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1.
Nat Commun ; 11(1): 4516, 2020 09 09.
Artigo em Inglês | MEDLINE | ID: mdl-32908137

RESUMO

Acinar metaplasia is an initial step in a series of events that can lead to pancreatic cancer. Here we perform single-cell RNA-sequencing of mouse pancreas during the progression from preinvasive stages to tumor formation. Using a reporter gene, we identify metaplastic cells that originated from acinar cells and express two transcription factors, Onecut2 and Foxq1. Further analyses of metaplastic acinar cell heterogeneity define six acinar metaplastic cell types and states, including stomach-specific cell types. Localization of metaplastic cell types and mixture of different metaplastic cell types in the same pre-malignant lesion is shown. Finally, single-cell transcriptome analyses of tumor-associated stromal, immune, endothelial and fibroblast cells identify signals that may support tumor development, as well as the recruitment and education of immune cells. Our findings are consistent with the early, premalignant formation of an immunosuppressive environment mediated by interactions between acinar metaplastic cells and other cells in the microenvironment.


Assuntos
Células Acinares/patologia , Carcinoma Ductal Pancreático/genética , Regulação Neoplásica da Expressão Gênica , Pâncreas/patologia , Neoplasias Pancreáticas/genética , Lesões Pré-Cancerosas/genética , Animais , Animais Geneticamente Modificados , Biópsia , Carcinoma Ductal Pancreático/mortalidade , Carcinoma Ductal Pancreático/patologia , Carcinoma Ductal Pancreático/cirurgia , Diferenciação Celular , Modelos Animais de Doenças , Feminino , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/metabolismo , Heterogeneidade Genética , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Humanos , Estimativa de Kaplan-Meier , Masculino , Metaplasia/genética , Camundongos , Mutação , Pâncreas/citologia , Pâncreas/cirurgia , Pancreatectomia , Neoplasias Pancreáticas/mortalidade , Neoplasias Pancreáticas/patologia , Neoplasias Pancreáticas/cirurgia , Lesões Pré-Cancerosas/patologia , Proteínas Proto-Oncogênicas p21(ras)/genética , RNA-Seq , Análise de Célula Única , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Microambiente Tumoral/genética
2.
PLoS One ; 15(8): e0237403, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32790713

RESUMO

Genome duplication leads to an emergence of gene paralogs that are essentially free to undergo the process of neofunctionalization, subfunctionalization or degeneration (gene loss). Onecut1 (Oc1) and Onecut2 (Oc2) transcription factors, encoded by paralogous genes in mammals, are expressed in precursors of horizontal cells (HCs), retinal ganglion cells and cone photoreceptors. Previous studies have shown that ablation of either Oc1 or Oc2 gene in the mouse retina results in a decreased number of HCs, while simultaneous deletion of Oc1 and Oc2 leads to a complete loss of HCs. Here we study the genetic redundancy between Oc1 and Oc2 paralogs and focus on how the dose of Onecut transcription factors influences abundance of individual retinal cell types and overall retina physiology. Our data show that reducing the number of functional Oc alleles in the developing retina leads to a gradual decrease in the number of HCs, progressive thinning of the outer plexiform layer and diminished electrophysiology responses. Taken together, these observations indicate that in the context of HC population, the alleles of Oc1/Oc2 paralogous genes are mutually interchangeable, function additively to support proper retinal function and their molecular evolution does not follow one of the typical routes after gene duplication.


Assuntos
Fator 6 Nuclear de Hepatócito/genética , Proteínas de Homeodomínio/genética , Retina/fisiologia , Fatores de Transcrição/genética , Alelos , Células Amácrinas/metabolismo , Células Amácrinas/patologia , Animais , Células Ependimogliais/metabolismo , Células Ependimogliais/patologia , Olho/crescimento & desenvolvimento , Olho/patologia , Loci Gênicos , Genótipo , Fator 6 Nuclear de Hepatócito/metabolismo , Proteínas de Homeodomínio/metabolismo , Camundongos , Camundongos Transgênicos , Retina/citologia , Retina/patologia , Células Bipolares da Retina/metabolismo , Células Bipolares da Retina/patologia , Células Fotorreceptoras Retinianas Cones/metabolismo , Células Fotorreceptoras Retinianas Cones/patologia , Células Ganglionares da Retina/citologia , Células Ganglionares da Retina/metabolismo , Fatores de Transcrição/metabolismo
3.
Nat Commun ; 11(1): 4267, 2020 08 26.
Artigo em Inglês | MEDLINE | ID: mdl-32848148

RESUMO

While footprinting analysis of ATAC-seq data can theoretically enable investigation of transcription factor (TF) binding, the lack of a computational tool able to conduct different levels of footprinting analysis has so-far hindered the widespread application of this method. Here we present TOBIAS, a comprehensive, accurate, and fast footprinting framework enabling genome-wide investigation of TF binding dynamics for hundreds of TFs simultaneously. We validate TOBIAS using paired ATAC-seq and ChIP-seq data, and find that TOBIAS outperforms existing methods for bias correction and footprinting. As a proof-of-concept, we illustrate how TOBIAS can unveil complex TF dynamics during zygotic genome activation in both humans and mice, and propose how zygotic Dux activates cascades of TFs, binds to repeat elements and induces expression of novel genetic elements.


Assuntos
Sequenciamento de Cromatina por Imunoprecipitação/métodos , Fatores de Transcrição/metabolismo , Ativação Transcricional , Zigoto/metabolismo , Animais , Sítios de Ligação/genética , Desenvolvimento Embrionário/genética , Epigênese Genética , Feminino , Genoma Humano , Proteínas de Homeodomínio/metabolismo , Humanos , Cinética , Camundongos , Regiões Promotoras Genéticas , Estudo de Prova de Conceito , Ligação Proteica/genética , Especificidade da Espécie
4.
PLoS One ; 15(8): e0236811, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32745141

RESUMO

In this study, we aimed to investigate the molecular biomarkers that are pivotal for the development and progression of gastric cancer (GC). We analyzed clinical specimens using RNA sequencing to identify the target genes. We found that the expression of HOXC6 mRNA was upregulated with the progression of cancer, which was validated by quantitative real time PCR and RNA in-situ hybridization. To compare the protein expression of HOXC6, we evaluated GC and normal gastric tissue samples using western blot analysis and immunohistochemistry. We detected significantly higher levels of HOXC6 in the GC tissues than in the normal controls at both mRNA and protein levels. The expression levels of HOXC6 mRNA in patients with advanced gastric cancer (AGC) were significantly higher than those in patients with early gastric cancer (EGC). Kaplan-Meier curves showed that high expression of HOXC6 mRNA is significantly associated with poor clinical prognosis. Our findings suggest that HOXC6 mRNA may be a novel biomarker and can be potentially valuable in predicting the prognosis of GC patients. Especially, HOXC6 mRNA in-situ hybridization may be a diagnostic tool for predicting prognosis of individual GC patients.


Assuntos
Proteínas de Homeodomínio , Neoplasias Gástricas , Idoso , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Progressão da Doença , Feminino , Regulação Neoplásica da Expressão Gênica , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Humanos , Imuno-Histoquímica , Hibridização In Situ , Linfonodos/patologia , Masculino , Pessoa de Meia-Idade , Prognóstico , RNA Mensageiro/metabolismo , Neoplasias Gástricas/diagnóstico , Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/secundário
5.
Dev Genes Evol ; 230(4): 305-314, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32671457

RESUMO

Crinoids are considered as the most basal extant echinoderms. They retain aboral nervous system with a nerve center, which has been degraded in the eleutherozoan echinoderms. To investigate the evolution of patterning of the nervous systems in crinoids, we examined temporal and spatial expression patterns of three neural patterning-related homeobox genes, six3, pax6, and otx, throughout the development of a feather star Anneissia japonica. These genes were involved in the patterning of endomesodermal tissues instead of the ectodermal neural tissues in the early planktonic stages. In the stages after larval attachment, the expression of these genes was mainly observed in the podia and the oral nervous systems instead of the aboral nerve center. Our results indicate the involvement of these three genes in the formation of oral nervous system in the common ancestor of the echinoderms and suggest that the aboral nerve center is not evolutionally related to the brain of other bilaterians.


Assuntos
Equinodermos/crescimento & desenvolvimento , Proteínas do Olho/metabolismo , Proteínas de Homeodomínio/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Fatores de Transcrição Otx/metabolismo , Fator de Transcrição PAX6/metabolismo , Animais , Padronização Corporal/genética , Equinodermos/genética , Equinodermos/metabolismo , Evolução Molecular , Proteínas do Olho/genética , Regulação da Expressão Gênica no Desenvolvimento/genética , Proteínas de Homeodomínio/genética , Larva/genética , Larva/metabolismo , Proteínas do Tecido Nervoso/genética , Sistema Nervoso/crescimento & desenvolvimento , Sistema Nervoso/metabolismo , Neurônios , Fatores de Transcrição Otx/genética , Fator de Transcrição PAX6/genética
6.
Proc Natl Acad Sci U S A ; 117(28): 16313-16323, 2020 07 14.
Artigo em Inglês | MEDLINE | ID: mdl-32601209

RESUMO

Peroxiredoxins are central to cellular redox homeostasis and signaling. They serve as peroxide scavengers, sensors, signal transducers, and chaperones, depending on conditions and context. Typical 2-Cys peroxiredoxins are known to switch between different oligomeric states, depending on redox state, pH, posttranslational modifications, and other factors. Quaternary states and their changes are closely connected to peroxiredoxin activity and function but so far have been studied, almost exclusively, outside the context of the living cell. Here we introduce the use of homo-FRET (Förster resonance energy transfer between identical fluorophores) fluorescence polarization to monitor dynamic changes in peroxiredoxin quaternary structure inside the crowded environment of living cells. Using the approach, we confirm peroxide- and thioredoxin-related quaternary transitions to take place in cellulo and observe that the relationship between dimer-decamer transitions and intersubunit disulfide bond formation is more complex than previously thought. Furthermore, we demonstrate the use of the approach to compare different peroxiredoxin isoforms and to identify mutations and small molecules affecting the oligomeric state inside cells. Mutagenesis experiments reveal that the dimer-decamer equilibrium is delicately balanced and can be shifted by single-atom structural changes. We show how to use this insight to improve the design of peroxiredoxin-based redox biosensors.


Assuntos
Peroxirredoxinas/química , Linhagem Celular , Transferência Ressonante de Energia de Fluorescência , Proteínas de Homeodomínio/química , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Humanos , Peróxido de Hidrogênio/metabolismo , Peróxido de Hidrogênio/farmacologia , Proteínas Luminescentes/química , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Mutação , Peroxirredoxinas/genética , Peroxirredoxinas/metabolismo , Multimerização Proteica/efeitos dos fármacos , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo
7.
Plant Mol Biol ; 104(3): 249-261, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32715397

RESUMO

Secondary cell wall not only provides rigidity and mechanical resistance to plants, but also has a large impact on plant growth and adaptation to environments. Biosynthesis of secondary cell wall is regulated by a complicated signaling transduction network; however, it is still unclear how the transcriptional regulation of secondary cell wall biosynthesis works at the molecular level. Here, we report in rice that OVATE family proteins 6 (OsOFP6) is a positive regulator in modulating expression of the genes related to biosynthesis of the secondary cell wall. Transgenic plants with knock-down of OsOFP6 by RNA interference showed increased leaf angle, which resulted from the thinner secondary cell wall with reduced amounts of cellulose and lignin, whilst overexpression of OsOFP6 in rice led to the thinker secondary cell wall with increased lignin content. Protein-protein interaction analysis revealed that OsOFP6 interacts with Oryza sativa homeobox 15 (OSH15), a class I KNOX protein. The interaction of OsOFP6 and OSH15 enhanced the transcriptional activity of OSH15 which binds to the promoter of OsIRX9 (Oryza sativa IRREGULAR XYLEM 9). Taken together, our study provides insights into the function of OsOFP6 in regulating leaf angle and the control of biosynthesis of secondary cell wall.


Assuntos
Parede Celular/metabolismo , Oryza/metabolismo , Folhas de Planta/metabolismo , Proteínas de Plantas/metabolismo , Proteínas Repressoras/metabolismo , Fatores de Transcrição/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Parede Celular/ultraestrutura , Celulose/metabolismo , Regulação da Expressão Gênica de Plantas , Proteínas de Homeodomínio/metabolismo , Lignina/metabolismo , Oryza/genética , Folhas de Planta/anatomia & histologia , Folhas de Planta/citologia , Proteínas de Plantas/genética , Plantas Geneticamente Modificadas/genética , Regiões Promotoras Genéticas , Domínios e Motivos de Interação entre Proteínas , Interferência de RNA , Proteínas Repressoras/genética , Fatores de Transcrição/genética , Xilema/metabolismo
8.
Nat Commun ; 11(1): 3339, 2020 07 03.
Artigo em Inglês | MEDLINE | ID: mdl-32620764

RESUMO

Chromosomal NUP98-PHF23 translocation is associated with an aggressive form of acute myeloid leukemia (AML) and poor survival rate. Here, we report the molecular mechanisms by which NUP98-PHF23 recognizes the histone mark H3K4me3 and is inhibited by small molecule compounds, including disulfiram that directly targets the PHD finger of PHF23 (PHF23PHD). Our data support a critical role for the PHD fingers of NUP98-PHF23, and related NUP98-KDM5A and NUP98-BPTF fusions in driving leukemogenesis, and demonstrate that blocking this interaction in NUP98-PHF23 expressing AML cells leads to cell death through necrotic and late apoptosis pathways. An overlap of NUP98-KDM5A oncoprotein binding sites and H3K4me3-positive loci at the Hoxa/b gene clusters and Meis1 in ChIP-seq, together with NMR analysis of the H3K4me3-binding sites of the PHD fingers from PHF23, KDM5A and BPTF, suggests a common PHD finger-dependent mechanism that promotes leukemogenesis by this type of NUP98 fusions. Our findings highlight the direct correlation between the abilities of NUP98-PHD finger fusion chimeras to associate with H3K4me3-enriched chromatin and leukemic transformation.


Assuntos
Cromatina/metabolismo , Proteínas de Homeodomínio/metabolismo , Leucemia Mieloide/metabolismo , Complexo de Proteínas Formadoras de Poros Nucleares/metabolismo , Proteínas de Fusão Oncogênica/metabolismo , Doença Aguda , Antígenos Nucleares/genética , Antígenos Nucleares/metabolismo , Cromatina/genética , Dissulfiram/farmacologia , Histonas/metabolismo , Proteínas de Homeodomínio/genética , Humanos , Leucemia Mieloide/genética , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Complexo de Proteínas Formadoras de Poros Nucleares/genética , Proteínas de Fusão Oncogênica/genética , Dedos de Zinco PHD/genética , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Proteína 2 de Ligação ao Retinoblastoma/genética , Proteína 2 de Ligação ao Retinoblastoma/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Translocação Genética/efeitos dos fármacos , Translocação Genética/genética
9.
Proc Natl Acad Sci U S A ; 117(28): 16509-16515, 2020 07 14.
Artigo em Inglês | MEDLINE | ID: mdl-32601200

RESUMO

Facioscapulohumeral muscular dystrophy (FSHD), characterized by progressive muscle weakness and deterioration, is genetically linked to aberrant expression of DUX4 in muscle. DUX4, in its full-length form, is cytotoxic in nongermline tissues. Here, we designed locked nucleic acid (LNA) gapmer antisense oligonucleotides (AOs) to knock down DUX4 in immortalized FSHD myoblasts and the FLExDUX4 FSHD mouse model. Using a screening method capable of reliably evaluating the knockdown efficiency of LNA gapmers against endogenous DUX4 messenger RNA in vitro, we demonstrate that several designed LNA gapmers selectively and effectively reduced DUX4 expression with nearly complete knockdown. We also found potential functional benefits of AOs on muscle fusion and structure in vitro. Finally, we show that one of the LNA gapmers was taken up and induced effective silencing of DUX4 upon local treatment in vivo. The LNA gapmers developed here will help facilitate the development of FSHD therapies.


Assuntos
Terapia Genética , Proteínas de Homeodomínio/genética , Distrofia Muscular Facioescapuloumeral/terapia , Oligonucleotídeos Antissenso/administração & dosagem , Animais , Modelos Animais de Doenças , Técnicas de Silenciamento de Genes , Proteínas de Homeodomínio/metabolismo , Humanos , Camundongos , Distrofia Muscular Facioescapuloumeral/genética , Distrofia Muscular Facioescapuloumeral/metabolismo , Mioblastos/metabolismo , Oligonucleotídeos Antissenso/genética , Oligonucleotídeos Antissenso/metabolismo
10.
Life Sci ; 258: 118145, 2020 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-32717270

RESUMO

AIMS: Studies on diabetes mellitus have shown that the phosphoinositide 3-kinase (PI3K)/serine threonine kinase (AKT)/forkhead box protein O1 (FoxO1) signaling pathway can regulate insulin secretion by modulating the expression of pancreatic and duodenal homeobox-1 (PDX-1). Therefore, it was hypothesized that the pathway also played an important role in functional abnormalities of pancreatic islets after severe burns. This study aimed to explore the role and mechanism of the PI3K/AKT/FoxO1/PDX-1 signaling pathway in functional changes of pancreatic islets in rats post severe burns. MAIN METHODS: Rats were grouped, subjected to full thickness burn injuries involving 50% total body surface area (TBSA), and injected intraperitoneally with BPV (HOpic) (0.6 mg/kg) or DMSO (0.55 mg/kg) once a day for three days. Glucose metabolism related indexes were measured by the glucometer, transmission electron microscopy (TEM) and enzyme-linked immunosorbent assay (ELISA). PI3K/AKT/FoxO1/PDX-1 signaling pathway related indexes were detected through immunofluorescence, western blot and RT-qPCR analyses. KEY FINDINGS: Dysglycemia and impaired insulin secretion occurred in rats, the activity of the PI3K/AKT/FoxO1/PDX-1 signaling pathway in the islets fell, and PDX-1 was translocated from the nucleus to the cytoplasm post severe burns. When BPV (HOpic) was used, glucose metabolism and insulin secretion were improved, the activity of the PI3K/AKT/FoxO1/PDX-1 signaling pathway in the islets was up-regulated, and PDX-1 was redistributed from the cytoplasm to the nucleus. SIGNIFICANCE: The activity of the PI3K/AKT/FoxO1/PDX-1 signaling pathway declined following severe burns. When its activity was up-regulated, insulin secretion could be improved, thus ameliorating hyperglycemia.


Assuntos
Queimaduras/patologia , Proteínas de Homeodomínio/metabolismo , Ilhotas Pancreáticas/patologia , Proteínas do Tecido Nervoso/metabolismo , Fosfatidilinositol 3-Quinase/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transativadores/metabolismo , Animais , Queimaduras/metabolismo , Secreção de Insulina , Ilhotas Pancreáticas/metabolismo , Masculino , Ratos , Ratos Wistar , Transdução de Sinais
11.
Int J Nanomedicine ; 15: 4483-4500, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32606690

RESUMO

Purpose: Tumor metastasis and drug resistance have always been vital aspects to cancer mortality and prognosis. To compromise metastasis and drug resistance, a nanoparticle IPPD-PHF2 (IR780/PLGA-PEI(Dox)-PHF2) has been engineered to accomplish efficient targeted epigenotherapy forced by PHF2-induced MET (mesenchymal to epithelial transition). Materials and Methods: IPPD-PHF2 nanoparticle was synthesized and characterized by several analytical techniques. The transfection efficiency of IPP-PHF2 (IR780/PLGA-PEI-PHF2) was compared with PP-PHF2 (PLGA-PEI-PHF2) in vitro by WB and in vivo by IHC, and the cytotoxicity of IPP was compared with Lipo2000 in vitro by CCK8 assay. The inhibition of cancer cell migration caused by PHF2-upregulation was tested by wound healing assay, and the enhanced chemotherapeutic sensitivity was detected by flow cytometry. Tumor-targeting property of IPPD-PHF2 was proved by fluorescent imaging in vivo with MDA-MB-231 tumor-bearing nude mice. Except for fluorescent imaging ability, considerable photoacoustic signals of IPPD-PHF2 at tumor sites were verified. The anti-tumor activity of IPPD-PHF2 was investigated using in vivo human breast cancer MDA-MB-231 cell models. Results: Tumor-targeting nanoparticle IPPD-PHF2 had an average size of about 319.2 nm, a stable zeta potential at about 38 mV. The encapsulation efficiency of doxorubicin was around 39.28%, and the adsorption capacity of plasmids was about 64.804 µg/mg. Significant up-regulation of PHF2 induced MET and caused reduced migration as well as enhanced chemotherapeutic sensitivity. Either IPPD (IR780/PLGA-PEI(Dox)) or IPP-PHF2 (IR780/PLGA-PEI-PHF2) presented minor therapeutic effects, whereas IPPD-PHF2 specifically accumulated within tumors, showed extraordinary transfection efficiency specifically in tumor sites, acted as inhibitors of metastasis and proliferation, and presented good multimodality imaging potentials in vivo. Conclusion: IPPD-PHF2 NPs is a promising tool to bring epigenotherapy into a more practical era, and the potential application of harm-free multimodality imaging guidance is of great value.


Assuntos
Antineoplásicos/uso terapêutico , Epigênese Genética , Nanopartículas/química , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Transfecção , Animais , Linhagem Celular Tumoral , Doxorrubicina/farmacologia , Doxorrubicina/uso terapêutico , Feminino , Proteínas de Homeodomínio/metabolismo , Humanos , Indóis/química , Camundongos Nus , Nanopartículas/ultraestrutura , Metástase Neoplásica , Técnicas Fotoacústicas , Polietilenoimina/química , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/química
12.
Gene ; 757: 144945, 2020 Oct 05.
Artigo em Inglês | MEDLINE | ID: mdl-32649979

RESUMO

HOX genes are important regulatory genes patterning head formation, including development of the ear. Microtia is a congenital ear anomaly characterized by lacking all or part of the structures of the outer ear. To date, only four HOXA2 mutations were reported in families with autosomal-recessive or dominant microtia, with or without hearing impairment. More identified mutations are needed to confirm the correlation between genotype and phenotype. Here, we collect two Chinese families with non-syndromic bilateral microtia. Next generation sequencing identified two heterozygous nonsense HOXA2 mutations, one in each family. One mutation (c.637A > T, p.Lys213*) is newly reported, while the other one (c.703C > T,p.Gln235*) is consistent with a previous report. In mouse, Hoxa2 can bind to a long-range enhancer and regulate expression of the Hmx1 gene, which is a crucial transcription factor in eye and ear development. Using dual luciferase reporter assays, we showed that both HOXA2 mutations have impaired activation of the long-range enhancer of HMX1. In the present study, we report the first two bilateral non-syndromic microtia cases with HOXA2 mutations of Chinese origin and identify a novel mutation. Our results also provide molecular insights about how these nonsense HOXA2 mutations could affect the activation of its downstream target HMX1 by interacting with the long-range enhancer.


Assuntos
Microtia Congênita/genética , Proteínas de Homeodomínio/genética , Mutação com Perda de Função , Células Cultivadas , Microtia Congênita/patologia , Feminino , Genes Dominantes , Células HEK293 , Proteínas de Homeodomínio/metabolismo , Humanos , Masculino , Linhagem , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
13.
Cell Prolif ; 53(8): e12835, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32557953

RESUMO

OBJECTIVES: HOXD3 is associated with progression of multiple types of cancer. This study aimed to identify the association of YY1 with HOXD3-ITGA2 axis in the progression of hepatocellular carcinoma. MATERIALS AND METHODS: Bioinformatics assay was used to identify the effect of YY1, HOXD3 and ITGA2 expression in HCC tissues. The function of YY1 and HOXD3 in HCCs was determined by qRT-PCR, MTT, apoptosis, Western blotting, colony formation, immunohistochemistry, and wound-healing and transwell invasion assays. The relationship between YY1 and HOXD3 or HOXD3 and ITGA2 was explored by RNA-Seq, ChIP-PCR, dual luciferase reports and Pearson's assays. The interactions between YY1 and HDAC1 were determined by immunofluorescence microscopy and Co-IP. RESULTS: Herein, we showed that the expression of YY1, HOXD3 and ITGA2 associated with the histologic and pathologic stages of HCC. Moreover, YY1, recruiting HDAC1, can directly target HOXD3 to regulate progression of HCCs. The relationship between YY1 and HOXD3 was unknown until uncovered by our present investigation. Furthermore, HOXD3 bound to promoter region of ITGA2 and up-regulated the expression, thus activating the ERK1/2 signalling and inducing HCCs proliferation, metastasis and migration in the vitro and vivo. CONCLUSIONS: Therefore, HOXD3, a target of YY1, facilitates HCC progression via activation of the ERK1/2 signalling by promoting ITGA2. This finding provides a new whole way to HCC therapy by serving YY1-HOXD3-ITGA2 regulatory axis as a potential therapeutic target for HCC therapy.


Assuntos
Carcinoma Hepatocelular/genética , Regulação Neoplásica da Expressão Gênica/genética , Histona Desacetilase 1/metabolismo , Proteínas de Homeodomínio/metabolismo , Neoplasias Hepáticas/genética , Fatores de Transcrição/metabolismo , Fator de Transcrição YY1/metabolismo , Apoptose/fisiologia , Movimento Celular/fisiologia , Proliferação de Células/fisiologia , Histona Desacetilase 1/genética , Humanos , RNA Longo não Codificante/metabolismo
14.
Life Sci ; 256: 117969, 2020 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-32553926

RESUMO

AIMS: Cannabinoids are the chemical compounds with a high affinity for cannabinoid receptors affecting the central nervous system through the release of neurotransmitters. However, the current knowledge related to the role of such compounds in the regulation of cellular aging is limited. This study aimed to investigate the effect of cannabidiol and tetrahydrocannabinol on the function of aged pancreatic islets. MAIN METHODS: The expression of p53, p38, p21, p16, and Glut2 genes and ß-galactosidase activity were measured as hallmarks of cell aging applying real-time PCR, ELISA, and immunocytochemistry techniques. Pdx1 protein expression, insulin release, and oxidative stress markers were compared between young and aged rat pancreatic islet cells. KEY FINDINGS: Upon the treatment of aged pancreatic islets cells with cannabidiol and tetrahydrocannabinol, the expression of p53, p38, p21 and the activity of ß-galactosidase were reduced. Cannabidiol and tetrahydrocannabinol increase insulin release, Pdx1, Glut2, and thiol molecules expression, while the oxidative stress parameters were decreased. The enhanced expression of Pdx1 and insulin release in aged pancreatic islet cells reflects the extension of cell healthy aging due to the significant reduction of ROS. SIGNIFICANCE: This study provides evidence for the involvement of cannabidiol and tetrahydrocannabinol in the oxidation process of cellular aging.


Assuntos
Canabinoides/farmacologia , Senescência Celular , Ilhotas Pancreáticas/citologia , Espécies Reativas de Oxigênio/metabolismo , Animais , Biomarcadores/metabolismo , Canabidiol/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Senescência Celular/efeitos dos fármacos , Dronabinol/farmacologia , Proteínas de Homeodomínio/metabolismo , Secreção de Insulina/efeitos dos fármacos , Masculino , Estresse Oxidativo/efeitos dos fármacos , Ratos Wistar , Transativadores/metabolismo
15.
Anat Sci Int ; 95(4): 523-539, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32476103

RESUMO

Type 1 diabetes mellitus (T1DM) is a chronic metabolic disease caused by the destruction of pancreatic ß-cells. Human dental pulp stem cells represent a promising source for cell-based therapies, owing to their easy, minimally invasive surgical access, and high proliferative capacity. It was reported that human dental pulp stem cells can differentiate into a pancreatic cell lineage in vitro; however, few studies have investigated their effects on diabetes. Our study aimed to investigate the therapeutic potential of intravenous and intrapancreatic transplantation of human dental pulp stem cells in a rat model of streptozotocin-induced type 1 diabetes. Forty Sprague Dawley male rats were randomly categorized into four groups: control, diabetic (STZ), intravenous treatment group (IV), and intrapancreatic treatment group (IP). Human dental pulp stem cells (1 × 106 cells) or vehicle were injected into the pancreas or tail vein 7 days after streptozotocin injection. Fasting blood glucose levels were monitored weekly. Glucose tolerance test, rat and human serum insulin and C-peptide, pancreas histology, and caspase-3, vascular endothelial growth factor, and Ki67 expression in pancreatic tissues were assessed 28 days post-transplantation. We found that both IV and IP transplantation of human dental pulp stem cells reduced blood glucose and increased levels of rat and human serum insulin and C-peptide. The cells engrafted and survived in the streptozotocin-injured pancreas. Islet-like clusters and scattered human dental pulp stem cells expressing insulin were observed in the pancreas of diabetic rats with some difference in the distribution pattern between the two injection routes. RT-PCR analyses revealed the expression of the human-specific pancreatic ß-cell genes neurogenin 3 (NGN3), paired box 4 (PAX4), glucose transporter 2 (GLUT2), and insulin in the pancreatic tissues of both the IP and IV groups. In addition, the transplanted cells downregulated the expression of caspase-3 and upregulated the expression of vascular endothelial growth factor and Ki67, suggesting that the injected cells exerted pro-angiogenetic and antiapoptotic effects, and promoted endogenous ß-cell replication. Our study is the first to show that human dental pulp stem cells can migrate and survive within streptozotocin-injured pancreas, and induce antidiabetic effects through the differentiation and replacement of lost ß-cells and paracrine-mediated pancreatic regeneration. Thus, human dental pulp stem cells may have therapeutic potential to treat patients with long term T1DM.


Assuntos
Polpa Dentária/citologia , Diabetes Mellitus Experimental/terapia , Diabetes Mellitus Tipo 1/terapia , Pâncreas/fisiologia , Transplante de Células-Tronco , Células-Tronco/fisiologia , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Caspase 3/metabolismo , Diferenciação Celular , Movimento Celular , Sobrevivência Celular , Modelos Animais de Doenças , Transportador de Glucose Tipo 2/metabolismo , Proteínas de Homeodomínio/metabolismo , Humanos , Antígeno Ki-67/metabolismo , Masculino , Proteínas do Tecido Nervoso/metabolismo , Fatores de Transcrição Box Pareados/metabolismo , Pâncreas/citologia , Pâncreas/metabolismo , Ratos , Ratos Sprague-Dawley , Regeneração , Estreptozocina
16.
PLoS One ; 15(6): e0229549, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32497078

RESUMO

Dysfunctions in the GABAergic system lead to various pathological conditions and impaired inhibitory function is one of the causes behind neuropathies characterized by neuronal hyper excitability. The Dlx homeobox genes are involved in the development of nervous system, neural crest, branchial arches and developing appendages. Dlx genes also take part in neuronal migration and differentiation during development, more precisely, in the migration and differentiation of GABAergic neurons. Functional analysis of dlx genes has mainly been carried out in developing zebrafish embryos and larvae, however information regarding the expression and roles of these genes in the adult zebrafish brain is still lacking. The extensive neurogenesis that takes place in the adult zebrafish brain, makes them a good model for the visualization of mechanisms involving dlx genes during adulthood in physiological conditions and during regeneration of the nervous system. We have identified the adult brain regions where transcripts of dlx1a, dlx2a, dlx5a and dlx6a genes are normally found and have confirmed that within telencephalic domains, there is high overlapping expression of the four dlx paralogs with a marker for GABAergic neurons. Co-localization analyses carried with the Tg(dlx6a-1.4kbdlx5a/dlx6a:GFP) reporter line have also shown that in some areas of the diencephalon, cells expressing the dlx5a/6a bigene may have a neural stem cell identity. Furthermore, investigations in a response to stab wound lesions, have demonstrated a possible participation of the dlx5a/6a bigene, most likely of dlx5a, during regeneration of the adult zebrafish brain. These observations suggest a possible participation of dlx-expressing cells during brain regeneration in adult zebrafish and also provide information on the role of dlx genes under normal physiological conditions in adults.


Assuntos
Encéfalo/fisiologia , Regulação da Expressão Gênica , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Regeneração , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Peixe-Zebra/genética , Peixe-Zebra/fisiologia , Animais , Encéfalo/citologia , Encéfalo/metabolismo , Células-Tronco Neurais/metabolismo
17.
Nat Commun ; 11(1): 2984, 2020 06 12.
Artigo em Inglês | MEDLINE | ID: mdl-32533114

RESUMO

ADNP (Activity Dependent Neuroprotective Protein) is a neuroprotective protein whose aberrant expression has been frequently linked to neural developmental disorders, including the Helsmoortel-Van der Aa syndrome (also called the ADNP syndrome). However, its role in neural development and pathology remains unclear. Here, we show that ADNP is required for neural induction and differentiation by enhancing Wnt signaling. Mechanistically, ADNP functions to stabilize ß-Catenin through binding to its armadillo domain which prevents its association with key components of the degradation complex: Axin and APC. Loss of ADNP promotes the formation of the degradation complex and ß-Catenin degradation via ubiquitin-proteasome pathway, resulting in down-regulation of key neuroectoderm developmental genes. In addition, adnp gene disruption in zebrafish leads to defective neurogenesis and reduced Wnt signaling. Our work provides important insights into the role of ADNP in neural development and the pathology of the Helsmoortel-Van der Aa syndrome caused by ADNP gene mutation.


Assuntos
Diferenciação Celular/genética , Proteínas de Homeodomínio/genética , Células-Tronco Embrionárias Murinas/metabolismo , Proteínas do Tecido Nervoso/genética , Neurônios/metabolismo , Via de Sinalização Wnt/genética , beta Catenina/genética , Animais , Células Cultivadas , Regulação da Expressão Gênica no Desenvolvimento , Células HEK293 , Proteínas de Homeodomínio/metabolismo , Humanos , Hibridização In Situ/métodos , Camundongos , Camundongos Knockout , Células-Tronco Embrionárias Murinas/citologia , Mutação , Proteínas do Tecido Nervoso/metabolismo , Neurônios/citologia , Ligação Proteica , Peixe-Zebra/embriologia , Peixe-Zebra/genética , Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismo , beta Catenina/metabolismo
18.
Nature ; 582(7811): 271-276, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32499640

RESUMO

A major factor in the progression to heart failure in humans is the inability of the adult heart to repair itself after injury. We recently demonstrated that the early postnatal mammalian heart is capable of regeneration following injury through proliferation of preexisting cardiomyocytes1,2 and that Meis1, a three amino acid loop extension (TALE) family homeodomain transcription factor, translocates to cardiomyocyte nuclei shortly after birth and mediates postnatal cell cycle arrest3. Here we report that Hoxb13 acts as a cofactor of Meis1 in postnatal cardiomyocytes. Cardiomyocyte-specific deletion of Hoxb13 can extend the postnatal window of cardiomyocyte proliferation and reactivate the cardiomyocyte cell cycle in the adult heart. Moreover, adult Meis1-Hoxb13 double-knockout hearts display widespread cardiomyocyte mitosis, sarcomere disassembly and improved left ventricular systolic function following myocardial infarction, as demonstrated by echocardiography and magnetic resonance imaging. Chromatin immunoprecipitation with sequencing demonstrates that Meis1 and Hoxb13 act cooperatively to regulate cardiomyocyte maturation and cell cycle. Finally, we show that the calcium-activated protein phosphatase calcineurin dephosphorylates Hoxb13 at serine-204, resulting in its nuclear localization and cell cycle arrest. These results demonstrate that Meis1 and Hoxb13 act cooperatively to regulate cardiomyocyte maturation and proliferation and provide mechanistic insights into the link between hyperplastic and hypertrophic growth of cardiomyocytes.


Assuntos
Calcineurina/metabolismo , Proliferação de Células , Proteínas de Homeodomínio/metabolismo , Proteína Meis1/metabolismo , Miócitos Cardíacos/citologia , Animais , Animais Recém-Nascidos , Feminino , Deleção de Genes , Regulação da Expressão Gênica , Coração/fisiologia , Proteínas de Homeodomínio/genética , Masculino , Camundongos , Miocárdio/citologia , Ligação Proteica , Regeneração
19.
Mol Cell ; 78(6): 1114-1132.e10, 2020 06 18.
Artigo em Inglês | MEDLINE | ID: mdl-32446320

RESUMO

Bromodomain-containing protein 4 (BRD4) is a cancer therapeutic target in ongoing clinical trials disrupting primarily BRD4-regulated transcription programs. The role of BRD4 in cancer has been attributed mainly to the abundant long isoform (BRD4-L). Here we show, by isoform-specific knockdown and endogenous protein detection, along with transgene expression, the less abundant BRD4 short isoform (BRD4-S) is oncogenic while BRD4-L is tumor-suppressive in breast cancer cell proliferation and migration, as well as mammary tumor formation and metastasis. Through integrated RNA-seq, genome-wide ChIP-seq, and CUT&RUN association profiling, we identify the Engrailed-1 (EN1) homeobox transcription factor as a key BRD4-S coregulator, particularly in triple-negative breast cancer. BRD4-S and EN1 comodulate the extracellular matrix (ECM)-associated matrisome network, including type II cystatin gene cluster, mucin 5, and cathepsin loci, via enhancer regulation of cancer-associated genes and pathways. Our work highlights the importance of targeted therapies for the oncogenic, but not tumor-suppressive, activity of BRD4.


Assuntos
Neoplasias da Mama/metabolismo , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ciclo Celular/fisiologia , Fatores de Transcrição/metabolismo , Fatores de Transcrição/fisiologia , Animais , Neoplasias da Mama/genética , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Genes Homeobox , Proteínas de Homeodomínio/metabolismo , Humanos , Camundongos , Invasividade Neoplásica , Proteínas Nucleares/metabolismo , Isoformas de Proteínas/metabolismo , Proteínas/antagonistas & inibidores , Proteínas/metabolismo , Transcrição Genética/genética , Neoplasias de Mama Triplo Negativas/genética
20.
Nat Commun ; 11(1): 2380, 2020 05 13.
Artigo em Inglês | MEDLINE | ID: mdl-32404936

RESUMO

YAP1 gene fusions have been observed in a subset of paediatric ependymomas. Here we show that, ectopic expression of active nuclear YAP1 (nlsYAP5SA) in ventricular zone neural progenitor cells using conditionally-induced NEX/NeuroD6-Cre is sufficient to drive brain tumour formation in mice. Neuronal differentiation is inhibited in the hippocampus. Deletion of YAP1's negative regulators LATS1 and LATS2 kinases in NEX-Cre lineage in double conditional knockout mice also generates similar tumours, which are rescued by deletion of YAP1 and its paralog TAZ. YAP1/TAZ-induced mouse tumours display molecular and ultrastructural characteristics of human ependymoma. RNA sequencing and quantitative proteomics of mouse tumours demonstrate similarities to YAP1-fusion induced supratentorial ependymoma. Finally, we find that transcriptional cofactor HOPX is upregulated in mouse models and in human YAP1-fusion induced ependymoma, supporting their similarity. Our results show that uncontrolled YAP1/TAZ activity in neuronal precursor cells leads to ependymoma-like tumours in mice.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Neoplasias Encefálicas/metabolismo , Proteínas de Ciclo Celular/metabolismo , Ependimoma/metabolismo , Transativadores/metabolismo , Fatores de Transcrição/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Adulto , Animais , Encéfalo/metabolismo , Encéfalo/patologia , Encéfalo/ultraestrutura , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patologia , Proteínas de Ciclo Celular/genética , Criança , Ependimoma/genética , Ependimoma/patologia , Regulação Neoplásica da Expressão Gênica , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Humanos , Camundongos Knockout , Camundongos Transgênicos , Microscopia Eletrônica de Varredura , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Transativadores/genética , Fatores de Transcrição/genética , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismo
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