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1.
Proc Natl Acad Sci U S A ; 117(38): 23952-23959, 2020 09 22.
Artigo em Inglês | MEDLINE | ID: mdl-32900950

RESUMO

Glands of the uterus are essential for pregnancy establishment. Forkhead box A2 (FOXA2) is expressed specifically in the glands of the uterus and a critical regulator of glandular epithelium (GE) differentiation, development, and function. Mice with a conditional deletion of FOXA2 in the adult uterus, created using the lactotransferrin iCre (Ltf-iCre) model, have a morphologically normal uterus with glands, but lack FOXA2-dependent GE-expressed genes, such as leukemia inhibitory factor (LIF). Adult FOXA2 conditional knockout (cKO; Ltf iCre/+ Foxa2 f/f ) mice are infertile due to defective embryo implantation arising from a lack of LIF, a critical implantation factor of uterine gland origin. However, intraperitoneal injections of LIF can initiate embryo implantation in the uterus of adult FOXA2 cKO mice with pregnancies maintained to term. Here, we tested the hypothesis that FOXA2-regulated genes in the uterine glands impact development of the decidua, placenta, and fetus. On gestational day 8.5, the antimesometrial and mesometrial decidua transcriptome was noticeably altered in LIF-replaced FOXA2 cKO mice. Viable fetuses were reduced in FOXA2 cKO mice on gestational days 12.5 and 17.5. Sex-dependent differences in fetal weight, placenta histoarchitecture, and the placenta and metrial gland transcriptome were observed between control and FOXA2 cKO mice. The transcriptome of the placenta with a female fetus was considerably more altered than the placenta with a male fetus in FOXA2 cKO dams. These studies reveal previously unrecognized sexually dimorphic effects of FOXA2 and uterine glands on fetoplacental development with potential impacts on offspring health into adulthood.


Assuntos
Feto/metabolismo , Fator 3-beta Nuclear de Hepatócito , Placenta/metabolismo , Caracteres Sexuais , Útero/metabolismo , Animais , Decídua/metabolismo , Feminino , Fator 3-beta Nuclear de Hepatócito/genética , Fator 3-beta Nuclear de Hepatócito/metabolismo , Masculino , Camundongos , Camundongos Knockout , Gravidez , Transcriptoma/genética
2.
Oral Dis ; 26(4): 756-765, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-31957176

RESUMO

OBJECTIVES: FOXA2 gene methylation links to the progression of cancers, but has not been documented in oral cancer. Herein, we explore the role of FOXA2 in the migration of oral cancer cells. MATERIAL AND METHODS: Methylation-specific PCR was applied for gene methylation. Wound healing and transwell experiments were tested for cell migration. FOXA2 expression in oral cancer tissues was addressed by immunohistochemistry, followed by statistical analysis of its association with clinical manifestations and patient survival. RESULTS: FOXA2 bound to the promoter of CDH1 and enhanced the expression of its gene product E-cadherin, and decreased the cancer cell migration activity. High FOXA2 expression in oral cancer tissues was associated with high E-cadherin expression, decreased lymph node metastasis, and increased patient survival. CONCLUSION: FOXA2-E-cadherin link is involved in regulation of oral cancer cell metastasis and provides a new insight for the tumor suppressor activity of FOXA2 in oral cancer.


Assuntos
Antígenos CD/genética , Caderinas/genética , Fator 3-beta Nuclear de Hepatócito/genética , Metástase Linfática , Neoplasias Bucais/genética , Movimento Celular , Metilação de DNA , Inativação Gênica , Humanos , Linfonodos/patologia , Neoplasias Bucais/patologia , Regiões Promotoras Genéticas
3.
Nat Commun ; 11(1): 342, 2020 01 17.
Artigo em Inglês | MEDLINE | ID: mdl-31953394

RESUMO

Precise control of hepatic glucose production (HGP) is pivotal to maintain systemic glucose homeostasis. HNF4α functions to stimulate transcription of key gluconeogenic genes. HNF4α harbors two promoters (P2 and P1) thought to be primarily active in fetal and adult livers, respectively. Here we report that the fetal version of HNF4α is required for HGP in the adult liver. This isoform is acutely induced upon fasting and chronically increased in type-2 diabetes (T2D). P2 isoform induction occurs in response to glucagon-stimulated upregulation of TET3, not previously shown to be involved in HGP. TET3 is recruited to the P2 promoter by FOXA2, leading to promoter demethylation and increased transcription. While TET3 overexpression augments HGP, knockdown of either TET3 or the P2 isoform alone in the liver improves glucose homeostasis in dietary and genetic mouse models of T2D. These studies unmask an unanticipated, conserved regulatory mechanism in HGP and offer potential therapeutic targets for T2D.


Assuntos
Diabetes Mellitus Tipo 2/metabolismo , Dioxigenases/metabolismo , Fator 4 Nuclear de Hepatócito/metabolismo , Fígado/metabolismo , Isoformas de Proteínas/metabolismo , Animais , Desmetilação do DNA , Metilação de DNA , Proteínas de Ligação a DNA/metabolismo , Dioxigenases/genética , Modelos Animais de Doenças , Jejum , Regulação da Expressão Gênica , Glucagon/metabolismo , Glucose/metabolismo , Fator 3-beta Nuclear de Hepatócito/genética , Fator 4 Nuclear de Hepatócito/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Regiões Promotoras Genéticas , Isoformas de Proteínas/genética , Ativação Transcricional , Transcriptoma , Regulação para Cima
4.
Life Sci ; 241: 117166, 2020 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-31843527

RESUMO

AIMS: Congenital diaphragmatic hernia (CDH) is a lethal birth defect characterized by congenital lung malformation, and the severity of pulmonary hypoplasia directly affects the prognosis of infants with CDH. Using a nitrofen-induced CDH rat model, we previously reported that Foxa2 expression was downregulated in CDH lungs by proteomics analysis. Here, we investigate the role of miR-130a-5p/Foxa2 axis in lung development of the nitrofen-induced CDH and evaluate its potential role in vivo prenatal therapy. MAIN METHODS: Nitrofen was orally administrated on embryonic day (E) 8.5 to establish a rat CDH model, and fetal lungs were collected on E13.5, E15.5, E17.5, E19.5 and E21.5. The binding sites of miR-130a-5p on Foxa2 mRNA were identified using bioinformatics prediction software and were validated via luciferase assay. The expression levels of miR-130a-5p and Foxa2 were detected using qRT-PCR, ISH, IHC and western blotting. The role of miR-130a-5p/Foxa2 axis in CDH-associated lung development was investigated in ex vivo lung explants. KEY FINDINGS: We found that Foxa2 was downregulated in CDH lung tissues, and Foxa2 upregulating improved CDH branching morphogenesis in ex vivo lung explants. Meanwhile, we also showed that miR-130a-5p was significantly upregulated in CDH lungs and thus inversely correlated with Foxa2. Increasing miR-130a-5p abundance with mimics decreases Foxa2-driven Shh/Gli1 signaling and inhibits branching morphogenesis in ex vivo lung explants. SIGNIFICANCE: This study was the first to show that the miR-130a-5p/Foxa2 axis played a crucial role in CDH-associated pulmonary hypoplasia. These findings may provide relevant insights into the prenatal diagnosis and prenatal therapy of CDH.


Assuntos
Desenvolvimento Fetal/genética , Regulação da Expressão Gênica no Desenvolvimento , Fator 3-beta Nuclear de Hepatócito/metabolismo , Hérnias Diafragmáticas Congênitas/patologia , Pulmão/citologia , MicroRNAs/genética , Organogênese , Animais , Proliferação de Células , Células Cultivadas , Feminino , Proteínas Hedgehog/genética , Proteínas Hedgehog/metabolismo , Fator 3-beta Nuclear de Hepatócito/genética , Herbicidas/toxicidade , Hérnias Diafragmáticas Congênitas/induzido quimicamente , Hérnias Diafragmáticas Congênitas/metabolismo , Pulmão/fisiologia , Masculino , Éteres Fenílicos/toxicidade , Gravidez , Ratos , Ratos Sprague-Dawley , Proteína GLI1 em Dedos de Zinco/genética , Proteína GLI1 em Dedos de Zinco/metabolismo
5.
PLoS One ; 14(12): e0226517, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31881038

RESUMO

Allergic asthma, characterized by chronic airway Th2-dominated inflammation, is associated with an increased risk of infection; however, the underlying mechanisms are unclear. Forkhead box protein A2 (Foxa2) plays a critical role in Th2 inflammation and is associated with pulmonary defenses. To determining the role of Foxa2 in Th2-dominated lung inflammation against the invading bacteria, we established a mouse OVA-sensitized model, an Escherichia coli lung invasion model, and mice with conditional deletion of Foxa2 in respiratory epithelial cells. The number of bacteria in the lung tissue was counted to assess clearance ability of lung. Lung inflammation and histopathology was evaluated using HE and PAS staining. It was found that OVA-sensitized mice had decreased E. coli clearance, reduced Foxa2 expression, and decreased DEFB1 secretion. Conditional deletion of Foxa2 in respiratory epithelial cells led to decreased clearance of E. coli and impaired secretion of DEFB1, similar to the OVA-induced allergic condition. The impaired secretion of DEFB1 may be responsible for the increased risk of infection in the Th2-dominated airway inflammation. Dual luciferase assay demonstrated that Foxa2 regulates DEFB1 expression by affecting its promoter activity in HBE cells. Our study indicated that Foxa2 plays an important role in Th2-dominated airway inflammation against invading bacteria. Conditional deletion of Foxa2 in respiratory epithelial cells can reduce pulmonary's defense against bacterial invasion by inhibiting DEFB1expression.


Assuntos
Asma/microbiologia , Infecções por Escherichia coli/prevenção & controle , Fator 3-beta Nuclear de Hepatócito/genética , beta-Defensinas/metabolismo , Animais , Asma/induzido quimicamente , Asma/genética , Asma/imunologia , Linhagem Celular , Modelos Animais de Doenças , Infecções por Escherichia coli/genética , Infecções por Escherichia coli/metabolismo , Feminino , Deleção de Genes , Regulação da Expressão Gênica , Humanos , Camundongos , Ovalbumina/efeitos adversos , Células Th2/metabolismo
6.
Anal Cell Pathol (Amst) ; 2019: 3234812, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31781476

RESUMO

miRNAs are small non-coding RNA sequences of 18-25 nucleotides. They can regulate different cellular pathways by acting on tumor suppressors, oncogenes, or both. miRNAs are mostly tissue-specific, and their expression varies depending on the cancer or the tissue in which they are found. hsa-miR-590-3p was found to be involved in several types of cancers. In this study, we identified potential downstream target genes of hsa-miR-590-3p computationally. Several bioinformatics tools and more than one approach were used to identify potential downstream target genes of hsa-miR-590-3p. CX3CL1, SOX2, N-cadherin, E-cadherin, and FOXA2 were utilized as potential downstream target genes of hsa-miR-590-3p. SNU449 and HepG2, hepatocellular carcinoma cell lines, were used to carry out various molecular techniques to further validate our in silico results. mRNA and protein expression levels of these genes were detected using RT-PCR and western blotting, respectively. Co-localization of hsa-miR-590-3p and its candidate downstream target gene, SOX2, was carried out using a miRNA in situ hybridization combined with immunohistochemistry staining through anti-SOX2. The results show that there is an inverse correlation between hsa-miR-590-3p expression and SOX2 protein expression in SNU449. Subsequently, we suggest that SOX2 can be a direct downstream target of has-miR-590-3p indicating that it may have a role in the self-renewal and self-maintenance of cancer cells. We also suggest that CX3CL1, E-cadherin, N-cadherin, and FOXA2 show a lot of potential as downstream target genes of hsa-miR-590-3p signifying its role in epithelial-mesenchymal transition. Studying the expression of hsa-miR-590-3p downstream targets can enrich our understanding of the cancer pathogenesis and how it can be used as a therapeutic tool.


Assuntos
Carcinoma Hepatocelular/genética , Genes Neoplásicos , Neoplasias Hepáticas/genética , MicroRNAs/genética , Sequência de Bases , Biomarcadores Tumorais/metabolismo , Caderinas/genética , Caderinas/metabolismo , Linhagem Celular Tumoral , Quimiocina CX3CL1/genética , Quimiocina CX3CL1/metabolismo , Regulação Neoplásica da Expressão Gênica , Fator 3-beta Nuclear de Hepatócito/genética , Fator 3-beta Nuclear de Hepatócito/metabolismo , Humanos , Mesoderma/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Fatores de Transcrição SOXB1/genética , Fatores de Transcrição SOXB1/metabolismo , Versicanas/genética , Versicanas/metabolismo , Vimentina/metabolismo
7.
Biosci Rep ; 39(11)2019 11 29.
Artigo em Inglês | MEDLINE | ID: mdl-31701999

RESUMO

MicroRNA (MiR)-942 regulates the development of a variety of tumors, however, its function in breast cancer (BCa) has been less reported. Therefore, the present study investigated the regulatory effects of miR-942 on BCa cells. The expression of miR-942 in whole blood samples and BCa cell lines was detected by quantitative real-time (qRT)-PCR. Direct target gene for miR-942 was confirmed by dual-luciferase reporter assay. FOXA2 expression in adjacent tissues was detected by qRT-PCR. The effects of miR-942, or miR-942 with FOXA2, on the cell viability, proliferation, apoptosis, migration and invasion of BCa cells were determined by cell counting kit-8 (CCK-8), colony formation assay, flow cytometry, wound scratch and Transwell, respectively. The levels of N-Cadherin, E-Cadherin and Snail were determined by Western blot. Kaplan-Meier was used to explore the relationship among the expressions of miR-942 and FOXA2 and the prognosis of BCa patients. MiR-942 had high expressed in BCa, while its low expression significantly suppressed the cell viability, proliferation, migration and invasion of BCa, but increased cell apoptosis. Down-regulation of N-Cadherin and Snail and up-regulation of E-Cadherin were also induced by low-expression of miR-942. FOXA2, which was proved as the direct target gene for miR-942 and was low-expressed in BCa, partially reversed the effect of overexpressed miR-942 on promoting cell viability, proliferation, migration and invasion, and suppressed cell apoptosis. A lower survival rate was observed in BCa patients with a high expression of miR-942 and a low expression of FOXA2. MiR-942 promoted the progression of BCa by down-regulating the expression of FOXA2.


Assuntos
Neoplasias da Mama/genética , Fator 3-beta Nuclear de Hepatócito/genética , MicroRNAs/genética , Apoptose/genética , Caderinas/genética , Estudos de Casos e Controles , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Sobrevivência Celular/genética , Células Cultivadas , Regulação para Baixo/genética , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Células MCF-7 , Masculino , Prognóstico , Regulação para Cima/genética
8.
Aging (Albany NY) ; 11(21): 9280-9294, 2019 11 05.
Artigo em Inglês | MEDLINE | ID: mdl-31689237

RESUMO

BACKGROUND: Intrahepatic cholangiocarcinoma (ICC) is characterized by a highly aggressive nature and a dismal outcome. FOXA2 is an archetypal transcription factor involved in cholangiocyte proliferation. RESULTS: FOXA2 expression was negatively correlated with tumor stage (p = 0.024). Univariate and multivariate analyses showed that low FoxA2 expression was associated with tumor relapse and survival. At 20 weeks after TAA administration, FoxA2-/- mice displayed significant manifestations of neoplasia, while WT mice did not.RNA sequencing analysis showed that the expression of genes in the MAPK signaling pathway was significantly higher in FoxA2-/- mice. IHC and Western blot results showed that p-ERK1/2, CREB1 and RAS were highly expressed in FoxA2-/- mice. Furthermore, using in vitro experiments with siRNA, we found that low expression of FoxA2 could exacerbate the metastatic potential of ICC. The expression of p-ERK1/2 and RAS, which are key mediators of the MAPK signaling pathway, was significantly increased. CONCLUSION: Low FOXA2 expression negatively affected the prognosis of patients with ICC. Loss of FoxA2 expression could promote intrahepatic bile duct neoplasia partly via activation of the MAPK signaling pathway. MATERIALS AND METHODS: In all, the data of 85 patients with ICC were retrospectively collected and analyzed. TAA was used to induce ICC in FoxA2-/- mice and WT mice. RNA-sequencing analysis was used to identify the expression of different genes.


Assuntos
Neoplasias dos Ductos Biliares/metabolismo , Colangiocarcinoma/metabolismo , Fator 3-beta Nuclear de Hepatócito/deficiência , Sistema de Sinalização das MAP Quinases , Adulto , Idoso , Animais , Neoplasias dos Ductos Biliares/genética , Neoplasias dos Ductos Biliares/patologia , Ductos Biliares Intra-Hepáticos/metabolismo , Ductos Biliares Intra-Hepáticos/patologia , Proliferação de Células , Transformação Celular Neoplásica , Colangiocarcinoma/genética , Colangiocarcinoma/patologia , Progressão da Doença , Feminino , Expressão Gênica , Fator 3-beta Nuclear de Hepatócito/genética , Humanos , Sistema de Sinalização das MAP Quinases/genética , Masculino , Camundongos , Camundongos Knockout , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/genética , Recidiva Local de Neoplasia/metabolismo , Recidiva Local de Neoplasia/patologia , Neoplasias Experimentais/genética , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/patologia , Prognóstico , Estudos Retrospectivos , Fatores de Risco , Tioacetamida/toxicidade
9.
Int J Biol Sci ; 15(12): 2561-2575, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31754329

RESUMO

Diabetes mellitus is characterized by pancreatic ß cell dysfunction. Previous studies have indicated that epidermal growth factor (EGF) and microRNA-124a (miR-124a) play opposite roles in insulin biosynthesis and secretion by beta cells. However, the underlying mechanisms remain poorly understood. In the present study, we demonstrated that EGF could inhibit miR-124a expression in beta cell lines through downstream signaling pathways, including mitogen-activated protein kinase kinase (MEK) and phosphatidylinositol 3-kinase (PI3K) cascades. Further, the transcription factor ETS2, a member of the ETS (E26 transformation-specific) family, was identified to be responsible for the EGF-mediated suppression of miR-124a expression, which was dependent on ETS2 phosphorylation at threonine 72. Activation of ETS2 decreased miR-124a promoter transcriptional activity through the putative conserved binding sites AGGAANA/TN in three miR-124a promoters located in different chromosomes. Of note, ETS2 played a positive role in regulating beta cell function-related genes, including miR-124a targets, Forkhead box a2 (FOXA2) and Neurogenic differentiation 1 (NEUROD1), which may have partly been through the inhibition of miR-124 expression. Knockdown and overexpression of ETS2 led to the prevention and promotion of insulin biosynthesis respectively, while barely affecting the secretion ability. These results suggest that EGF may induce the activation of ETS2 to inhibit miR-124a expression to maintain proper beta cell functions and that ETS2, as a novel regulator of insulin production, is a potential therapeutic target for diabetes mellitus treatment.


Assuntos
Fator de Crescimento Epidérmico/fisiologia , Células Secretoras de Insulina/metabolismo , MicroRNAs/metabolismo , Proteínas Quinases Ativadas por Mitógeno/fisiologia , Fosfatidilinositol 3-Quinase/fisiologia , Proteína Proto-Oncogênica c-ets-2/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Sítios de Ligação , Diabetes Mellitus/genética , Diabetes Mellitus/metabolismo , Fator de Crescimento Epidérmico/genética , Fator de Crescimento Epidérmico/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Fator 3-beta Nuclear de Hepatócito/genética , Fator 3-beta Nuclear de Hepatócito/metabolismo , Insulina/metabolismo , Ilhotas Pancreáticas/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , MicroRNAs/genética , Proteínas Quinases Ativadas por Mitógeno/genética , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Fosfatidilinositol 3-Quinase/genética , Fosfatidilinositol 3-Quinase/metabolismo , Fosforilação , Regiões Promotoras Genéticas , Transdução de Sinais , Treonina/metabolismo
10.
Development ; 146(20)2019 10 15.
Artigo em Inglês | MEDLINE | ID: mdl-31575644

RESUMO

During early embryogenesis, mechanical constraints and localized biochemical signals co-occur around anteroposterior axis determination and symmetry breaking. Their relative roles, however, are hard to tease apart in vivo Using brachyury (Bra), a primitive streak and mesendoderm marker in mouse embryoid bodies (EBs), we studied how contact, biochemical cues and neighboring cell cues affect the positioning of a primitive streak-like locus and thus determine the anteroposterior axis. We show that a Bra-competent layer must be formed in the EB before Bra expression initiates, and that Bra onset locus position is biased by contact points of the EB with its surrounding, probably through modulation of chemical cues rather than by mechanical signaling. We can push or pull Bra onset away from contact points by introducing a separate localized Wnt signal source, or maneuver Bra onset to a few loci or to an isotropic peripheral pattern. Furthermore, we show that Foxa2-positive cells are predictive of the future location of Bra onset, demonstrating an earlier symmetry-breaking event. Our analysis of factors affecting symmetry breaking and spatial fate choice during this developmental process could prove valuable for in vitro differentiation and organoid formation.


Assuntos
Corpos Embrioides/citologia , Corpos Embrioides/metabolismo , Animais , Diferenciação Celular/genética , Diferenciação Celular/fisiologia , Células Cultivadas , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/metabolismo , Proteínas Fetais/genética , Proteínas Fetais/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Fator 3-beta Nuclear de Hepatócito/genética , Fator 3-beta Nuclear de Hepatócito/metabolismo , Camundongos , Linha Primitiva/citologia , Linha Primitiva/metabolismo , Proteínas com Domínio T/genética , Proteínas com Domínio T/metabolismo
11.
EMBO J ; 38(20): e102161, 2019 10 15.
Artigo em Inglês | MEDLINE | ID: mdl-31531882

RESUMO

Differentiation of normal and tumor cells is controlled by regulatory networks enforced by lineage-determining transcription factors (TFs). Among them, TFs such as FOXA1/2 bind naïve chromatin and induce its accessibility, thus establishing new gene regulatory networks. Pancreatic ductal adenocarcinoma (PDAC) is characterized by the coexistence of well- and poorly differentiated cells at all stages of disease. How the transcriptional networks determining such massive cellular heterogeneity are established remains to be determined. We found that FOXA2, a TF controlling pancreas specification, broadly contributed to the cis-regulatory networks of PDACs. Despite being expressed in both well- and poorly differentiated PDAC cells, FOXA2 displayed extensively different genomic distributions and controlled distinct gene expression programs. Grade-specific functions of FOXA2 depended on its partnership with TFs whose expression varied depending on the differentiation grade. These data suggest that FOXA2 contributes to the regulatory networks of heterogeneous PDAC cells via interactions with alternative partner TFs.


Assuntos
Diferenciação Celular , Regulação Neoplásica da Expressão Gênica , Fator 1-beta Nuclear de Hepatócito/metabolismo , Fator 3-beta Nuclear de Hepatócito/metabolismo , Proteínas de Homeodomínio/metabolismo , Neoplasias Pancreáticas/patologia , Elementos Reguladores de Transcrição , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/metabolismo , Carcinoma Ductal Pancreático/patologia , Movimento Celular , Proliferação de Células , Redes Reguladoras de Genes , Fator 1-beta Nuclear de Hepatócito/genética , Fator 3-alfa Nuclear de Hepatócito/genética , Fator 3-alfa Nuclear de Hepatócito/metabolismo , Fator 3-beta Nuclear de Hepatócito/genética , Proteínas de Homeodomínio/genética , Humanos , Gradação de Tumores , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Células Tumorais Cultivadas
12.
Am J Med Genet A ; 179(9): 1783-1790, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31294511

RESUMO

Rare individuals with 20p11.2 proximal deletions have been previously reported, with a variable phenotype that includes heterotaxy, biliary atresia, midline brain defects associated with panhypopituitarism, intellectual disability, scoliosis, and seizures. Deletions have ranged in size from 277 kb to 11.96 Mb. We describe a newborn with a de novo 2.7 Mb deletion of 20p11.22p11.21 that partially overlaps previously reported deletions and encompasses FOXA2. Her clinical findings further expand the 20p11.2 deletion phenotype to include severe midline cranial and intracranial defects such as aqueductal stenosis with hydrocephalus, mesencephalosynapsis with diencephalic-mesencephalic junction dysplasia, and pyriform aperture stenosis. We also report one individual with a missense variant in FOXA2 who had abnormal glucose homeostasis, panhypopituitarism, and endodermal organ dysfunction. Together, these findings support the critical role of FOXA2 in panhypopituitarism and midline defects.


Assuntos
Encéfalo/anormalidades , Constrição Patológica/genética , Fator 3-beta Nuclear de Hepatócito/genética , Hipopituitarismo/genética , Encéfalo/diagnóstico por imagem , Encéfalo/fisiopatologia , Deleção Cromossômica , Cromossomos Humanos Par 20/genética , Constrição Patológica/diagnóstico por imagem , Constrição Patológica/fisiopatologia , Predisposição Genética para Doença , Humanos , Hidrocefalia/diagnóstico por imagem , Hidrocefalia/genética , Hidrocefalia/fisiopatologia , Hipopituitarismo/diagnóstico por imagem , Hipopituitarismo/fisiopatologia , Recém-Nascido , Mutação de Sentido Incorreto/genética , Fenótipo , Córtex Piriforme/diagnóstico por imagem , Córtex Piriforme/fisiopatologia
13.
Cell Rep ; 28(2): 382-393.e7, 2019 07 09.
Artigo em Inglês | MEDLINE | ID: mdl-31291575

RESUMO

Transcriptional regulatory mechanisms of lineage priming in embryonic development are largely uncharacterized because of the difficulty of isolating transient progenitor populations. Directed differentiation of human pluripotent stem cells (hPSCs) combined with gene editing provides a powerful system to define precise temporal gene requirements for progressive chromatin changes during cell fate transitions. Here, we map the dynamic chromatin landscape associated with sequential stages of pancreatic differentiation from hPSCs. Our analysis of chromatin accessibility dynamics led us to uncover a requirement for FOXA2, known as a pioneer factor, in human pancreas specification not previously shown from mouse knockout studies. FOXA2 knockout hPSCs formed reduced numbers of pancreatic progenitors accompanied by impaired recruitment of GATA6 to pancreatic enhancers. Furthermore, FOXA2 is required for proper chromatin remodeling and H3K4me1 deposition during enhancer priming. This work highlights the power of combining hPSC differentiation, genome editing, and computational genomics for discovering transcriptional mechanisms during development.


Assuntos
Fator 3-beta Nuclear de Hepatócito/fisiologia , Pâncreas/fisiologia , Diferenciação Celular/fisiologia , Linhagem Celular , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/metabolismo , Células-Tronco Embrionárias/fisiologia , Fator 3-beta Nuclear de Hepatócito/genética , Fator 3-beta Nuclear de Hepatócito/metabolismo , Humanos , Masculino , Pâncreas/citologia , Pâncreas/metabolismo , Transcriptoma
14.
Nucleic Acids Res ; 47(17): 9069-9086, 2019 09 26.
Artigo em Inglês | MEDLINE | ID: mdl-31350899

RESUMO

Pioneer transcription factors (PTF) can recognize their binding sites on nucleosomal DNA and trigger chromatin opening for recruitment of other non-pioneer transcription factors. However, critical properties of PTFs are still poorly understood, such as how these transcription factors selectively recognize cell type-specific binding sites and under which conditions they can initiate chromatin remodelling. Here we show that early endoderm binding sites of the paradigm PTF Foxa2 are epigenetically primed by low levels of active chromatin modifications in embryonic stem cells (ESC). Priming of these binding sites is supported by preferential recruitment of Foxa2 to endoderm binding sites compared to lineage-inappropriate binding sites, when ectopically expressed in ESCs. We further show that binding of Foxa2 is required for chromatin opening during endoderm differentiation. However, increased chromatin accessibility was only detected on binding sites which are synergistically bound with other endoderm transcription factors. Thus, our data suggest that binding site selection of PTFs is directed by the chromatin environment and that chromatin opening requires collaboration of PTFs with additional transcription factors.


Assuntos
Cromatina/metabolismo , Fator 3-beta Nuclear de Hepatócito/metabolismo , Células-Tronco Embrionárias Murinas/metabolismo , Animais , Sítios de Ligação/genética , Diferenciação Celular/genética , Montagem e Desmontagem da Cromatina/genética , Endoderma/citologia , Fator de Transcrição GATA4/genética , Fator de Transcrição GATA4/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/genética , Fator 3-beta Nuclear de Hepatócito/genética , Código das Histonas , Histonas/metabolismo , Camundongos , Camundongos Knockout , Modelos Genéticos , Células-Tronco Embrionárias Murinas/citologia , Transdução de Sinais
15.
Hepatobiliary Pancreat Dis Int ; 18(6): 546-556, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31230960

RESUMO

BACKGROUND: Our previous study showed that overexpression of hepatocyte nuclear factor 4α (HNF4α) could directly promote mesenchymal stem cells (MSCs) to differentiate into hepatocyte-like cells. However, the efficiency of hepatic differentiation remains low. The purpose of our study was to establish an MSC cell line that overexpressed HNF4α and FOXA2 genes to obtain an increased hepatic differentiation efficiency and hepatocyte-like cells with more mature hepatocyte functions. METHODS: Successful establishment of high-level HNF4α and FOXA2 co-overexpression in human induced hepatocyte-like cells (hiHep cells) was verified by flow cytometry, immunofluorescence and RT-PCR. Measurements of albumin (ALB), urea, glucose, indocyanine green (ICG) uptake and release, cytochrome P450 (CYP) activity and gene expression were used to analyze mature hepatic functions of hiHep cells. RESULTS: hiHep cells efficiently express HNF4α and FOXA2 genes and proteins, exhibit typical epithelial morphology and acquire mature hepatocyte-like cell functions, including ALB secretion, urea production, ICG uptake and release, and glycogen storage. hiHep cells can be activated by CYP inducers. The percentage of both ALB and α-1-antitrypsin (AAT)-positive cells was approximately 72.6%. The expression levels of hepatocyte-specific genes (ALB, AAT, and CYP1A1) and liver drug transport-related genes (ABCB1, ABCG2, and SLC22A18) in hiHep cells were significantly higher than those in MSCs-Vector cells. The hiHep cells did not form tumors after subcutaneous xenograft in BALB/c nude mice after 2 months. CONCLUSION: This study provides an accessible, feasible and efficient strategy to generate hiHep cells from MSCs.


Assuntos
Diferenciação Celular , Fator 3-beta Nuclear de Hepatócito/metabolismo , Fator 4 Nuclear de Hepatócito/metabolismo , Hepatócitos/metabolismo , Células-Tronco Mesenquimais/metabolismo , Animais , Biomarcadores/metabolismo , Linhagem Celular , Proliferação de Células , Forma Celular , Feminino , Fator 3-beta Nuclear de Hepatócito/genética , Fator 4 Nuclear de Hepatócito/genética , Humanos , Camundongos Endogâmicos BALB C , Camundongos Nus , Transdução de Sinais , Transfecção , Regulação para Cima
16.
Anticancer Res ; 39(5): 2351-2360, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31092427

RESUMO

BACKGROUND/AIM: Hepatic hemangiomas (HH) can show an aggressive course with significant complications. Prognostic markers that identify an aggressive course are entirely absent. Since we have showed that Hedgehog signaling is overexpressed in aggressive hemangiomas of the skin. Here, we hypothesize that it is also altered in aggressive HH. MATERIALS AND METHODS: Immunohistological staining for GLUT1 and quantitative PCR was performed in seven specimens with aggressive HH. For comparison, we included specimens of kaposiform hemangioendothelioma (KHE), skin hemangioma and normal liver tissue. RESULTS: Overexpression of the Hedgehog signaling components SHH and GLI2 and its target gene FOXA2 in HH were similar to those found in aggressive skin hemangioma and KHE, their expression being significantly higher than in mild skin hemangioma. High expression levels of SHH and FOXA2 positively correlated with HH, but not with normal liver tissue. CONCLUSION: Hedgehog signaling is up-regulated in aggressive HH. This finding may lead to a biomarker allowing early intervention.


Assuntos
Proteínas Hedgehog/genética , Hemangioma/genética , Fator 3-beta Nuclear de Hepatócito/genética , Neoplasias Hepáticas/genética , Proteínas Nucleares/genética , Proteína Gli2 com Dedos de Zinco/genética , Biomarcadores Tumorais/genética , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Transportador de Glucose Tipo 1/genética , Hemangioendotelioma/genética , Hemangioendotelioma/patologia , Hemangioma/patologia , Humanos , Lactente , Recém-Nascido , Síndrome de Kasabach-Merritt/genética , Síndrome de Kasabach-Merritt/patologia , Fígado/metabolismo , Neoplasias Hepáticas/patologia , Masculino , Sarcoma de Kaposi/genética , Sarcoma de Kaposi/patologia , Transdução de Sinais/genética
17.
Mol Cell ; 75(1): 154-171.e5, 2019 07 11.
Artigo em Inglês | MEDLINE | ID: mdl-31056445

RESUMO

The epigenetic information present in mammalian gametes and whether it is transmitted to the progeny are relatively unknown. We find that many promoters in mouse sperm are occupied by RNA polymerase II (Pol II) and Mediator. The same promoters are accessible in GV and MII oocytes and preimplantation embryos. Sperm distal ATAC-seq sites containing motifs for various transcription factors are conserved in monkeys and humans. ChIP-seq analyses confirm that Foxa1, ERα, and AR occupy distal enhancers in sperm. Accessible sperm enhancers containing H3.3 and H2A.Z are also accessible in oocytes and preimplantation embryos. Furthermore, their interactions with promoters in the gametes persist during early development. Sperm- or oocyte-specific interactions mediated by CTCF and cohesin are only present in the paternal or maternal chromosomes, respectively, in the zygote and 2-cell stages. These interactions converge in both chromosomes by the 8-cell stage. Thus, mammalian gametes contain complex patterns of 3D interactions that can be transmitted to the zygote after fertilization.


Assuntos
Fator de Ligação a CCCTC/genética , Fator 3-beta Nuclear de Hepatócito/genética , Oócitos/metabolismo , Espermatozoides/metabolismo , Zigoto/metabolismo , Animais , Sequência de Bases , Fator de Ligação a CCCTC/metabolismo , Cromatina/química , Cromatina/metabolismo , Sequência Conservada , Embrião de Mamíferos , Desenvolvimento Embrionário/genética , Elementos Facilitadores Genéticos , Receptor alfa de Estrogênio/genética , Receptor alfa de Estrogênio/metabolismo , Receptor beta de Estrogênio/genética , Receptor beta de Estrogênio/metabolismo , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Fator 3-beta Nuclear de Hepatócito/metabolismo , Humanos , Macaca mulatta , Masculino , Camundongos , Oócitos/citologia , Oócitos/crescimento & desenvolvimento , Receptores Androgênicos/genética , Receptores Androgênicos/metabolismo , Homologia de Sequência do Ácido Nucleico , Espermatozoides/citologia , Espermatozoides/crescimento & desenvolvimento , Dedos de Zinco/genética , Zigoto/citologia , Zigoto/crescimento & desenvolvimento
18.
Cell Rep ; 27(3): 708-718.e10, 2019 04 16.
Artigo em Inglês | MEDLINE | ID: mdl-30995470

RESUMO

Studies in vertebrates have outlined conserved molecular control of definitive endoderm (END) development. However, recent work also shows that key molecular aspects of human END regulation differ even from rodents. Differentiation of human embryonic stem cells (ESCs) to END offers a tractable system to study the molecular basis of normal and defective human-specific END development. Here, we interrogated dynamics in chromatin accessibility during differentiation of ESCs to END, predicting DNA-binding proteins that may drive this cell fate transition. We then combined single-cell RNA-seq with parallel CRISPR perturbations to comprehensively define the loss-of-function phenotype of those factors in END development. Following a few candidates, we revealed distinct impairments in the differentiation trajectories for mediators of TGFß signaling and expose a role for the FOXA2 transcription factor in priming human END competence for human foregut and hepatic END specification. Together, this single-cell functional genomics study provides high-resolution insight on human END development.


Assuntos
Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas/genética , RNA Guia/metabolismo , Fatores de Transcrição/metabolismo , Diferenciação Celular , Cromatina/metabolismo , Endoderma/citologia , Endoderma/metabolismo , Fator 3-beta Nuclear de Hepatócito/antagonistas & inibidores , Fator 3-beta Nuclear de Hepatócito/genética , Fator 3-beta Nuclear de Hepatócito/metabolismo , Células-Tronco Embrionárias Humanas/citologia , Células-Tronco Embrionárias Humanas/metabolismo , Humanos , Células-Tronco Pluripotentes/citologia , Células-Tronco Pluripotentes/metabolismo , Interferência de RNA , Fatores de Transcrição SOXF/genética , Fatores de Transcrição SOXF/metabolismo , Transdução de Sinais , Análise de Célula Única , Proteína Smad4/genética , Proteína Smad4/metabolismo , Fatores de Transcrição/antagonistas & inibidores , Fatores de Transcrição/genética , Fator de Crescimento Transformador beta/metabolismo
19.
Genes Dev ; 33(11-12): 656-668, 2019 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-30923168

RESUMO

Transcription factors (TFs) are dosage-sensitive master regulators of gene expression, with haploinsufficiency frequently leading to life-threatening disease. Numerous mechanisms have evolved to tightly regulate the expression and activity of TFs at the transcriptional, translational, and posttranslational levels. A subset of long noncoding RNAs (lncRNAs) is spatially correlated with transcription factors in the genome, but the regulatory relationship between these lncRNAs and their neighboring TFs is unclear. We identified a regulatory feedback loop between the TF Foxa2 and a downstream lncRNA, Falcor (Foxa2-adjacent long noncoding RNA). Foxa2 directly represses Falcor expression by binding to its promoter, while Falcor functions in cis to positively regulate the expression of Foxa2. In the lung, loss of Falcor is sufficient to lead to chronic inflammatory changes and defective repair after airway epithelial injury. Moreover, disruption of the Falcor-Foxa2 regulatory feedback loop leads to altered cell adhesion and migration, in turn resulting in chronic peribronchial airway inflammation and goblet cell metaplasia. These data reveal that the lncRNA Falcor functions within a regulatory feedback loop to fine-tune the expression of Foxa2, maintain airway epithelial homeostasis, and promote regeneration.


Assuntos
Células Epiteliais/metabolismo , Fator 3-beta Nuclear de Hepatócito/genética , Pulmão/citologia , Pulmão/metabolismo , RNA Longo não Codificante/genética , Animais , Adesão Celular , Linhagem Celular , Movimento Celular , Feminino , Regulação da Expressão Gênica , Fator 3-beta Nuclear de Hepatócito/metabolismo , Homeostase , Humanos , Masculino , Camundongos , Regiões Promotoras Genéticas , Regeneração , Transcrição Genética
20.
Mol Biol Rep ; 46(2): 2355-2362, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30778924

RESUMO

A large proportion of the transcriptome is sex biased in a wide range of taxa. Sexually dimorphic genes expression is highly tissue-dependent. Although gastric cancer exhibits sex bias to some extent, sexually dimorphic gene expression in the stomach is yet to be fully understood. The aim of the present study was to compare the expression levels of 12 genes in the gastric tissue between age-matched healthy men and women of different age groups. A total of 70 human antrum gastric tissue samples were obtained by endoscopy. The difference in expression levels of the 12 intended genes between two genders was investigated using quantitative Real-Time PCR, following total RNA extraction. The results indicated that the expression levels of both the GKN2 (7.2-fold, p < 0.001) and FOXA2 (3.7-fold, p = 0.003) were significantly higher in men compared to those in women. In addition, FOXA1 gene expression was age-dependent only in women. Interestingly, the expression level of FOXA1 was significantly higher in premenopausal women compared to postmenopausal women (2.53-fold, p = 0.016). The expression levels of some of the investigated genes in this study were sex-dependent in the stomach. This sexual dimorphism in gene expression might influence the differential susceptibility to the gastric cancer between the sexes.


Assuntos
Proteínas de Transporte/genética , Fator 3-beta Nuclear de Hepatócito/genética , Adulto , Fatores Etários , Idoso , Proteínas de Transporte/metabolismo , Feminino , Expressão Gênica/genética , Perfilação da Expressão Gênica/métodos , Fator 3-alfa Nuclear de Hepatócito/genética , Fator 3-alfa Nuclear de Hepatócito/metabolismo , Fator 3-beta Nuclear de Hepatócito/metabolismo , Humanos , Masculino , Pessoa de Meia-Idade , Caracteres Sexuais , Estômago/fisiologia , Neoplasias Gástricas , Transcriptoma/genética
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