binding DDR1. CONCLUSION: COL5A2 is a potential prognostic marker of CRC.
RESUMO
Introduction: Glycomics, which aims to define the glycome of a biological system to better assess the biological attributes of the glycans, has attracted increasing interest. However, the complexity and diversity of glycans present challenging barriers to glycome definition. Technological advances are major drivers in glycomics.Areas covered: This review summarizes the main methods and emphasizes the most recent advances in mass spectrometry-based methods regarding glycomics following the general workflow in glycomic analysis.Expert opinion: Recent mass spectrometry-based technological advances have significantly lowered the barriers in glycomics. The field of glycomics is moving toward both generic and precise analysis.
Assuntos
Glicômica/métodos , Espectrometria de Massas/métodos , Animais , Humanos , Polissacarídeos/químicaRESUMO
Efficient detection of aberrant glycoproteins in serum is particularly important for biomarker discovery. However, direct quantitation of glycoproteins in serum remains technically challenging because of the extraordinary complexity of the serum proteome. In the current work, we proposed a straightforward and highly efficient strategy by using the nonglycopeptides releasing from the specifically enriched glycoproteins for targeted glycoprotein quantification. With this so-called nonglycopeptide-based mass spectrometry (NGP-MS) strategy, a powerful and nondiscriminatory pipeline for hepatocellular carcinoma (HCC) glycoprotein biomarker discovery, verification, and validation has been developed. First, a data set of 234 NGPs was strictly established for multiple-reaction monitoring (MRM) quantification in serum. Second, the NGPs enriched from 20 HCC serum mixtures and 20 normal serum mixtures were labeled with mTRAQ reagents (Δ0 and Δ8, respectively) to find the differentially expressed glycoproteins in HCC. A total of 97 glycoprotein candidates were preliminarily screened and submitted for absolute quantitation with NGP-based stable-isotope-labeled (SID)-MRM in the individual samples of 38 HCC serum and 24 normal controls. Finally, 21 glycoproteins were absolutely quantified with high quality. The diagnostic sensitivity results showed that three glycoproteins, ß-2-glycoprotein 1 (APOH), α-1-acid glycoprotein 2 (ORM2), and complement C3 (C3), could be used for the discrimination between HCC patients and healthy people. A novel glycoprotein biomarker panel [APOH, ORM2, C3, and α-fetoprotein (AFP)] has proven to outperform AFP, the known HCC serum biomarker, alone, in this study. We believe that this strategy and the panel of glycoproteins might hold great clinical value for HCC detection in the future.
Assuntos
Carcinoma Hepatocelular/sangue , Glicoproteínas/sangue , Neoplasias Hepáticas/sangue , Espectrometria de Massas/métodos , Biomarcadores/sangue , Humanos , alfa-Fetoproteínas/metabolismoRESUMO
Protein phosphorylation is a reversible and important post-translational modification. Identification of phosphopeptides without enrichment is difficult for the low-abundance of phosphopeptides in real complex biological samples. Therefore, the effective and selective concentration of phosphopeptides prior to proteomic identification by mass spectrometer is necessary. In this study, we synthesized a novel titanium-based immobilized metal ion affinity chromatography material for highly selective enrichment of phosphopeptides. To improve material hydrophilia to the maximum extent, titanium ions were immobilized on the 4-armed Poly(ethylene oxide)(4µ-PEO-Ti4+), a totally soluble polymer with large molecular weight (20000â¯g/mol). The 4µ-PEO-Ti4+ was used to enrich phosphopeptides from tryptic digests of standard proteins and real complex biological samples, followed by MALDI-TOF MS analysis. In enrichment of phosphopeptides from 4â¯pmol ß-casein, the 4µ-PEO-Ti4+ performed the best property with starting material of 99-132⯵g, loading buffer of 50% ACN/5% TFA (v/v), elution buffer of 10% NH3·H2O (v/v) and elution time of 30â¯min. The 4µ-PEO-Ti4+ has a superior detection sensitivity as low as 2â¯fmol for phosphopeptides. The high selectivity of 4µ-PEO-Ti4+ allows a deep enrichment of phosphopeptides of ß-casein from a mixture with BSA of 1000-fold abundant. The 4µ-PEO-Ti4+ shows great stability and endurability and can be recycled up to at least 5 times. In addition, 4µ-PEO-Ti4+ could detect 10 and 15 phosphopeptides from non-fat milk and nonenzymatic human saliva, respectively. In total, 4µ-PEO-Ti4+ is a novel excellent material which shows great sensitive and selective enrichment of low-abundance phosphopeptides in real complex biological samples.
Assuntos
Fosfopeptídeos/isolamento & purificação , Polietilenoglicóis/química , Titânio/química , Sequência de Aminoácidos , Animais , Caseínas/química , Caseínas/isolamento & purificação , Cromatografia de Afinidade/métodos , Feminino , Humanos , Leite/química , Fragmentos de Peptídeos/isolamento & purificação , Proteólise , Saliva/química , Proteínas e Peptídeos Salivares/isolamento & purificação , Tripsina/químicaRESUMO
Benefiting from high sensitivity and great ability to measure multiple samples simultaneously, isobaric tandem Mass spectrometry (MS2) quantification has been widely applied for protein biomarker screening. Here, a newly developed isobaric MS2 quantification method named triplex quantification by isobaric termini labeling (Triplex-QITL) was established. This method enables the accurate comparison of various fragment ions (reporter ions, amino acid fragments and N-/C-terminal fragments) based quantification to be operated in a single run. To our knowledge, this is the first time that this kind of comparison is achieved. In Triplex-QITL, proteins were first digested with Lys-C to produce peptides with lysine (K) at the C-termini, then dimethylation reagents and mTRAQ reagents were used to label the N-termini and C-termini of the peptides respectively. N- and C-terminal fragment ion pairs, reporter ions from mTRAQ (113,117,121) and a1 ion pairs were simultaneously generated in MS2 spectra. In simple sample experiment, not much difference in using various fragment ions for quantification was observed. When analyzing SW480 cell lysate, comparing with a1 ions, about two times of reproducible quantification results were achieved by reporter ions and N- and C-terminal ions. Meanwhile the measured quantification results were much closer to the expected results even in large ratios (1:10:10) using N- and C-terminal ions. Finally, Triplex-QITL was successfully applied to profile metastatic differences of three hepatocellular carcinoma (HCC) cell lines. In all, Triplex-QITL shows a promising future in quantitative proteomics.
Assuntos
Proteômica/métodos , Espectrometria de Massas em Tandem/métodos , Sequência de Aminoácidos , Linhagem Celular Tumoral , Cromatografia Líquida de Alta Pressão/métodos , Humanos , Marcação por Isótopo/métodos , Neoplasias/química , Neoplasias/patologia , Peptídeos/análise , Proteínas/análise , Proteoma/análise , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodosRESUMO
Amino acids are known regulators of cellular signaling and physiology, but how they are sensed intracellularly is not fully understood. Herein, we report that each aminoacyl-tRNA synthetase (ARS) senses its cognate amino acid sufficiency through catalyzing the formation of lysine aminoacylation (K-AA) on its specific substrate proteins. At physiologic levels, amino acids promote ARSs bound to their substrates and form K-AAs on the É-amine of lysines in their substrates by producing reactive aminoacyl adenylates. The K-AA marks can be removed by deacetylases, such as SIRT1 and SIRT3, employing the same mechanism as that involved in deacetylation. These dynamically regulated K-AAs transduce signals of their respective amino acids. Reversible leucylation on ras-related GTP-binding protein A/B regulates activity of the mammalian target of rapamycin complex 1. Glutaminylation on apoptosis signal-regulating kinase 1 suppresses apoptosis. We discovered non-canonical functions of ARSs and revealed systematic and functional amino acid sensing and signal transduction networks.
Assuntos
Aminoacilação , Espaço Intracelular/metabolismo , Lisina/metabolismo , Transdução de Sinais , Aminoacil-tRNA Sintetases/metabolismo , Apoptose , Biocatálise , Células HEK293 , Humanos , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Especificidade por SubstratoRESUMO
The precise and large-scale identification of intact glycopeptides is a critical step in glycoproteomics. Owing to the complexity of glycosylation, the current overall throughput, data quality and accessibility of intact glycopeptide identification lack behind those in routine proteomic analyses. Here, we propose a workflow for the precise high-throughput identification of intact N-glycopeptides at the proteome scale using stepped-energy fragmentation and a dedicated search engine. pGlyco 2.0 conducts comprehensive quality control including false discovery rate evaluation at all three levels of matches to glycans, peptides and glycopeptides, improving the current level of accuracy of intact glycopeptide identification. The N-glycoproteome of samples metabolically labeled with 15N/13C were analyzed quantitatively and utilized to validate the glycopeptide identification, which could be used as a novel benchmark pipeline to compare different search engines. Finally, we report a large-scale glycoproteome dataset consisting of 10,009 distinct site-specific N-glycans on 1988 glycosylation sites from 955 glycoproteins in five mouse tissues.Protein glycosylation is a heterogeneous post-translational modification that generates greater proteomic diversity that is difficult to analyze. Here the authors describe pGlyco 2.0, a workflow for the precise one step identification of intact N-glycopeptides at the proteome scale.
Assuntos
Glicopeptídeos/análise , Proteômica/métodos , Ferramenta de Busca , Espectrometria de Massas em Tandem/métodos , Animais , Isótopos de Carbono , Glicopeptídeos/metabolismo , Glicosilação , Ensaios de Triagem em Larga Escala/métodos , Humanos , Masculino , Camundongos Endogâmicos C57BL , Isótopos de Nitrogênio , Polissacarídeos/análise , Polissacarídeos/metabolismo , Processamento de Proteína Pós-Traducional , Controle de Qualidade , Software , Fluxo de TrabalhoRESUMO
The von Hippel-Lindau (VHL) is deficient in â¼70% of clear-cell renal cell carcinomas (ccRCC), which contributes to the carcinogenesis and drug resistance of ccRCC. Here we show that VHL-deficient ccRCC cells present enhanced cytotoxicity of anthracyclines in a hypoxia-inducible factor-independent manner. By subtractive proteomic analysis coupling with RNAi or overexpression verification, aldehyde dehydrogenase 2 (ALDH2) is found to be transcriptionally regulated by VHL and contributes to enhanced anthracyclines cytotoxicity in ccRCC cells. Furthermore, VHL regulates ALDH2 expression by directly binding the promoter of -130 bp to -160 bp to activate the transcription of hepatocyte nuclear factor 4 alpha (HNF-4α). In addition, a positive correlation is found among the protein expressions of VHL, HNF-4α and ALDH2 in ccRCC samples. These findings will deepen our understanding of VHL function and shed light on precise treatment for ccRCC patients.
Assuntos
Aldeído-Desidrogenase Mitocondrial/genética , Antraciclinas/uso terapêutico , Carcinoma de Células Renais/tratamento farmacológico , Carcinoma de Células Renais/genética , Regulação para Baixo/genética , Neoplasias Renais/tratamento farmacológico , Neoplasias Renais/genética , Proteína Supressora de Tumor Von Hippel-Lindau/metabolismo , Aldeído-Desidrogenase Mitocondrial/metabolismo , Animais , Antraciclinas/farmacologia , Antraciclinas/toxicidade , Carcinoma de Células Renais/patologia , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Regulação para Baixo/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Fator 4 Nuclear de Hepatócito/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Neoplasias Renais/patologia , Masculino , Camundongos Nus , Proteínas de Neoplasias/metabolismo , Proteômica , Transcrição Gênica/efeitos dos fármacosRESUMO
The intra-articular injection of adipose-derived stem cells (ASCs) is a novel potential therapy for patients with osteoarthritis (OA). However, the efficacy of ASCs from different regions of the body remains unknown. This study investigated whether ASCs from subcutaneous or visceral adipose tissue provide the same improvement of OA. Mouse and human subcutaneous and visceral adipose tissue were excised for ASC isolation. Morphology, proliferation, surface markers and adipocyte differentiation of subcutaneous ASCs (S-ASCs) and visceral ASCs (V-ASCs) were analysed. A surgically induced rat model of OA was established, and 4 weeks after the operation, S-ASCs, V-ASCs or phosphate-buffered saline (PBS, control) were injected into the articular cavity. Histology, immunohistochemistry and gene expression analyses were performed 6 weeks after ASC injection. The ability of ASCs to differentiate into chondrocytes was assessed by in vitro chondrogenesis, and the immunosuppressive activity of ASCs was evaluated by co-culturing with macrophages. The proliferation of V-ASCs was significantly greater than that of S-ASCs, but S-ASCs had the greater adipogenic capacity than V-ASCs. In addition, the infracted cartilage treated with S-ASCs showed significantly greater improvement than cartilage treated with PBS or V-ASCs. Moreover, S-ASCs showed better chondrogenic potential and immunosuppression in vitro. Subcutaneous adipose tissue is an effective cell source for cell therapy of OA as it promotes stem cell differentiation into chondrocytes and inhibits immunological reactions.
Assuntos
Gordura Intra-Abdominal/citologia , Osteoartrite do Joelho/terapia , Transplante de Células-Tronco , Células-Tronco/citologia , Gordura Subcutânea/citologia , Animais , Anti-Inflamatórios/metabolismo , Cartilagem Articular/patologia , Condrogênese/genética , Modelos Animais de Doenças , Progressão da Doença , Regulação da Expressão Gênica , Humanos , Injeções Intra-Articulares , Interleucina-6/metabolismo , Lipopolissacarídeos , Óxido Nítrico Sintase Tipo II/metabolismo , Osteoartrite do Joelho/patologia , Fenótipo , Ratos Sprague-Dawley , Células-Tronco/metabolismo , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Besides the conventional carbon sources, acetyl-CoA has recently been shown to be generated from acetate in various types of cancers, where it promotes lipid synthesis and tumour growth. The underlying mechanism, however, remains largely unknown. We find that acetate induces a hyperacetylated state of histone H3 in hypoxic cells. Acetate predominately activates lipogenic genes ACACA and FASN expression by increasing H3K9, H3K27 and H3K56 acetylation levels at their promoter regions, thus enhancing de novo lipid synthesis, which combines with its function as the metabolic precursor for fatty acid synthesis. Acetyl-CoA synthetases (ACSS1, ACSS2) are involved in this acetate-mediated epigenetic regulation. More importantly, human hepatocellular carcinoma with high ACSS1/2 expression exhibit increased histone H3 acetylation and FASN expression. Taken together, this study demonstrates that acetate, in addition to its ability to induce fatty acid synthesis as an immediate metabolic precursor, also functions as an epigenetic metabolite to promote cancer cell survival under hypoxic stress.
Assuntos
Acetatos/metabolismo , Carcinoma Hepatocelular/metabolismo , Hipóxia/metabolismo , Lipogênese , Neoplasias Hepáticas/metabolismo , Acetato-CoA Ligase/metabolismo , Acetilação , Sobrevivência Celular , Epigênese Genética , Ácido Graxo Sintase Tipo I/metabolismo , Regulação Neoplásica da Expressão Gênica , Células Hep G2 , Histona Acetiltransferases/metabolismo , Histonas/metabolismo , HumanosRESUMO
N-glycosylation is one of the most prominent and abundant posttranslational modifications of proteins. It is estimated that over 50% of mammalian proteins undergo glycosylation. However, the analysis of N-glycoproteins has been limited by the available analytical technology. In this study, we comprehensively mapped the N-glycosylation sites in the mouse brain proteome by combining complementary methods, which included seven protease treatments, four enrichment techniques and two fractionation strategies. Altogether, 13492 N-glycopeptides containing 8386 N-glycosylation sites on 3982 proteins were identified. After evaluating the performance of the above methods, we proposed a simple and efficient workflow for large-scale N-glycosylation site mapping. The optimized workflow yielded 80% of the initially identified N-glycosylation sites with considerably less effort. Analysis of the identified N-glycoproteins revealed that many of the mouse brain proteins are N-glycosylated, including those proteins in critical pathways for nervous system development and neurological disease. Additionally, several important biomarkers of various diseases were found to be N-glycosylated. These data confirm that N-glycosylation is important in both physiological and pathological processes in the brain, and provide useful details about numerous N-glycosylation sites in brain proteins.
Assuntos
Encéfalo/metabolismo , Glicoproteínas/metabolismo , Proteoma , Animais , Biomarcadores/metabolismo , Biologia Computacional , Glicosilação , Concentração de Íons de Hidrogênio , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Processamento de Proteína Pós-Traducional , Proteômica , Espectrometria de Massas em TandemRESUMO
AIMS: In our previous study, eEF1A1 was identified to be a new target for protecting brain ischemia injury, but the mechanism remains largely unknown. In this study, we screened the downstream cellular protein molecules interacted with eEF1A1 and found mechanism of eEF1A1 in brain ischemia protection. METHODS AND RESULTS: Through co-immunoprecipitation and mass spectrometry for searching the interaction of proteins with eEF1A1 in bEnd3 cells, HSC70 was identified to be a binding protein of eEF1A1, which was further validated by Western blot and immunofluorescence. eEF1A1 or HSC70 knockdown, respectively, increased OGD-induced apoptosis of brain vascular endothelial cells, which was detected by Annexin V-FITC/PI staining. HSC70 or eEF1A1 knockdown enhances phosphorylated JNK, phosphorylation of c-JUN (Ser63, Ser73), cleaved caspase-9, and cleaved caspase-3 expression, which could be rescued by JNK inhibitor. CONCLUSION: In summary, our data suggest that the presence of chaperone forms of interaction between eEF1A1 and HSC70 in brain vascular endothelial cells, eEF1A1 and HSC70 can play a protective role in the process of ischemic stroke by inhibiting the JNK signaling pathway activation.
Assuntos
Células Endoteliais/metabolismo , Proteínas de Choque Térmico HSC70/metabolismo , MAP Quinase Quinase 4/metabolismo , Fator 1 de Elongação de Peptídeos/metabolismo , Análise de Variância , Animais , Anexina A5/metabolismo , Apoptose/efeitos dos fármacos , Encéfalo/citologia , Células Cultivadas , Células Endoteliais/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Citometria de Fluxo , Proteínas de Choque Térmico HSC70/genética , Imunoprecipitação , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/fisiologia , Espectrometria de Massas , Camundongos , Microscopia Confocal , Fator 1 de Elongação de Peptídeos/genética , RNA Interferente Pequeno/farmacologia , Fatores de TempoRESUMO
The tumor suppressor gene AT-rich interactive domain-containing protein 1A (ARID1A) was frequently mutated in cancers. The modulation mechanism of ARID1A for PI3K/AKT signaling in gastric cancer (GC) remains elusive. Here, we found that depletion of endogenous ARID1A enhanced the in vitro proliferation, colony formation, cellular growth, nutrient uptake and in vivo xenograft tumor growth of GC cells. PI3K/AKT activation by ARID1A-silencing was profiled using a phospho-protein antibody array. The phosphorylation of PDK1, AKT, GSK3ß and 70S6K, and the protein and mRNA expressions of PI3K and PDK1, were upregulated by ARID1A-silencing. Chromatin immunoprecipitation and luciferase reporter assay revealed that ARID1A-involved SWI/SNF complex inhibited PIK3CA and PDK1 transcription by direct binding to their promoters. Serial deletion mutation analyses revealed that the ARID1A central region containing the HIC1-binding domain, but not the ARID DNA-binding domain and the C-terminal domain, was essential for the inhibition of GC cell growth, PI3K/AKT pathway phosphorylation and its transcriptional modulation activity of PIK3CA and PDK1. The proliferation, cellular growth and glucose consumption of ARID1A-deficient GC cells were efficiently prohibited by allosteric inhibitors mk2206 and LY294002, which targeting AKT and PI3K, respectively. Both inhibitors also downregulated the phosphorylation of PI3K/AKT pathway in ARID1A-deficient GC cells. Such cells were sensitized to the treatment of LY294002, and AT7867, another inhibitor of AKT and p70S6K. The administration of LY294002 alone inhibited the in vivo growth of ARID1A- deficient GC cells in mouse xenograft model. Our study provides a novel insight into the modulatory function and mechanism of ARID1A in PI3K/AKT signaling in GC.