RESUMO
The mechanistic target of rapamycin (mTOR) is a central regulator of cell growth and an attractive anticancer target that integrates diverse signals to control cell proliferation. Previous studies using mTOR inhibitors have shown that mTOR targeting suppresses gene expression and cell proliferation. To date, however, mTOR-targeted therapies in cancer have seen limited efficacy, and one key issue is related to the development of evasive resistance. In this manuscript, through the use of a gene targeting mouse model, we have found that inducible deletion of mTOR in hematopoietic stem cells (HSCs) results in a loss of quiescence and increased proliferation. Adaptive to the mTOR loss, mTOR-/- HSCs increase chromatin accessibility and activate global gene expression, contrary to the effects of short-term inhibition by mTOR inhibitors. Mechanistically, such genomic changes are due to a rewiring and adaptive activation of the ERK/MNK/eIF4E signaling pathway that enhances the protein translation of RNA polymerase II, which in turn leads to increased c-Myc gene expression, allowing the HSCs to thrive despite the loss of a functional mTOR pathway. This adaptive mechanism can also be utilized by leukemia cells undergoing long-term mTOR inhibitor treatment to confer resistance to mTOR drug targeting. The resistance can be counteracted by MNK, CDK9, or c-Myc inhibition. These results provide insights into the physiological role of mTOR in mammalian stem cell regulation and implicate a mechanism of evasive resistance in the context of mTOR targeting.
Assuntos
Proliferação de Células/efeitos dos fármacos , Células-Tronco Hematopoéticas/metabolismo , Sirolimo/farmacologia , Serina-Treonina Quinases TOR/antagonistas & inibidores , Serina-Treonina Quinases TOR/genética , Animais , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/genética , Cromatina/metabolismo , Sequenciamento de Cromatina por Imunoprecipitação , Quinase 9 Dependente de Ciclina/metabolismo , Fator de Iniciação 4E em Eucariotos/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Marcação de Genes , Genes myc/genética , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/genética , Camundongos , Camundongos Knockout , Fosforilação/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , RNA Polimerase II/metabolismo , Serina-Treonina Quinases TOR/metabolismoRESUMO
Hematopoietic stem and progenitor cell (HSPC) formation and lineage differentiation involve gene expression programs orchestrated by transcription factors and epigenetic regulators. Genetic disruption of the chromatin remodeler chromodomain-helicase-DNA-binding protein 7 (CHD7) expanded phenotypic HSPCs, erythroid, and myeloid lineages in zebrafish and mouse embryos. CHD7 acts to suppress hematopoietic differentiation. Binding motifs for RUNX and other hematopoietic transcription factors are enriched at sites occupied by CHD7, and decreased RUNX1 occupancy correlated with loss of CHD7 localization. CHD7 physically interacts with RUNX1 and suppresses RUNX1-induced expansion of HSPCs during development through modulation of RUNX1 activity. Consequently, the RUNX1:CHD7 axis provides proper timing and function of HSPCs as they emerge during hematopoietic development or mature in adults, representing a distinct and evolutionarily conserved control mechanism to ensure accurate hematopoietic lineage differentiation.
Assuntos
Subunidade alfa 2 de Fator de Ligação ao Core , Proteínas de Ligação a DNA , Hematopoese , Animais , Diferenciação Celular , Linhagem Celular , Subunidade alfa 2 de Fator de Ligação ao Core/química , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Feminino , Células-Tronco Hematopoéticas , Humanos , Masculino , Camundongos , Baço/citologia , Peixe-ZebraRESUMO
Invasive growth of glioblastoma makes residual tumor unremovable by surgery and leads to disease relapse. Temozolomide is widely used first-line chemotherapy drug to treat glioma patients, but development of temozolomide resistance is almost inevitable. Ferroptosis, an iron-dependent form of non-apoptotic cell death, is found to be related to temozolomide response of gliomas. However, whether inducing ferroptosis could affect invasive growth of glioblastoma cells and which ferroptosis-related regulators were involved in temozolomide resistance are still unclear. In this study, we treated glioblastoma cells with RSL3, a ferroptosis inducer, in vitro (cell lines) and in vivo (subcutaneous and orthotopic animal models). The treated glioblastoma cells with wild-type or mutant IDH1 were subjected to RNA sequencing for transcriptomic profiling. We then analyze data from our RNA sequencing and public TCGA glioma database to identify ferroptosis-related biomarkers for prediction of prognosis and temozolomide resistance in gliomas. Analysis of transcriptome data from RSL3-treated glioblastoma cells suggested that RSL3 could inhibit glioblastoma cell growth and suppress expression of genes involved in cell cycle. RSL3 effectively reduced mobility of glioblastoma cells through downregulation of critical genes involved in epithelial-mesenchymal transition. Moreover, RSL3 in combination with temozolomide showed suppressive efficacy on glioblastoma cell growth, providing a promising therapeutic strategy for glioblastoma treatment. Although temozolomide attenuated invasion of glioblastoma cells with mutant IDH1 more than those with wild-type IDH1, the combination of RSL3 and temozolomide similarly impaired invasive ability of glioblastoma cells in spite of IDH1 status. Finally, we noticed that both ferritin heavy chain 1 and ferritin light chain predicted unfavorable prognosis of glioma patients and were significantly correlated with mRNA levels of methylguanine methyltransferase as well as temozolomide resistance. Altogether, our study provided rationale for combination of RSL3 with temozolomide to suppress glioblastoma cells and revealed ferritin heavy chain 1 and ferritin light chain as biomarkers to predict prognosis and temozolomide resistance of glioma patients.
Assuntos
Neoplasias Encefálicas , Ferroptose , Glioblastoma , Glioma , Animais , Apoferritinas/farmacologia , Apoferritinas/uso terapêutico , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos/genética , Glioblastoma/tratamento farmacológico , Glioblastoma/genética , Glioblastoma/metabolismo , Glioma/tratamento farmacológico , Glioma/genética , Glioma/metabolismo , Temozolomida/farmacologia , Temozolomida/uso terapêuticoRESUMO
Cardiopulmonary bypass (CPB), though indispensable in many cardiac surgery procedures, has several undesirable consequences. The aim of this study was to identify potential genes that may reduce the inflammatory response and complications after CPB. The GSE132176 dataset was selected from the Gene Expression Omnibus (GEO) database and included 10 patients with tetralogy of Fallot and 10 patients with an atrial septal defect who underwent CPB surgery. TSV files were downloaded after GEO2R processing. Protein-protein interaction analysis of common differentially expressed genes (DEGs) was performed using the Search Tool for the Retrieval of Interacting Genes. Gene modules and hub genes were visualized in the protein-protein interaction network using Cytoscape. Enrichment analysis was performed for all important DEGs, modular genes, and hub genes. A total of 72 DEGs were screened, including two functional and one hub gene module. FOS modular genes were primarily enriched in NGF-stimulated transcription, spinal cord injury, and PID AP1 pathway. The ATF3 modular gene was mainly enriched in cytomegalovirus infection and transcriptional misregulation in cancer. Hub gene modules were primarily enriched in the PID AP1 pathway, positive regulation of pri-miRNA transcription by RNA polymerase II, and the PID ATF2 pathway. FOS, JUN, ATF3, and EGR1 were the four most important hub genes; the top three hub genes were involved in the formation of AP-1 and enriched in the AP-1 pathway. Finally, we measured the expression levels of these four genes in patients undergoing CPB via qRT-PCR, and the results were consistent with those obtained in bioinformatic analysis. FOS, JUN, ATF3, and EGR1 and the AP-1 pathway may play key roles in inflammation and complications caused by CPB.
Assuntos
Biologia Computacional , Perfilação da Expressão Gênica , Ponte Cardiopulmonar/efeitos adversos , Biologia Computacional/métodos , Perfilação da Expressão Gênica/métodos , Humanos , Inflamação/genética , Inflamação/prevenção & controle , Fator de Transcrição AP-1RESUMO
Identifying signaling pathways that regulate hematopoietic stem and progenitor cell (HSPC) formation in the embryo will guide efforts to produce and expand HSPCs ex vivo. Here we show that sterile tonic inflammatory signaling regulates embryonic HSPC formation. Expression profiling of progenitors with lymphoid potential and hematopoietic stem cells (HSCs) from aorta/gonad/mesonephros (AGM) regions of midgestation mouse embryos revealed a robust innate immune/inflammatory signature. Mouse embryos lacking interferon γ (IFN-γ) or IFN-α signaling and zebrafish morphants lacking IFN-γ and IFN-Ï activity had significantly fewer AGM HSPCs. Conversely, knockdown of IFN regulatory factor 2 (IRF2), a negative regulator of IFN signaling, increased expression of IFN target genes and HSPC production in zebrafish. Chromatin immunoprecipitation (ChIP) combined with sequencing (ChIP-seq) and expression analyses demonstrated that IRF2-occupied genes identified in human fetal liver CD34(+) HSPCs are actively transcribed in human and mouse HSPCs. Furthermore, we demonstrate that the primitive myeloid population contributes to the local inflammatory response to impact the scale of HSPC production in the AGM region. Thus, sterile inflammatory signaling is an evolutionarily conserved pathway regulating the production of HSPCs during embryonic development.
Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Células-Tronco Hematopoéticas/citologia , Imunidade Inata/genética , Imunidade Inata/imunologia , Transdução de Sinais , Animais , Antígenos Ly/genética , Antígenos Ly/metabolismo , Proliferação de Células/genética , Células Cultivadas , Citocinas/imunologia , Embrião de Mamíferos , Embrião não Mamífero , Perfilação da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Inflamação/genética , Inflamação/imunologia , Interferons/genética , Interferons/metabolismo , Camundongos , Peixe-Zebra/embriologiaRESUMO
Circular RNAs (circRNAs) are essential regulators associated with many cardiac conditions, including myocardial infarction (MI). This study aimed to explore circRNA expression during MI development in an animal model and in hypoxia/reoxygenation (H/R)-treated cardiomyocytes. Microarray and real-time quantitative PCR showed that the circRNA PVT1 (circPVT1) was expressed at high levels in MI tissues and H/R-triggered cardiomyocytes. Loss-of-function assays were utilized for examining the influence of circPVT1 on cardiac function and cardiomyocyte properties. Cardiac function was measured by echocardiography at 7 d after MI. Reduced circPVT1 expression significantly decreased MI-triggered myocardial infarct size by 60% and prevented MI-triggered reductions in fractional shortening (%FS) and ejection fraction (EF%). Results of LDH, CCK-8, EdU staining, colony formation assays, and flow cytometry showed that circPVT1 silencing restored cell viability and proliferation while decreased apoptosis. Mechanistic experiments indicated that microRNAs (miR)-125b and miR-200a associated with circPVT1. We demonstrated that circPVT1 functioned as a competitive endogenous RNA (ceRNA) to sponge both miR-125b and miR-200a. Gain-of-function assays showed that miR-125b and miR-200a upregulation partially eliminated the effects of circPVT1 on cardiomyocyte properties. In addition, we found that the previously reported p53/TRAF6, SIRT7, Keap1/Nrf2, and PDCD4 pathways were regulated by the circPVT1/miR-125b/miR-200a axis. In conclusion, our study suggests that circPVT1 protects the myocardium from MI and H/R injury by preventing miR-125b- and miR-200a-mediated apoptotic signaling.
Assuntos
MicroRNAs/genética , Interferência de RNA/fisiologia , RNA Circular/genética , RNA Longo não Codificante/genética , Traumatismo por Reperfusão/genética , Animais , Apoptose/genética , Proliferação de Células/genética , Sobrevivência Celular/genética , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Miocárdio/metabolismo , Miócitos Cardíacos/metabolismo , Ratos , Transdução de Sinais/genética , Regulação para Cima/genéticaRESUMO
BACKGROUND Interleukin-36 has been demonstrated to be involved in inflammatory responses. Inflammatory responses due to ischemia-reperfusion injury following cardiopulmonary bypass (CPB) can cause heart dysfunction or damage. MATERIAL AND METHODS The CPB models were constructed in IL-36R-/-, IL-36RN-/-, and wild-type SD rats. Ultrasonic cardiography and ELISA were used to evaluate the cardiac function and measuring myocardial biomarker levels in different groups. TUNEL assay was used to evaluate apoptosis. Western blot assays and RT-PCR were performed to measure the expression of chemokines and secondary inflammatory cytokines in the heart. Oxidative stress in tissue and cultured cells was assessed using a DCFH-DA fluorescence probe and quantification of superoxide dismutase activity. RESULTS Improved systolic function and decreased serum levels of myocardial damage biomarkers were found in IL-36R-/- rats compared to WT rats, while worse cardiac function and cardiomyocyte IR injury were observed in IL-36RN-/- rats compared to WT rats. TUNEL staining and Western blot analyses found that cardiomyocyte apoptosis and inflammation were significantly lower in the hearts of IL-36R-/- rats compared with that of WT rats. Oxidative stress was significantly lower in IL-36R-/- rats compared to WT rats. iNOS expression was significantly reduced, while eNOS expression was increased in the hearts of IL-36R-/- rats. Silencing of IL-36R expression in vitro activated SIRT1/FOXO1/p53 signaling in cardiomyocytes. CONCLUSIONS IL-36R deficiency in cardiomyocytes repressed infiltration of bone marrow-derived inflammatory cells and oxidative stress dependent on SIRT1-FOXO1 signaling, thus protecting cardiomyocytes and improving cardiac function in CPB model rats.
Assuntos
Ponte Cardiopulmonar/efeitos adversos , Traumatismo por Reperfusão Miocárdica/imunologia , Miocárdio/patologia , Miócitos Cardíacos/patologia , Receptores de Interleucina/deficiência , Animais , Modelos Animais de Doenças , Técnicas de Inativação de Genes , Humanos , Masculino , Traumatismo por Reperfusão Miocárdica/patologia , Miocárdio/citologia , Miocárdio/imunologia , Miócitos Cardíacos/imunologia , Proteínas do Tecido Nervoso/metabolismo , Estresse Oxidativo/genética , Estresse Oxidativo/imunologia , Ratos , Ratos Transgênicos , Receptores de Interleucina/genética , Receptores de Interleucina/metabolismo , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Sirtuína 1/metabolismoRESUMO
Runx1 and Cbfß are critical for the establishment of definitive hematopoiesis and are implicated in leukemic transformation. Despite the absolute requirements for these factors in the development of hematopoietic stem cells and lymphocytes, their roles in the development of bone marrow progenitor subsets have not been defined. Here, we demonstrate that Cbfß is essential for the development of Flt3(+) macrophage-dendritic cell (DC) progenitors in the bone marrow and all DC subsets in the periphery. Besides the loss of DC progenitors, pan-hematopoietic Cbfb-deficient mice also lack CD105(+) erythroid progenitors, leading to severe anemia at 3 to 4 months of age. Instead, Cbfb deficiency results in aberrant progenitor differentiation toward granulocyte-macrophage progenitors (GMPs), resulting in a myeloproliferative phenotype with accumulation of GMPs in the periphery and cellular infiltration of the liver. Expression of the transcription factor Irf8 is severely reduced in Cbfb-deficient progenitors, and overexpression of Irf8 restors DC differentiation. These results demonstrate that Runx proteins and Cbfß restrict granulocyte lineage commitment to facilitate multilineage hematopoietic differentiation and thus identify their novel tumor suppressor function in myeloid leukemia.
Assuntos
Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Subunidade beta de Fator de Ligação ao Core/metabolismo , Células Dendríticas/metabolismo , Transtornos Mieloproliferativos/metabolismo , Células-Tronco/metabolismo , Tirosina Quinase 3 Semelhante a fms/metabolismo , Anemia/genética , Anemia/metabolismo , Anemia/patologia , Animais , Células da Medula Óssea/metabolismo , Diferenciação Celular/genética , Células Cultivadas , Subunidade alfa 1 de Fator de Ligação ao Core/genética , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Subunidade alfa 3 de Fator de Ligação ao Core/genética , Subunidade alfa 3 de Fator de Ligação ao Core/metabolismo , Subunidade beta de Fator de Ligação ao Core/genética , Citometria de Fluxo , Expressão Gênica , Granulócitos/metabolismo , Hematopoese/genética , Fatores Reguladores de Interferon/genética , Fatores Reguladores de Interferon/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Transtornos Mieloproliferativos/genética , Transtornos Mieloproliferativos/patologia , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Cancer is one of the major diseases that seriously threaten human health. Timely screening is beneficial to the cure of cancer. There are some shortcomings in current diagnosis methods, so it is very important to find a low-cost, fast, and nondestructive cancer screening technology. In this study, we demonstrated that serum Raman spectroscopy combined with a convolutional neural network model can be used for the diagnosis of four types of cancer including gastric cancer, colon cancer, rectal cancer, and lung cancer. Raman spectra database containing four types of cancer and healthy controls was established and a one-dimensional convolutional neural network (1D-CNN) was constructed. The classification accuracy of the Raman spectra combined with the 1D-CNN model was 94.5%. A convolutional neural network (CNN) is regarded as a black box, and the learning mechanism of the model is not clear. Therefore, we tried to visualize the CNN features of each convolutional layer in the diagnosis of rectal cancer. Overall, Raman spectroscopy combined with the CNN model is an effective tool that can be used to distinguish different cancer from healthy controls.
Assuntos
Neoplasias do Colo , Neoplasias Pulmonares , Neoplasias Retais , Neoplasias Gástricas , Humanos , Neoplasias Gástricas/diagnóstico , Análise Espectral Raman , Redes Neurais de Computação , Neoplasias Pulmonares/diagnósticoRESUMO
As a transcription factor in the RUNT domain core-binding factor family, RUNX1 is crucial in multiple stages of hematopoiesis, and its mutation can cause familial platelet disorder with a predisposition to acute myeloid leukemia. Previous work has established that RUNX1 is involved in the maturation of megakaryocytes (MKs) and the production of platelets. Recent studies have shown that there exists a subpopulation of hematopoietic stem cells (HSCs) with relatively high expression of von Willebrand factor and CD41 at the apex of the HSC hierarchy, termed MK-HSCs, which can give rise to MKs without going through the traditional differentiation trajectory from HSC via MPP (multipotent progenitors) and MEP (megakaryocyte-erythroid progenitor). Here, by using Runx1F/FMx1-Cre mouse model, we discovered that the MK-HSC to MK direct differentiation can occur within 1 cell division, and RUNX1 is an important regulator in the process. Runx1 knockout results in a drastic decrease in platelet counts and a severe defect in the differentiation from MK-HSCs to MKs. Single cell RNA sequencing (RNAseq) analysis shows that MK-HSCs have a distinct gene expression signature compared with non-MK-HSCs, and Runx1 deletion alters the platelet and MK-related gene expression in MK-HSCs. Furthermore, bulk RNAseq and Cut&Run analyses show that RUNX1 binds to multiple essential MK or platelet developmental genes, such as Spi1, Selp, and Itga2b and regulates their expressions in MK-HSCs. Thus, by modulating the expression of MK-related genes, RUNX1 governs the direct differentiation from MK-HSCs to MKs and platelets.
Assuntos
Subunidade alfa 2 de Fator de Ligação ao Core , Megacariócitos , Animais , Camundongos , Megacariócitos/metabolismo , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Células-Tronco Hematopoéticas/metabolismo , Hematopoese , Diferenciação Celular/genéticaRESUMO
mTOR, as a serine/threonine kinase, is a widely pursued anticancer target. Multiple clinical trials of mTOR kinase inhibitors are ongoing, but their specificity and safety features remain lacking. Here, we have employed an inducible kinase-inactive D2338A mTOR knock-in mouse model (mTOR-/KI) together with a mTOR conditional knockout model (mTOR-/-) to assess the kinase-dependent/-independent function of mTOR in hematopoiesis and the on-/off-target effects of mTOR kinase inhibitor AZD2014. Despite exhibiting many similar phenotypes to mTOR-/- mice in hematopoiesis, the mTOR-/KI mice survived longer and showed differences in hematopoietic progenitor cells compared to mTOR-/- mice, suggesting a kinase-independent function of mTOR in hematopoiesis. Gene expression signatures in hematopoietic stem cells (HSCs) further revealed both kinase-dependent and independent effects of mTOR. AZD2014, a lead mTOR kinase inhibitor, appeared to work mostly on-target in suppressing mTOR kinase activity, mimicking that of mTOR-/KI HSCs in transcriptome analysis, but it also induced a small set of off-target responses in mTOR-/KI HSCs. In murine and human myeloid leukemia, besides kinase-inhibitory on-target effects, AZD2014 displayed similar off-target and growth-inhibitory cytostatic effects. These studies provide new insights into kinase-dependent/-independent effects of mTOR in hematopoiesis and present a genetic means for precisely assessing the specificity of mTOR kinase inhibitors.
Assuntos
Morfolinas , Serina-Treonina Quinases TOR , Camundongos , Humanos , Animais , Serina-Treonina Quinases TOR/metabolismo , Morfolinas/farmacologia , Benzamidas/farmacologia , Pirimidinas/farmacologia , HematopoeseRESUMO
BACKGROUND: Acute myeloid leukemia (AML) is a hematologic malignancy with genetic alterations. RUNX1, which is an essential transcription factor for hematopoiesis, is frequently mutated in AML. Loss-of-function mutation of RUNX1 is correlated with poor prognosis of AML patients. It is urgent to reveal the underlying mechanism. RESEARCH DESIGN AND METHODS: TCGA AML, GSE106291, GSE142700, and GSE67609 datasets were used. R package was used to define differentially expressed miRNAs, miRNA target genes, RUNX1-related gene, RUNX directly regulating genes, and so on. The relationship of gene expression with overall survival was analyzed by Cox regression. KEGG and GO analyses were applied to the above-mentioned genesets and overlapped genes. Alteration and importance of MAPK pathway were validated in K562 cells by Western blotting and apoptosis assay in vitro. RESULTS: RUNX1 regulated MAPK pathway indirectly and directly. MAPK pathway was altered in K562-cell-induced mutated RUNX1, and these cells were more sensitive to AraC after p38 was inhibited. CONCLUSIONS: RUNX1 could modulate MAPK pathway, which may provide a potential therapeutic target for AML patients with RUNX1 mutations.
Assuntos
Subunidade alfa 2 de Fator de Ligação ao Core , Leucemia Mieloide Aguda , MicroRNAs , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Humanos , Leucemia Mieloide Aguda/patologia , Mutação , Transdução de SinaisRESUMO
As a frequent disease affecting the nervous system, cerebral infarction has emerged as a major cause of disability and elicits disorders in motor, sensation, and cognition as sequelae. No clear mechanism has been known in meridian massage despite it having been proved to be an effective therapeutic option. The study was carried out to explore the treatment of meridian massage on cerebral ischemia in rats and its effects on motor function restoration and nerve cell's ultrastructure in the ischemic territory. The alleviated nerve damages and recovered injured brain tissues were found in the cerebral infarction model of SD rats after meridian massage. Expressions of miR-206 and the brain-derived neurotrophic factor (BDNF) in the gastrocnemius muscle were all well observed. The effects of miR-206 on BDNF were testified by overexpressed and interfered miR-206 in the C2C12 myoblast. Moreover, at the molecular level, meridian massage downregulated miR-206 expression at an elevated level of BDNF. Consequently, meridian massage exerts a vital role in promoting cerebral ischemia restoration, which is expected to provide an addition to the application of traditional Chinese medicine (TCM) in the reconstruction and treatment of cerebral ischemia.
RESUMO
Hematopoietic stem cells (HSCs) adapt their metabolism to maintenance and proliferation; however, the mechanism remains incompletely understood. Here, we demonstrated that homeostatic HSCs exhibited high amino acid (AA) catabolism to reduce cellular AA levels, which activated the GCN2-eIF2α axis, a protein synthesis inhibitory checkpoint to restrain protein synthesis for maintenance. Furthermore, upon proliferation conditions, HSCs enhanced mitochondrial oxidative phosphorylation (OXPHOS) for higher energy production but decreased AA catabolism to accumulate cellular AAs, which inactivated the GCN2-eIF2α axis to increase protein synthesis and coupled with proteotoxic stress. Importantly, GCN2 deletion impaired HSC function in repopulation and regeneration. Mechanistically, GCN2 maintained proteostasis and inhibited Src-mediated AKT activation to repress mitochondrial OXPHOS in HSCs. Moreover, the glycolytic metabolite, NAD+ precursor nicotinamide riboside (NR), accelerated AA catabolism to activate GCN2 and sustain the long-term function of HSCs. Overall, our study uncovered direct links between metabolic alterations and translation control in HSCs during homeostasis and proliferation.
Assuntos
Fator de Iniciação 2 em Eucariotos , Proteostase , Aminoácidos/metabolismo , Fator de Iniciação 2 em Eucariotos/metabolismo , Células-Tronco Hematopoéticas/metabolismo , Fosforilação Oxidativa , FosforilaçãoRESUMO
AML1-ETO is the chimeric protein product of the t(8;21) in acute myeloid leukemia. The ETO portion of the fusion protein includes the eTAFH domain, which is homologous to several TATA binding protein-associated factors (TAFs) and interacts with E proteins (E2A and HEB). It has been proposed that AML1-ETO-mediated silencing of E protein function might be important for t(8;21) leukemogenesis. Here, we determined the solution structure of a complex between the AML1-ETO eTAFH domain and an interacting peptide from HEB. On the basis of the structure, key residues in AML1-ETO for HEB association were mutated. These mutations do not impair the ability of AML1-ETO to enhance the clonogenic capacity of primary mouse bone marrow cells and do not eliminate its ability to repress proliferation or granulocyte differentiation. Therefore, the eTAFH-E protein interaction appears to contribute relatively little to the activity of AML1-ETO.
Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Proteínas de Ligação a DNA/metabolismo , Leucemia Mieloide Aguda/metabolismo , Proteínas de Fusão Oncogênica/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Fatores de Transcrição/metabolismo , Animais , Células COS , Diferenciação Celular/imunologia , Divisão Celular/imunologia , Células Cultivadas , Chlorocebus aethiops , Subunidade alfa 2 de Fator de Ligação ao Core/química , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/genética , Granulócitos/citologia , Granulócitos/fisiologia , Leucemia Mieloide Aguda/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Ressonância Magnética Nuclear Biomolecular , Proteínas de Fusão Oncogênica/química , Proteínas de Fusão Oncogênica/genética , Estrutura Secundária de Proteína , Estrutura Terciária de Proteína , Proteínas Proto-Oncogênicas/química , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , Fator de Transcrição STAT6/metabolismo , Relação Estrutura-Atividade , Fatores de Transcrição/química , Fatores de Transcrição/genéticaRESUMO
BACKGROUND: Finding effective prognostic signatures is of great urgency due to the high risk of recurrence and progression of bladder cancer (BC). Although a lot of genetic alterations are involved in the carcinogenesis, none of them were referred in the current risk group stratifications. In this study, we aimed to find significant copy number variations (CNVs) to predict prognosis for BC patients. METHODS: CNVs with high aberration frequencies in BC were explored by array-based comparative genomic hybridization in 65 tumor samples. Candidates were validated in independent groups of BC tumor samples (n=219) and urine samples (n=123). 3D digital PCR was applied for detecting accurate gene copy numbers in BC urine. In order to explore the prognostic value of candidate CNVs, all enrolled patients were followed up for the disease-free survival (DFS). Cox proportional hazards regression analysis was performed to find the independent prognostic factors for DFS. RESULTS: CNVs of CEP63, FOSL2 and PAQR6 with high aberration frequencies (67.7%, 56.9% and 60.0%, respectively) were found in BC tumors. Copy numbers of CEP63, FOSL2 and PAQR6 were gained in 219 tumor samples. CNVs of CEP63 and FOSL2 were correlated with advanced tumor stage and high grade. Retrospective analysis (median follow-up time: 69 months) revealed that CNVs of CEP63 and FOSL2 were independent prognostic factors for DFS of non-muscle-invasive bladder cancer (NMIBC) patients, while CNVs of FOSL2 and PAQR6 were independent prognostic factors for DFS of muscle-invasive bladder cancer (MIBC) patients. Models for predicting DFS were constructed based on CNVs of three genes. Patients with high prognostic indexes tended to have poor DFS. Prognostic index can also help to identify those with worse outcomes among high risk NMIBC patients. Copy number gains of CEP63 and FOSL2 in urine were found to be significantly correlated with poor DFS of NMIBC patients. CONCLUSIONS: CNVs of CEP63, FOSL2 and PAQR6 were capable of predicting DFS and may serve as promising signatures for prognosis of BC.
RESUMO
BACKGROUND: Autosomal recessive dystrophic epidermolysis bullosa (RDEB) is an incurable and severe inherited skin disorder characterized by recurrent blistering at the sublamina densa beneath the cutaneous basement membrane. It is caused by biallelic loss-of-function mutation in the gene encoding type VII collagen (COL7A1). This study aimed to identify the causative variants of a Chinese RDEB patient and further provide prenatal diagnosis for the ongoing risk pregnancy of the proband's mother. METHODS: Clinical exome sequencing (CES) has been performed and an in-house pipeline was used to conduct a phenotype-driven data analysis. A minigene assay was used to verify the pathogenicity of a novel splice site variant in the COL7A1. RESULTS: Here we report two compound heterozygous variants in COL7A1, c.3867delT (p.G1290Efs*35) and c.5532+4_5532+5delAG, identified in a RDEB patient by CES. The minigene assay confirmed that thec.5532+4_5532+5delAGchange was a noncanonic splice site variant leading to in an in-frame deletion of exon 64. Prenatal diagnosis indicated that the present pregnancy of the patient's mother was not affected. CONCLUSION: Our study expands the mutation spectrum of COL7A1 and demonstrated that CES and minigene assays were efficient tools for RDEB molecular diagnoses.
Assuntos
Colágeno Tipo VII/genética , Epidermólise Bolhosa Distrófica/genética , Alelos , Pré-Escolar , Colágeno Tipo VII/metabolismo , Epidermólise Bolhosa Distrófica/patologia , Feminino , Células HeLa , Humanos , Mutação , Fenótipo , Sítios de Splice de RNARESUMO
Numerous long noncoding RNAs (lncRNAs) are aberrantly expressed in pancreatic cancer (PC); however, their functions and mechanisms in cancer progression are largely unknown. In this study, we identified a novel PC-associated lncRNA, RUNX1-IT1, that was significantly upregulated in PC patient samples from multiple centers and associated with poor prognosis. In vitro and in vivo, alterations in RUNX1-IT1 expression markedly affected PC proliferation, migration and invasion. RUNX1-IT1 contributed to the progression of PC by interacting with the adjacent gene RUNX1. Rescue experiments showed that RUNX1 reduced the cancer-promoting effect of RUNX1-IT1. RNA-seq analysis after silencing RUNX1-IT1 and RUNX1 highlighted alterations in the common target C-FOS. Mechanistically, we demonstrated that RUNX1-IT1 was a trans-acting factor that participated in the proliferation, migration and invasion of PC by recruiting RUNX1 to the C-FOS gene promoter. Furthermore, RUNX1-IT1 enhanced the transcription of the RUNX1 gene, indicating its potential as a cis-regulatory RNA involved in the upstream regulation of RUNX1. Overall, RUNX1-IT1 is a crucial oncogenic lncRNA that activates C-FOS expression by regulating and recruiting RUNX1 and is a potential prognostic biomarker and therapeutic target for PC.
Assuntos
Movimento Celular/genética , Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Proteínas Proto-Oncogênicas c-fos/genética , RNA Longo não Codificante/metabolismo , Transcrição Gênica , Animais , Sequência de Bases , Linhagem Celular Tumoral , Proliferação de Células/genética , Progressão da Doença , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Camundongos Nus , Invasividade Neoplásica , Metástase Neoplásica , Prognóstico , Regiões Promotoras Genéticas/genética , Ligação Proteica , Proteínas Proto-Oncogênicas c-fos/metabolismo , RNA Longo não Codificante/genética , Regulação para Cima/genéticaRESUMO
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
RESUMO
An amendment to this paper has been published and can be accessed via a link at the top of the paper.