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1.
Nat Commun ; 12(1): 1020, 2021 02 15.
Artículo en Inglés | MEDLINE | ID: mdl-33589611

RESUMEN

The extracellular matrix (ECM) is unique to each tissue and capable of guiding cell differentiation, migration, morphology, and function. The ECM proteome of different developmental stages has not been systematically studied in the human pancreas. In this study, we apply mass spectrometry-based quantitative proteomics strategies using N,N-dimethyl leucine isobaric tags to delineate proteome-wide and ECM-specific alterations in four age groups: fetal (18-20 weeks gestation), juvenile (5-16 years old), young adults (21-29 years old) and older adults (50-61 years old). We identify 3,523 proteins including 185 ECM proteins and quantify 117 of them. We detect previously unknown proteome and matrisome features during pancreas development and maturation. We also visualize specific ECM proteins of interest using immunofluorescent staining and investigate changes in ECM localization within islet or acinar compartments. This comprehensive proteomics analysis contributes to an improved understanding of the critical roles that ECM plays throughout human pancreas development and maturation.


Asunto(s)
Proteínas de la Matriz Extracelular/genética , Regulación del Desarrollo de la Expresión Génica , Páncreas/metabolismo , Proteoma/genética , Adolescente , Adulto , Niño , Preescolar , Cromatografía Liquida , Matriz Extracelular/genética , Matriz Extracelular/metabolismo , Proteínas de la Matriz Extracelular/clasificación , Proteínas de la Matriz Extracelular/metabolismo , Femenino , Feto , Técnica del Anticuerpo Fluorescente , Ontología de Genes , Humanos , Masculino , Persona de Mediana Edad , Anotación de Secuencia Molecular , Organogénesis/genética , Páncreas/crecimiento & desarrollo , Proteoma/clasificación , Proteoma/metabolismo , Proteómica/métodos , Espectrometría de Masas en Tándem
2.
Nat Commun ; 12(1): 833, 2021 02 05.
Artículo en Inglés | MEDLINE | ID: mdl-33547280

RESUMEN

The structure of proline prevents it from adopting an optimal position for rapid protein synthesis. Poly-proline-tract (PPT) associated ribosomal stalling is resolved by highly conserved eIF5A, the only protein to contain the amino acid hypusine. We show that de novo heterozygous EIF5A variants cause a disorder characterized by variable combinations of developmental delay, microcephaly, micrognathia and dysmorphism. Yeast growth assays, polysome profiling, total/hypusinated eIF5A levels and PPT-reporters studies reveal that the variants impair eIF5A function, reduce eIF5A-ribosome interactions and impair the synthesis of PPT-containing proteins. Supplementation with 1 mM spermidine partially corrects the yeast growth defects, improves the polysome profiles and restores expression of PPT reporters. In zebrafish, knockdown eif5a partly recapitulates the human phenotype that can be rescued with 1 µM spermidine supplementation. In summary, we uncover the role of eIF5A in human development and disease, demonstrate the mechanistic complexity of EIF5A-related disorder and raise possibilities for its treatment.


Asunto(s)
Discapacidades del Desarrollo/genética , Regulación del Desarrollo de la Expresión Génica , Microcefalia/genética , Micrognatismo/genética , Factores de Iniciación de Péptidos/genética , Proteínas de Unión al ARN/genética , Adolescente , Secuencia de Aminoácidos , Animales , Niño , Discapacidades del Desarrollo/metabolismo , Discapacidades del Desarrollo/patología , Embrión no Mamífero , Femenino , Humanos , Lisina/análogos & derivados , Lisina/genética , Lisina/metabolismo , Masculino , Microcefalia/metabolismo , Microcefalia/patología , Micrognatismo/metabolismo , Micrognatismo/patología , Factores de Iniciación de Péptidos/deficiencia , Péptidos/genética , Péptidos/metabolismo , Biosíntesis de Proteínas , Conformación Proteica , Isoformas de Proteínas/deficiencia , Isoformas de Proteínas/genética , Ribosomas/genética , Ribosomas/metabolismo , Saccharomyces cerevisiae/efectos de los fármacos , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/crecimiento & desarrollo , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Alineación de Secuencia , Homología de Secuencia de Aminoácido , Espermidina/farmacología , Pez Cebra , Proteínas de Pez Cebra/genética , Proteínas de Pez Cebra/metabolismo
3.
Nat Commun ; 12(1): 841, 2021 02 05.
Artículo en Inglés | MEDLINE | ID: mdl-33547291

RESUMEN

A new life begins with the unification of the maternal and paternal chromosomes upon fertilization. The parental chromosomes first become enclosed in two separate pronuclei near the surface of the fertilized egg. The mechanisms that then move the pronuclei inwards for their unification are only poorly understood in mammals. Here, we report two mechanisms that act in concert to unite the parental genomes in fertilized mouse eggs. The male pronucleus assembles within the fertilization cone and is rapidly moved inwards by the flattening cone. Rab11a recruits the actin nucleation factors Spire and Formin-2 into the fertilization cone, where they locally nucleate actin and further accelerate the pronucleus inwards. In parallel, a dynamic network of microtubules assembles that slowly moves the male and female pronuclei towards the cell centre in a dynein-dependent manner. Both mechanisms are partially redundant and act in concert to unite the parental pronuclei in the zygote's centre.


Asunto(s)
Núcleo Celular/metabolismo , Fertilización/genética , Forminas/genética , Proteínas de Microfilamentos/genética , Proteínas del Tejido Nervioso/genética , Cigoto/metabolismo , Proteínas de Unión al GTP rab/genética , Actinas/genética , Actinas/metabolismo , Animales , Núcleo Celular/ultraestructura , Femenino , Forminas/metabolismo , Regulación del Desarrollo de la Expresión Génica , Genes Reporteros , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Proteínas Luminiscentes/genética , Proteínas Luminiscentes/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos CBA , Proteínas de Microfilamentos/metabolismo , Microtúbulos/metabolismo , Microtúbulos/ultraestructura , Movimiento , Proteínas del Tejido Nervioso/metabolismo , Oocitos/metabolismo , Oocitos/ultraestructura , Espermatozoides/metabolismo , Espermatozoides/ultraestructura , Cigoto/ultraestructura , Proteínas de Unión al GTP rab/metabolismo
4.
Ecotoxicol Environ Saf ; 208: 111716, 2021 Jan 15.
Artículo en Inglés | MEDLINE | ID: mdl-33396047

RESUMEN

Although withdrawn from the market in the 1980s, polychlorinated biphenyls (PCBs) are still found ubiquitously in the aquatic environment and pose a serious risk to biota due to their teratogenic potential. In fish, early life-stages are often considered most sensitive with regard to their exposure to PCBs and other dioxin-like compounds. However, little is known about the molecular drivers of the frequently observed teratogenic effects. Therefore, the aims of our study were to: (1) characterize the baseline transcriptome profiles at different embryonic life-stages in zebrafish (Danio rerio); and (2) to identify the molecular response to PCB exposure and life-stage specific-effects of the chemical on associated processes. For both objectives, embryos were sampled at 12, 48, and 96 h post-fertilization (hpf) and subjected to Illumina sequence-by-synthesis and RNAseq analysis. Results revealed that with increasing age more genes and related pathways were upregulated both in terms of number and magnitude. Yet, other transcripts followed an opposite pattern with greater transcript abundance at the earlier time points. Additionally, embryos were exposed to PCB126, a potent agonist of the aryl hydrocarbon receptor (AHR). ClueGO network analysis revealed significant enrichment of genes associated with basic cell metabolism, communication, and homeostasis as well as eye development, muscle formation, and skeletal formation. We selected eight genes involved in the affected pathways for an in-depth characterization of their regulation throughout normal embryogenesis and after exposure to PCB126 by quantification of transcript abundances every 12 h until 118 hpf. Among these, fgf7 and c9 stood out because of their strong upregulation by PCB126 exposure at 48 and 96 hpf, respectively. Cyp2aa12 was upregulated from 84 hpf on. Fabp10ab, myhz1.1, col8a1a, sulf1, and opn1sw1 displayed specific regulation depending on the developmental stage. Overall, we demonstrate that (1) the developmental transcriptome of zebrafish is highly dynamic, and (2) dysregulation of gene expression by exposure to PCB126 was significant and in several cases not directly connected to AHR-signaling. Hence, this study improves the understanding of linkages between molecular events and apical outcomes that are of regulatory relevance.


Asunto(s)
Desarrollo Embrionario/efectos de los fármacos , Bifenilos Policlorados/toxicidad , Teratógenos/toxicidad , Contaminantes Químicos del Agua/toxicidad , Pez Cebra/embriología , Animales , Embrión no Mamífero/efectos de los fármacos , Desarrollo Embrionario/genética , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Receptores de Hidrocarburo de Aril/antagonistas & inhibidores , Transcriptoma , Pez Cebra/genética , Pez Cebra/metabolismo , Proteínas de Pez Cebra/antagonistas & inhibidores
5.
Sichuan Da Xue Xue Bao Yi Xue Ban ; 52(1): 82-86, 2021 Jan.
Artículo en Chino | MEDLINE | ID: mdl-33474894

RESUMEN

Objective: In order to better understand the role of mechanical stress in early tooth development, we examined the spatiotemporal expression patterns of mechanical-stress related regulatory protein (actin filament, or F-actin), non-muscle myosin ⅡB (NMⅡB) and vinculin at different stages of tooth development in mice. Methods: Mouse first mandible molars were used as the research model. Immunofluorescence staining was performed to detect the expression patterns of F-actin, NMⅡB and Vinculin, the key molecules constituting the chemical mechanical system, at bud, cap, early bell and late bell stages of tooth. Results: F-actin, NMⅡB and vinculin were all expressed in the tooth epithelium in an extensive or a limited way at different stages of tooth development, while F-actin was also expressed steadily in the mesenchymal cells. The quantitative analysis of fluorescence intensity showed that F-actin and NMⅡB exhibited significantly increase in the early stage of tooth development, but then dropped to their basal levels at the end of the late bell stage and the early bell stage respectively, with the differences of expression changes between successive developmental stages showing statistically significance ( P<0.05). Vinculin expression, however, remained at a relatively constant level except for the late bell stage when vinculin expression was slightly elevated compared to that of the early bell stage ( P<0.05). Conclusions: The findings suggest that mechanical stress is involved in early tooth development. F-actin may have an important role in dispersing and transmitting mechanical stress while NMⅡB may participate in tooth epithelial invagination and cusps formation. The findings also suggest that vinculin can respond to the mechanical stimuli and its interaction with cell adhesion molecules may play a role in tooth development. The mechanism of how actomyosin and cell adhesion interact with each other in regulating tooth development still needs further investigation.


Asunto(s)
Regulación del Desarrollo de la Expresión Génica , Odontogénesis , Animales , Epitelio , Ratones , Diente Molar , Odontogénesis/genética , Estrés Mecánico
6.
Nat Commun ; 12(1): 50, 2021 01 04.
Artículo en Inglés | MEDLINE | ID: mdl-33397914

RESUMEN

Gene flow is predicted to impede parallel adaptation via de novo mutation, because it can introduce pre-existing adaptive alleles from population to population. We test this using Hawaiian crickets (Teleogryllus oceanicus) in which 'flatwing' males that lack sound-producing wing structures recently arose and spread under selection from an acoustically-orienting parasitoid. Morphometric and genetic comparisons identify distinct flatwing phenotypes in populations on three islands, localized to different loci. Nevertheless, we detect strong, recent and ongoing gene flow among the populations. Using genome scans and gene expression analysis we find that parallel evolution of flatwing on different islands is associated with shared genomic hotspots of adaptation that contain the gene doublesex, but the form of selection differs among islands and corresponds to known flatwing demographics in the wild. We thus show how parallel adaptation can occur on contemporary timescales despite gene flow, indicating that it could be less constrained than previously appreciated.


Asunto(s)
Adaptación Fisiológica/genética , Flujo Génico , Gryllidae/genética , Gryllidae/fisiología , Animales , Regulación del Desarrollo de la Expresión Génica , Sitios Genéticos , Variación Genética , Genética de Población , Genoma de los Insectos , Geografía , Hawaii , Islas , Masculino , Selección Genética , Alas de Animales/anatomía & histología
7.
Hereditas ; 158(1): 4, 2021 Jan 04.
Artículo en Inglés | MEDLINE | ID: mdl-33397514

RESUMEN

BACKGROUND: The coronavirus disease 2019 (COVID-19) has spread rapidly around the world. In addition to common respiratory symptoms such as cough and fever, some patients also have cardiac injury, however, the mechanism of cardiac injury is not clear. In this study, we analyzed the RNA expression atlases of angiotensin-converting enzyme 2(ACE2), cathepsin B (CTSB) and cathepsin L (CTSL) in the human embryonic heart at single-cell resolution. RESULTS: The results showed that ACE2 was preferentially enriched in cardiomyocytes. Interestingly, serine protease transmembrane serine protease 2 (TMPRSS2) had less expression in cardiomyocytes, but CTSB and CTSL, which belonged to cell protease, could be found to be enriched in cardiomyocytes. The results of enrichment analysis showed that differentially expressed genes (DEGs) in ACE2-positive cardiomyocytes were mainly enriched in the processes of cardiac muscle contraction, regulation of cardiac conduction, mitochondrial respiratory chain, ion channel binding, adrenergic signaling in cardiomyocytes and viral transcription. CONCLUSIONS: Our study suggests that both atrial and ventricular cardiomyocytes are potentially susceptible to severe acute respiratory syndrome coronavirus-2(SARS-CoV-2), and SARS-CoV-2 may enter ventricular cardiomyocytes using CTSB/CTSL for S protein priming. This may be the partial cellular mechanism of cardiac injury in patients with COVID-19.


Asunto(s)
/prevención & control , Regulación del Desarrollo de la Expresión Génica , Corazón/embriología , Miocitos Cardíacos/metabolismo , Análisis de la Célula Individual/métodos , /genética , /virología , Catepsina B/genética , Catepsina L/genética , Ontología de Genes , Humanos , Miocitos Cardíacos/citología , Miocitos Cardíacos/virología , Pandemias , Serina Endopeptidasas/genética , Transducción de Señal/genética
8.
Nat Commun ; 12(1): 463, 2021 01 19.
Artículo en Inglés | MEDLINE | ID: mdl-33469025

RESUMEN

Splicing varies across brain regions, but the single-cell resolution of regional variation is unclear. We present a single-cell investigation of differential isoform expression (DIE) between brain regions using single-cell long-read sequencing in mouse hippocampus and prefrontal cortex in 45 cell types at postnatal day 7 ( www.isoformAtlas.com ). Isoform tests for DIE show better performance than exon tests. We detect hundreds of DIE events traceable to cell types, often corresponding to functionally distinct protein isoforms. Mostly, one cell type is responsible for brain-region specific DIE. However, for fewer genes, multiple cell types influence DIE. Thus, regional identity can, although rarely, override cell-type specificity. Cell types indigenous to one anatomic structure display distinctive DIE, e.g. the choroid plexus epithelium manifests distinct transcription-start-site usage. Spatial transcriptomics and long-read sequencing yield a spatially resolved splicing map. Our methods quantify isoform expression with cell-type and spatial resolution and it contributes to further our understanding of how the brain integrates molecular and cellular complexity.


Asunto(s)
Empalme Alternativo/fisiología , Regulación del Desarrollo de la Expresión Génica/fisiología , Hipocampo/metabolismo , Corteza Prefrontal/metabolismo , Isoformas de Proteínas/metabolismo , Animales , Animales Recién Nacidos , Biología Computacional , Femenino , Hipocampo/citología , Hipocampo/crecimiento & desarrollo , Ratones , Modelos Animales , Corteza Prefrontal/citología , Corteza Prefrontal/crecimiento & desarrollo , Isoformas de Proteínas/análisis , Isoformas de Proteínas/genética , Análisis de la Célula Individual/métodos , Análisis Espacial
9.
Nucleic Acids Res ; 49(2): 726-744, 2021 01 25.
Artículo en Inglés | MEDLINE | ID: mdl-33406262

RESUMEN

The establishment of the small intestinal (SI) lineage during human embryogenesis ensures functional integrity of the intestine after birth. The chromatin dynamics that drive SI lineage formation and regional patterning in humans are essentially unknown. To fill this knowledge void, we apply a cutting-edge genomic technology to a state-of-the-art human model of early SI development. Specifically, we leverage chromatin run-on sequencing (ChRO-seq) to define the landscape of active promoters, enhancers and gene bodies across distinct stages of directed differentiation of human pluripotent stem cells into SI spheroids with regional specification. Through comprehensive ChRO-seq analysis we identify candidate stage-specific chromatin activity states, novel markers and enhancer hotspots during the directed differentiation. Moreover, we propose a detailed transcriptional network associated with SI lineage formation or regional patterning. Our ChRO-seq analyses uncover a previously undescribed pattern of enhancer activity and transcription at HOX gene loci underlying SI regional patterning. We also validated this unique HOX dynamics by the analysis of single cell RNA-seq data from human fetal SI. Overall, the results lead to a new proposed working model for the regulatory underpinnings of human SI development, thereby adding a novel dimension to the literature that has relied almost exclusively on non-human models.


Asunto(s)
Ensamble y Desensamble de Cromatina , Regulación del Desarrollo de la Expresión Génica , Células Madre Embrionarias Humanas/metabolismo , Intestino Delgado/embriología , Modelos Biológicos , Animales , Diferenciación Celular , Línea Celular , Linaje de la Célula , Elementos de Facilitación Genéticos , Genes Homeobox , Células Madre Embrionarias Humanas/citología , Humanos , Intestino Delgado/metabolismo , Ratones , Organoides , ARN Mensajero/biosíntesis , ARN Mensajero/genética , Análisis de Secuencia de ARN/métodos , Análisis de la Célula Individual , Transcripción Genética
10.
Ecotoxicol Environ Saf ; 210: 111876, 2021 Mar 01.
Artículo en Inglés | MEDLINE | ID: mdl-33418158

RESUMEN

Evidence has shown that miRNAs could play a role in dental fluorosis, but there is no study has investigated the global expression miRNA profiles of fluoride-exposed enamel organ. In this study, we analysed the differentially expressed (DE) miRNAs between fluoride-treated and control enamel organ for the first time and found several candidate miRNAs and signaling pathways worthy of further research. Thirty Wistar rats were randomly distributed into three groups and exposed to drinking water with different fluoride contents for 10 weeks and during the gestation. The three groups were a control group (distilled water), medium fluoride group (75 mg/L NaF), and high fluoride group (150 mg/L NaF). On the embryonic day 19.5, the mandible was dissected for histological analysis, and the enamel organ of the mandibular first molar tooth germ was collected for miRNA sequencing (miRNA-seq) and quantitative real-time PCR analysis (qRT-PCR). Typical dental fluorosis was observed in the incisors of the prepregnant rats. In addition to the disorganized structure of enamel organ cells, 39 DE miRNAs were identified in the fluoride groups compared with the control group, and good agreement between the miRNA-seq data and qRT-PCR data was found. The functional annotation of the target genes of 39 DE miRNAs showed significant enrichment in metabolic process, cell differentiation, calcium signaling pathway, and mitogen-activated protein kinase(MAPK) signaling pathway terms. This study provides a theoretical reference for an extensive understanding of the mechanism of fluorosis and potential valuable miRNAs as therapeutic targets in fluorosis.


Asunto(s)
Órgano del Esmalte/efectos de los fármacos , Fluoruros/toxicidad , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , MicroARNs , Animales , Embrión de Mamíferos , Órgano del Esmalte/embriología , Órgano del Esmalte/metabolismo , Femenino , Fluorosis Dental , Ratas Wistar , Transcriptoma/efectos de los fármacos
11.
Nat Commun ; 12(1): 464, 2021 01 19.
Artículo en Inglés | MEDLINE | ID: mdl-33469015

RESUMEN

Conventional dendritic cells (cDC) are key activators of naive T cells, and can be targeted in adults to induce adaptive immunity, but in early life are considered under-developed or functionally immature. Here we show that, in early life, when the immune system develops, cDC2 exhibit a dual hematopoietic origin and, like other myeloid and lymphoid cells, develop in waves. Developmentally distinct cDC2 in early life, despite being distinguishable by fate mapping, are transcriptionally and functionally similar. cDC2 in early and adult life, however, are exposed to distinct cytokine environments that shape their transcriptional profile and alter their ability to sense pathogens, secrete cytokines and polarize T cells. We further show that cDC2 in early life, despite being distinct from cDC2 in adult life, are functionally competent and can induce T cell responses. Our results thus highlight the potential of harnessing cDC2 for boosting immunity in early life.


Asunto(s)
Inmunidad Adaptativa/fisiología , Diferenciación Celular/genética , Citocinas/metabolismo , Células Dendríticas/inmunología , Regulación del Desarrollo de la Expresión Génica/inmunología , Factores de Edad , Animales , Diferenciación Celular/inmunología , Separación Celular , Células Dendríticas/metabolismo , Femenino , Citometría de Flujo , Células Madre Hematopoyéticas/fisiología , Masculino , Ratones , Ratones Transgénicos , Modelos Animales , Cultivo Primario de Células , RNA-Seq , Análisis de la Célula Individual , Linfocitos T/inmunología , Transcriptoma/inmunología
12.
Nat Commun ; 12(1): 439, 2021 01 19.
Artículo en Inglés | MEDLINE | ID: mdl-33469032

RESUMEN

Developmental genes are often regulated by multiple elements with overlapping activity. Yet, in most cases, the relative function of those elements and their contribution to endogenous gene expression remain poorly characterized. An example of this phenomenon is that distinct sets of enhancers have been proposed to direct Fgf8 in the limb apical ectodermal ridge and the midbrain-hindbrain boundary. Using in vivo CRISPR/Cas9 genome engineering, we functionally dissect this complex regulatory ensemble and demonstrate two distinct regulatory logics. In the apical ectodermal ridge, the control of Fgf8 expression appears distributed between different enhancers. In contrast, we find that in the midbrain-hindbrain boundary, one of the three active enhancers is essential while the other two are dispensable. We further dissect the essential midbrain-hindbrain boundary enhancer to reveal that it is also composed by a mixture of essential and dispensable modules. Cross-species transgenic analysis of this enhancer suggests that its composition may have changed in the vertebrate lineage.


Asunto(s)
Desarrollo Embrionario/genética , Elementos de Facilitación Genéticos/genética , Factor 8 de Crecimiento de Fibroblastos/genética , Regulación del Desarrollo de la Expresión Génica , Ingeniería Genética/métodos , Animales , Sistemas CRISPR-Cas/genética , Ectodermo/embriología , Embrión de Mamíferos , Extremidades/embriología , Estudios de Factibilidad , Femenino , Factor 8 de Crecimiento de Fibroblastos/metabolismo , Redes Reguladoras de Genes , Masculino , Mesencéfalo/embriología , Ratones , Ratones Transgénicos , Rombencéfalo/embriología
13.
Nat Commun ; 12(1): 485, 2021 01 20.
Artículo en Inglés | MEDLINE | ID: mdl-33473133

RESUMEN

Schistosomes are parasitic flatworms causing one of the most prevalent infectious diseases from which millions of people are currently suffering. These parasites have high fecundity and their eggs are both the transmissible agents and the cause of the infection-associated pathology. Given its biomedical significance, the schistosome germline has been a research focus for more than a century. Nonetheless, molecular mechanisms that regulate its development are only now being understood. In particular, it is unknown what balances the fate of germline stem cells (GSCs) in producing daughter stem cells through mitotic divisions versus gametes through meiosis. Here, we perform single-cell RNA sequencing on juvenile schistosomes and capture GSCs during de novo gonadal development. We identify a genetic program that controls the proliferation and differentiation of GSCs. This program centers around onecut, a homeobox transcription factor, and boule, an mRNA binding protein. Their expressions are mutually dependent in the schistosome male germline, and knocking down either of them causes over-proliferation of GSCs and blocks germ cell differentiation. We further show that this germline-specific regulatory program is conserved in the planarian, schistosome's free-living evolutionary cousin, but the function of onecut has changed during evolution to support GSC maintenance.


Asunto(s)
Células Germinativas/metabolismo , Schistosoma mansoni/fisiología , Análisis de la Célula Individual/métodos , Células Madre/metabolismo , Animales , Diferenciación Celular , Proliferación Celular , Femenino , Regulación del Desarrollo de la Expresión Génica , Genes Homeobox , Masculino , Ratones , Planarias , Interferencia de ARN , ARN Mensajero , Proteínas de Unión al ARN , Schistosoma mansoni/citología , Esquistosomiasis mansoni/parasitología , Factores de Transcripción
14.
Nat Commun ; 12(1): 256, 2021 01 11.
Artículo en Inglés | MEDLINE | ID: mdl-33431871

RESUMEN

In humans, inactivating mutations in MLL4, which encodes a histone H3-lysine 4-methyltransferase, lead to Kabuki syndrome (KS). While dwarfism is a cardinal feature of KS, the underlying etiology remains unclear. Here we report that Mll4 regulates the development of growth hormone-releasing hormone (GHRH)-producing neurons in the mouse hypothalamus. Our two Mll4 mutant mouse models exhibit dwarfism phenotype and impairment of the developmental programs for GHRH-neurons. Our ChIP-seq analysis reveals that, in the developing mouse hypothalamus, Mll4 interacts with the transcription factor Nrf1 to trigger the expression of GHRH-neuronal genes. Interestingly, the deficiency of Mll4 results in a marked reduction of histone marks of active transcription, while treatment with the histone deacetylase inhibitor AR-42 rescues the histone mark signature and restores GHRH-neuronal production in Mll4 mutant mice. Our results suggest that the developmental dysregulation of Mll4-directed epigenetic control of transcription plays a role in the development of GHRH-neurons and dwarfism phenotype in mice.


Asunto(s)
Hormona Liberadora de Hormona del Crecimiento/biosíntesis , N-Metiltransferasa de Histona-Lisina/metabolismo , Hipotálamo/citología , Neuronas/metabolismo , Animales , Secuencia de Bases , Enanismo/metabolismo , Embrión de Mamíferos/metabolismo , Epigénesis Genética , Regulación del Desarrollo de la Expresión Génica , Células HEK293 , Humanos , Hipotálamo/embriología , Masculino , Ratones Noqueados , Modelos Biológicos , Factor Nuclear 1 de Respiración/metabolismo , Fenilbutiratos/farmacología , Factores de Transcripción/metabolismo
15.
Nat Commun ; 12(1): 410, 2021 01 18.
Artículo en Inglés | MEDLINE | ID: mdl-33462227

RESUMEN

Active DNA demethylation is required for sexual reproduction in plants but the molecular determinants underlying this epigenetic control are not known. Here, we show in Arabidopsis thaliana that the DNA glycosylases DEMETER (DME) and REPRESSOR OF SILENCING 1 (ROS1) act semi-redundantly in the vegetative cell of pollen to demethylate DNA and ensure proper pollen tube progression. Moreover, we identify six pollen-specific genes with increased DNA methylation as well as reduced expression in dme and dme;ros1. We further show that for four of these genes, reinstalling their expression individually in mutant pollen is sufficient to improve male fertility. Our findings demonstrate an essential role of active DNA demethylation in regulating genes involved in pollen function.


Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/fisiología , Desmetilación del ADN , Regulación de la Expresión Génica de las Plantas , N-Glicosil Hidrolasas/metabolismo , Proteínas Nucleares/metabolismo , Transactivadores/metabolismo , Proteínas de Arabidopsis/genética , Fertilidad/genética , Regulación del Desarrollo de la Expresión Génica , Mutación , N-Glicosil Hidrolasas/genética , Proteínas Nucleares/genética , Plantas Modificadas Genéticamente , Tubo Polínico/crecimiento & desarrollo , Transactivadores/genética
16.
Am J Hum Genet ; 108(1): 115-133, 2021 01 07.
Artículo en Inglés | MEDLINE | ID: mdl-33308444

RESUMEN

Signal peptide-CUB-EGF domain-containing protein 3 (SCUBE3) is a member of a small family of multifunctional cell surface-anchored glycoproteins functioning as co-receptors for a variety of growth factors. Here we report that bi-allelic inactivating variants in SCUBE3 have pleiotropic consequences on development and cause a previously unrecognized syndromic disorder. Eighteen affected individuals from nine unrelated families showed a consistent phenotype characterized by reduced growth, skeletal features, distinctive craniofacial appearance, and dental anomalies. In vitro functional validation studies demonstrated a variable impact of disease-causing variants on transcript processing, protein secretion and function, and their dysregulating effect on bone morphogenetic protein (BMP) signaling. We show that SCUBE3 acts as a BMP2/BMP4 co-receptor, recruits the BMP receptor complexes into raft microdomains, and positively modulates signaling possibly by augmenting the specific interactions between BMPs and BMP type I receptors. Scube3-/- mice showed craniofacial and dental defects, reduced body size, and defective endochondral bone growth due to impaired BMP-mediated chondrogenesis and osteogenesis, recapitulating the human disorder. Our findings identify a human disease caused by defective function of a member of the SCUBE family, and link SCUBE3 to processes controlling growth, morphogenesis, and bone and teeth development through modulation of BMP signaling.


Asunto(s)
Huesos/metabolismo , Proteínas de Unión al Calcio/metabolismo , Discapacidades del Desarrollo/metabolismo , Osteogénesis/fisiología , Transducción de Señal/fisiología , Animales , Proteína Morfogenética Ósea 2/metabolismo , Proteína Morfogenética Ósea 4/metabolismo , Proteínas Morfogenéticas Óseas/metabolismo , Línea Celular , Línea Celular Tumoral , Femenino , Regulación del Desarrollo de la Expresión Génica/fisiología , Células HEK293 , Células Hep G2 , Humanos , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Células MCF-7 , Masculino , Ratones , Ratones Endogámicos C3H , Ratones Endogámicos C57BL
18.
Mol Cell ; 81(2): 304-322.e16, 2021 01 21.
Artículo en Inglés | MEDLINE | ID: mdl-33357414

RESUMEN

Protein synthesis must be finely tuned in the developing nervous system as the final essential step of gene expression. This study investigates the architecture of ribosomes from the neocortex during neurogenesis, revealing Ebp1 as a high-occupancy 60S peptide tunnel exit (TE) factor during protein synthesis at near-atomic resolution by cryoelectron microscopy (cryo-EM). Ribosome profiling demonstrated Ebp1-60S binding is highest during start codon initiation and N-terminal peptide elongation, regulating ribosome occupancy of these codons. Membrane-targeting domains emerging from the 60S tunnel, which recruit SRP/Sec61 to the shared binding site, displace Ebp1. Ebp1 is particularly abundant in the early-born neural stem cell (NSC) lineage and regulates neuronal morphology. Ebp1 especially impacts the synthesis of membrane-targeted cell adhesion molecules (CAMs), measured by pulsed stable isotope labeling by amino acids in cell culture (pSILAC)/bioorthogonal noncanonical amino acid tagging (BONCAT) mass spectrometry (MS). Therefore, Ebp1 is a central component of protein synthesis, and the ribosome TE is a focal point of gene expression control in the molecular specification of neuronal morphology during development.


Asunto(s)
Proteínas de Unión al ADN/genética , Regulación del Desarrollo de la Expresión Génica , Neocórtex/metabolismo , Neuronas/metabolismo , Biosíntesis de Proteínas , Proteostasis/genética , Proteínas de Unión al ARN/genética , Subunidades Ribosómicas Grandes de Eucariotas/genética , Animales , Animales Recién Nacidos , Sitios de Unión , Moléculas de Adhesión Celular Neuronal/química , Moléculas de Adhesión Celular Neuronal/genética , Moléculas de Adhesión Celular Neuronal/metabolismo , Línea Celular Tumoral , Microscopía por Crioelectrón , Proteínas de Unión al ADN/química , Proteínas de Unión al ADN/metabolismo , Embrión de Mamíferos , Femenino , Masculino , Ratones , Neocórtex/citología , Neocórtex/crecimiento & desarrollo , Células-Madre Neurales/citología , Células-Madre Neurales/metabolismo , Neurogénesis/genética , Neuronas/citología , Cultivo Primario de Células , Unión Proteica , Conformación Proteica en Hélice alfa , Conformación Proteica en Lámina beta , Dominios y Motivos de Interacción de Proteínas , Proteínas de Unión al ARN/química , Proteínas de Unión al ARN/metabolismo , Subunidades Ribosómicas Grandes de Eucariotas/metabolismo , Subunidades Ribosómicas Grandes de Eucariotas/ultraestructura , Partícula de Reconocimiento de Señal/química , Partícula de Reconocimiento de Señal/genética , Partícula de Reconocimiento de Señal/metabolismo
19.
Gene ; 772: 145349, 2021 Mar 10.
Artículo en Inglés | MEDLINE | ID: mdl-33338511

RESUMEN

Inverted cuttings of Populus yunnanensis are characterized by enlarged stems and dwarfed new shoots, and phytohormones play a crucial role in the response to inversion. The polar auxin transport (PAT) system is distinct from the transport systems of other hormones and is controlled by three major transporter gene families: pin-formed (PIN), auxin-resistant/like aux (AUX/LAX) and ATP-binding cassette transporters of the B class (ABCB). Here, we identified these three families in P. trichocarpa, P. euphratica and P. yunnanensis through a genome-wide analysis. The Populus PIN, AUX/LAX and ABCB gene families comprised 15, 8 and 31 members, respectively. Most PAT genes in Populus and Arabidopsis were identified as clear sister pairs, and some had unique motifs. Transcriptome profiling revealed that the expression of most PAT genes was unrelated to cutting inversion and that only several genes showed altered expression when cuttings were inverted. The auxin content difference at positions was opposite in upright and inverted cutting bodies during rooting, which obeyed the original plant polarity. However, during plant growth, the two direction types exhibited similar auxin movements in the cutting bodies, and the opposite auxin changes were observed in new shoots. Four PAT genes with a positive response to cutting inversion, PyuPIN10, PyuPIN11, PyuLAX6 and PyuABCB27, showed diverse expression patterns between upright and inverted cuttings during rooting and plant growth. Furthermore, PAT gene expression retained its polarity, which differs from the results found for auxin flow during plant growth. The inconformity indicated that a new downward auxin flow in addition to the old upward flow might be established during the growth of inverted cuttings. Some highly polar PAT genes were involved in the maintenance of original auxin polarity, which might cause the enlarged stems of inverted cuttings. This work lays a foundation for understanding the roles of auxin transport in plant responses to inversion.


Asunto(s)
Perfilación de la Expresión Génica/métodos , Ácidos Indolacéticos/metabolismo , Proteínas de Transporte de Membrana/genética , Populus/crecimiento & desarrollo , Regulación del Desarrollo de la Expresión Génica , Regulación de la Expresión Génica de las Plantas , Proteínas de Transporte de Membrana/metabolismo , Familia de Multigenes , Filogenia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Populus/clasificación , Populus/genética , Populus/metabolismo , Secuenciación Completa del Genoma
20.
Gene ; 772: 145355, 2021 Mar 10.
Artículo en Inglés | MEDLINE | ID: mdl-33340562

RESUMEN

Calanthe tsoongiana is a rare orchid species native to China. Asymbiotic seed germination is of great importance in the ex situ conservation of this species. Based on morphological characteristics and anatomical structures, the C. tsoongiana developmental process from seeds to seedlings was divided into four stages (SA, PB, PC and PD), and subsequently, changes in endogenous hormone contents and gene expression were assessed using RNA-seq analysis. K-means analysis divided the DEGs into eight clusters. The gene expression decreased markedly between the imbibed seed and globular protocorm stages, with this being the most notably enriched cluster. During the seed germination period, DEGs were dominated by ATP metabolic processes, respiration and photosynthesis. A small change in gene expression was found in the globular protocorm versus the finger-like protocorm stages. During the last developmental stage, DEGs were significantly enriched in lignin catabolic processes and plant-type secondary cell wall biogenesis. DEG homologs, such as TSA1, DAO, NCED1, STM, and CUC2, were related to phytohormones and the morphogenesis of shoots, leaves and roots. Particularly, interactions between CUC2 and STM as well as AS1 and STM were likely involved in protocorm formation and development. Furthermore, TSA1 and DAO were distinctly validated and implicated in the synthesis and metabolism of auxin, which has a pivotal role in plant development. Our study is the first to combine morphological and transcriptome analysis to examine the process of protocorm formation and development. The results provide a foundation for understanding the mechanisms of seed germination and protocorm development of C. tsoongiana.


Asunto(s)
Perfilación de la Expresión Génica/métodos , Orchidaceae/crecimiento & desarrollo , Proteínas de Plantas/genética , Regulación del Desarrollo de la Expresión Génica , Regulación de la Expresión Génica de las Plantas , Germinación , Orchidaceae/genética , Fotosíntesis , Semillas/genética , Semillas/crecimiento & desarrollo , Análisis de Secuencia de ARN
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