Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 25
Filtrar
1.
PLoS Genet ; 8(3): e1002609, 2012.
Artículo en Inglés | MEDLINE | ID: mdl-22479204

RESUMEN

Insulin-like growth factor I receptor (Igf1r) signaling controls proliferation, differentiation, growth, and cell survival in many tissues; and its deregulated activity is involved in tumorigenesis. Although important during fetal growth and postnatal life, a function for the Igf pathway during preimplantation development has not been described. We show that abrogating Igf1r signaling with specific inhibitors blocks trophectoderm formation and compromises embryo survival during murine blastocyst formation. In normal embryos total Igf1r is present throughout the membrane, whereas the activated form is found exclusively at cell contact sites, colocalizing with E-cadherin. Using genetic domain switching, we show a requirement for E-cadherin to maintain proper activation of Igf1r. Embryos expressing exclusively a cadherin chimera with N-cadherin extracellular and E-cadherin intracellular domains (NcEc) fail to form a trophectoderm and cells die by apoptosis. In contrast, homozygous mutant embryos expressing a reverse-structured chimera (EcNc) show trophectoderm survival and blastocoel cavitation, indicating a crucial and non-substitutable role of the E-cadherin ectodomain for these processes. Strikingly, blastocyst formation can be rescued in homozygous NcEc embryos by restoring Igf1r signaling, which enhances cell survival. Hence, perturbation of E-cadherin extracellular integrity, independent of its cell-adhesion function, blocked Igf1r signaling and induced cell death in the trophectoderm. Our results reveal an important and yet undiscovered function of Igf1r during preimplantation development mediated by a unique physical interaction between Igf1r and E-cadherin indispensable for proper receptor activation and anti-apoptotic signaling. We provide novel insights into how ligand-dependent Igf1r activity is additionally gated to sense developmental potential in utero and into a bifunctional role of adhesion molecules in contact formation and signaling.


Asunto(s)
Blastocisto , Cadherinas , Desarrollo Embrionario/genética , Receptor IGF Tipo 1 , Transducción de Señal/genética , Animales , Apoptosis , Blastocisto/citología , Blastocisto/metabolismo , Cadherinas/genética , Cadherinas/metabolismo , Adhesión Celular/genética , Comunicación Celular/genética , Supervivencia Celular , Células Madre Embrionarias , Regulación del Desarrollo de la Expresión Génica , Homocigoto , Ligandos , Ratones , Mutación , Estructura Terciaria de Proteína/genética , Receptor IGF Tipo 1/genética , Receptor IGF Tipo 1/metabolismo
2.
Commun Biol ; 7(1): 31, 2024 01 05.
Artículo en Inglés | MEDLINE | ID: mdl-38182651

RESUMEN

The stability of cellular phenotypes in developing organisms depends on error-free transmission of epigenetic and genetic information during mitosis. Methylation of cytosine residues in genomic DNA is a key epigenetic mark that modulates gene expression and prevents genome instability. Here, we report on a genetic test of the relationship between DNA replication and methylation in the context of the developing vertebrate organism instead of cell lines. Our analysis is based on the identification of hypomorphic alleles of dnmt1, encoding the DNA maintenance methylase Dnmt1, and pole1, encoding the catalytic subunit of leading-strand DNA polymerase epsilon holoenzyme (Pole). Homozygous dnmt1 mutants exhibit genome-wide DNA hypomethylation, whereas the pole1 mutation is associated with increased DNA methylation levels. In dnmt1/pole1 double-mutant zebrafish larvae, DNA methylation levels are restored to near normal values, associated with partial rescue of mutant-associated transcriptional changes and phenotypes. Hence, a balancing antagonism between DNA replication and maintenance methylation buffers against replicative errors contributing to the robustness of vertebrate development.


Asunto(s)
Metilación de ADN , Pez Cebra , Animales , Pez Cebra/genética , Alelos , ADN , Epigénesis Genética
3.
Proc Natl Acad Sci U S A ; 107(38): 16613-8, 2010 Sep 21.
Artículo en Inglés | MEDLINE | ID: mdl-20823228

RESUMEN

The thymus is essential for T-cell development. Here, we focus on the role of the transcription factor Foxn1 in the development and function of thymic epithelial cells (TECs) of the mouse. TECs are of endodermal origin; they initially express Foxn1 and give rise to orthotopic (thoracic) and additional (cervical) thymi. Using Foxn1-directed cytoablation, we show that during embryogenesis, cervical thymi develop a few days after the thoracic lobes, and that bipotent epithelial progenitors of cortical and medullary compartments express Foxn1. We also show that following acute selective near-total ablation during embryogenesis, complete regeneration of TECs does not occur, providing an animal model for human thymic aplasia syndromes. Finally, we address the functional role of Foxn1-negative TECs that arise postnatally in the mouse. Lineage tracing shows that such Foxn1-negative TECs are descendants of Foxn1-positive progenitors; furthermore, Foxn1-directed subacute intoxication of TECs by polyglutamine-containing EGFP proteins indicates that a presumptive Foxn1-independent lineage does not contribute to thymopoietic function of the adult thymus. Our findings therefore support the notion that Foxn1 is the essential transcription factor regulating the differentiation of TECs and that its expression marks the major functional lineage of TECs in embryonic and adult thymic tissue.


Asunto(s)
Factores de Transcripción Forkhead/metabolismo , Linfopoyesis/fisiología , Timo/embriología , Animales , Secuencia de Bases , Cartilla de ADN/genética , Células Madre Embrionarias/citología , Células Madre Embrionarias/metabolismo , Células Epiteliales/citología , Células Epiteliales/metabolismo , Femenino , Factores de Transcripción Forkhead/deficiencia , Factores de Transcripción Forkhead/genética , Regulación del Desarrollo de la Expresión Génica , Humanos , Linfopoyesis/genética , Ratones , Ratones Noqueados , Ratones Transgénicos , Embarazo , Timo/anomalías , Timo/citología , Timo/metabolismo
4.
Nat Cell Biol ; 25(9): 1265-1278, 2023 09.
Artículo en Inglés | MEDLINE | ID: mdl-37652981

RESUMEN

Despite advances in four-factor (4F)-induced reprogramming (4FR) in vitro and in vivo, how 4FR interconnects with senescence remains largely under investigated. Here, using genetic and chemical approaches to manipulate senescent cells, we show that removal of p16High cells resulted in the 4FR of somatic cells into totipotent-like stem cells. These cells expressed markers of both pluripotency and the two-cell embryonic state, readily formed implantation-competent blastoids and, following morula aggregation, contributed to embryonic and extraembryonic lineages. We identified senescence-dependent regulation of nicotinamide N-methyltransferase as a key mechanism controlling the S-adenosyl-L-methionine levels during 4FR that was required for expression of the two-cell genes and acquisition of an extraembryonic potential. Importantly, a partial 4F epigenetic reprogramming in old mice was able to reverse several markers of liver aging only in conjunction with the depletion of p16High cells. Our results show that the presence of p16High senescent cells limits cell plasticity, whereas their depletion can promote a totipotent-like state and histopathological tissue rejuvenation during 4F reprogramming.


Asunto(s)
Plasticidad de la Célula , Reprogramación Celular , Animales , Ratones , Reprogramación Celular/genética , Envejecimiento/genética , Implantación del Embrión , Epigenómica
5.
Am J Pathol ; 177(2): 840-53, 2010 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-20558576

RESUMEN

Tissue inhibitors of metalloproteinases (TIMPs) are a family of closely related proteins that inhibit matrix metalloproteinases (MMPs). In the central nervous system (CNS), TIMPs 2, 3, and 4 are constitutively expressed at high levels, whereas TIMP1 can be induced by various stimuli. Here, we studied the effects of constitutive expression of TIMP1 in the CNS in transgenic mice. Transgene expression started prenatally and persisted throughout lifetime at high levels. Since MMP activity has been implicated in CNS development, in proper function of the adult CNS, and in inflammatory disorders, we investigated Timp1-induced CNS alterations. Despite sufficient MMP inhibition, high expressor transgenic mice had a normal phenotype. The absence of compensatory up-regulation of MMP genes in the CNS of Timp1 transgenic mice indicates that development, learning, and memory functions do not require the entire MMP arsenal. To elucidate the effects of strong Timp1 expression in CNS inflammation, we induced experimental allergic encephalomyelitis. We observed a Timp1 dose-dependent mitigation of both experimental allergic encephalomyelitis symptoms and histological lesions in the CNS of transgenic mice. All in all, our data demonstrate that (1) long-term CNS expression of TIMP1 with complete suppression of gelatinolytic activity does not interfere with physiological brain function and (2) TIMP1 might constitute a promising candidate for long-term therapeutic treatment of inflammatory CNS diseases such as multiple sclerosis.


Asunto(s)
Sistema Nervioso Central/metabolismo , Encefalomielitis Autoinmune Experimental/metabolismo , Inhibidor Tisular de Metaloproteinasa-1/metabolismo , Animales , Conducta Animal/fisiología , Sistema Nervioso Central/patología , Femenino , Masculino , Metaloproteinasas de la Matriz/genética , Metaloproteinasas de la Matriz/metabolismo , Ratones , Ratones Transgénicos , Pruebas Neuropsicológicas , Embarazo , Inhibidor Tisular de Metaloproteinasa-1/genética
7.
Proc Natl Acad Sci U S A ; 105(13): 5177-82, 2008 Apr 01.
Artículo en Inglés | MEDLINE | ID: mdl-18362329

RESUMEN

The giant cytosolic protease tripeptidyl peptidase II (TPPII) has been implicated in the regulation of proliferation and survival of malignant cells, particularly lymphoma cells. To address its functions in normal cellular and systemic physiology we have generated TPPII-deficient mice. TPPII deficiency activates cell type-specific death programs, including proliferative apoptosis in several T lineage subsets and premature cellular senescence in fibroblasts and CD8(+) T cells. This coincides with up-regulation of p53 and dysregulation of NF-kappaB. Prominent degenerative alterations at the organismic level were a decreased lifespan and symptoms characteristic of immunohematopoietic senescence. These symptoms include accelerated thymic involution, lymphopenia, impaired proliferative T cell responses, extramedullary hematopoiesis, and inflammation. Thus, TPPII is important for maintaining normal cellular and systemic physiology, which may be relevant for potential therapeutic applications of TPPII inhibitors.


Asunto(s)
Envejecimiento/inmunología , Apoptosis/inmunología , Serina Endopeptidasas/deficiencia , Serina Endopeptidasas/metabolismo , Aminopeptidasas , Animales , Diferenciación Celular/inmunología , Células Cultivadas , Dipeptidil-Peptidasas y Tripeptidil-Peptidasas , Fibroblastos , Eliminación de Gen , Linfopenia/enzimología , Linfopenia/genética , Linfopenia/patología , Ratones , Ratones Noqueados , FN-kappa B/metabolismo , Fenotipo , Serina Endopeptidasas/genética , Linfocitos T/citología , Linfocitos T/enzimología , Linfocitos T/inmunología , Timo/citología , Timo/enzimología , Timo/inmunología , Proteína p53 Supresora de Tumor/metabolismo
8.
Dev Cell ; 3(2): 171-81, 2002 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-12194849

RESUMEN

Using Cre/loxP, we conditionally inactivated the beta-catenin gene in cells of structures that exhibit important embryonic organizer functions: the visceral endoderm, the node, the notochord, and the definitive endoderm. Mesoderm formation was not affected in the mutant embryos, but the node was missing, patterning of the head and trunk was affected, and no notochord or somites were formed. Surprisingly, deletion of beta-catenin in the definitive endoderm led to the formation of multiple hearts all along the anterior-posterior (A/P) axis of the embryo. Ectopic hearts developed in parallel with the normal heart in regions of ectopic Bmp2 expression. We provide evidence that ablation of beta-catenin in embryonic endoderm changes cell fate from endoderm to precardiac mesoderm, consistent with the existence of bipotential mesendodermal progenitors in mouse embryos.


Asunto(s)
Diferenciación Celular/genética , Linaje de la Célula/genética , Coristoma/genética , Proteínas del Citoesqueleto/deficiencia , Embrión de Mamíferos/anomalías , Eliminación de Gen , Cardiopatías Congénitas/genética , Ratones Noqueados/anomalías , Transactivadores/deficiencia , Animales , Tipificación del Cuerpo/genética , Células Cultivadas , Quimera/anomalías , Quimera/genética , Quimera/metabolismo , Coristoma/metabolismo , Coristoma/fisiopatología , Citocinas , Proteínas del Citoesqueleto/genética , Proteínas de Unión al ADN/metabolismo , Embrión de Mamíferos/citología , Embrión de Mamíferos/metabolismo , Endodermo/citología , Endodermo/metabolismo , Femenino , Factor de Transcripción GATA4 , Regulación del Desarrollo de la Expresión Génica/fisiología , Cabeza/anomalías , Cardiopatías Congénitas/metabolismo , Cardiopatías Congénitas/fisiopatología , Proteínas de Homeodominio/metabolismo , Péptidos y Proteínas de Señalización Intercelular , Mesodermo/citología , Mesodermo/metabolismo , Ratones , Ratones Noqueados/genética , Ratones Noqueados/metabolismo , Mutación/genética , Proteínas/metabolismo , Transactivadores/genética , Factores de Transcripción/metabolismo , beta Catenina
9.
Dev Cell ; 4(3): 395-406, 2003 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-12636920

RESUMEN

The vertebral column derives from somites generated by segmentation of presomitic mesoderm (PSM). Somitogenesis involves a molecular oscillator, the segmentation clock, controlling periodic Notch signaling in the PSM. Here, we establish a novel link between Wnt/beta-catenin signaling and the segmentation clock. Axin2, a negative regulator of the Wnt pathway, is directly controlled by Wnt/beta-catenin and shows oscillating expression in the PSM, even when Notch signaling is impaired, alternating with Lfng expression. Moreover, Wnt3a is required for oscillating Notch signaling activity in the PSM. We propose that the segmentation clock is established by Wnt/beta-catenin signaling via a negative-feedback mechanism and that Wnt3a controls the segmentation process in vertebrates.


Asunto(s)
Relojes Biológicos/fisiología , Tipificación del Cuerpo/fisiología , Embrión de Mamíferos/embriología , Embrión no Mamífero , Proteínas/metabolismo , Somitos/metabolismo , Columna Vertebral/embriología , Vertebrados/embriología , Animales , Proteína Axina , Relojes Biológicos/genética , Proteínas del Citoesqueleto/genética , Proteínas del Citoesqueleto/metabolismo , Embrión de Mamíferos/metabolismo , Retroalimentación/fisiología , Feto , Regulación del Desarrollo de la Expresión Génica/genética , Glicosiltransferasas/genética , Glicosiltransferasas/metabolismo , Péptidos y Proteínas de Señalización Intracelular , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Mesodermo/citología , Mesodermo/metabolismo , Ratones , Ratones Transgénicos , Proteínas/genética , Transducción de Señal/genética , Somitos/citología , Columna Vertebral/citología , Columna Vertebral/metabolismo , Transactivadores/genética , Transactivadores/metabolismo , Vertebrados/metabolismo , Proteínas Wnt , Proteína Wnt3 , Proteína Wnt3A , beta Catenina
11.
Sci Rep ; 8(1): 11095, 2018 07 23.
Artículo en Inglés | MEDLINE | ID: mdl-30038304

RESUMEN

The numbers of thymic epithelial cells (TECs) and thymocytes steadily increase during embryogenesis. To examine this dynamic, we generated several TEC-specific transgenic mouse lines, which express fluorescent proteins in the nucleus, the cytosol and in the membranes under the control of the Foxn1 promoter. These tools enabled us to determine TEC numbers in tissue sections by confocal fluorescent microscopy, and in the intact organ by light-sheet microscopy. Compared to histological procedures, flow cytometric analysis of thymic cellularity is shown to underestimate the numbers of TECs by one order of magnitude; using enzymatic digestion of thymic tissue, the loss of cortical TECs (cTECs) is several fold greater than that of medullary TECs (mTECs), although different cTEC subsets appear to be still present in the final preparation. Novel reporter lines driven by Psmb11 and Prss16 promoters revealed the trajectory of differentiation of cTEC-like cells, and, owing to the additional facility of conditional cell ablation, allowed us to follow the recovery of such cells after their depletion during embryogenesis. Multiparametric histological analyses indicate that the new transgenic reporter lines not only reveal the unique morphologies of different TEC subsets, but are also conducive to the analysis of the complex cellular interactions in the thymus.


Asunto(s)
Epitelio/embriología , Timo/embriología , Animales , Comunicación Celular , Microambiente Celular , Células Epiteliales/citología , Células Epiteliales/metabolismo , Epitelio/metabolismo , Expresión Génica , Genes Reporteros , Ratones Transgénicos , Células del Estroma/citología , Células del Estroma/metabolismo , Timo/metabolismo
12.
Cell Rep ; 18(12): 2815-2824, 2017 03 21.
Artículo en Inglés | MEDLINE | ID: mdl-28329675

RESUMEN

Wnt/ß-catenin signaling is required for embryonic stem cell (ESC) pluripotency by inducing mesodermal differentiation and inhibiting neuronal differentiation; however, how ß-catenin counter-regulates these differentiation pathways is unknown. Here, we show that lysine 49 (K49) of ß-catenin is trimethylated (ß-catMe3) by Ezh2 or acetylated (ß-catAc) by Cbp. Significantly, ß-catMe3 acts as a transcriptional co-repressor of the neuronal differentiation genes sox1 and sox3, whereas ß-catAc acts as a transcriptional co-activator of the key mesodermal differentiation gene t-brachyury (t-bra). Furthermore, ß-catMe3 and ß-catAc are alternatively enriched on repressed or activated genes, respectively, during ESC and adult stem cell differentiation into neuronal or mesodermal progenitor cell lineages. Importantly, expression of a ß-catenin K49A mutant results in major defects in ESC differentiation. We conclude that ß-catenin K49 trimethylation and acetylation are key elements in regulating ESC pluripotency and differentiation potential.


Asunto(s)
Células Madre Embrionarias/citología , Células Madre Embrionarias/metabolismo , Lisina/metabolismo , Células Madre Pluripotentes/citología , Células Madre Pluripotentes/metabolismo , beta Catenina/química , beta Catenina/metabolismo , Acetilación , Animales , Anticuerpos Monoclonales/metabolismo , Diferenciación Celular/genética , Proteína Potenciadora del Homólogo Zeste 2/metabolismo , Regulación de la Expresión Génica , Metilación , Mutación/genética , Complejo Represivo Polycomb 2/metabolismo , Regiones Promotoras Genéticas/genética , Ratas Endogámicas Lew
13.
Mech Dev ; 119(2): 127-35, 2002 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-12464426

RESUMEN

Neural crest cells are essential for proper development of a variety of tissues and structures, including peripheral and autonomic nervous systems, facial skeleton, aortic arches and pharyngeal glands like the thymus and parathyroids. Previous work has shown that bone morphogenic protein (BMP) signalling is required for the production of migratory neural crest cells that contribute to the neurogenic and skeletogenic lineages. We show here that BMP-dependent neural crest cells are also required for development of the embryonic aortic arches and pharynx-derived glands. Blocking formation or migration of this crest cell population from the caudal hindbrain resulted in strong phenotypes in the cardiac outflow tract and the thymus. Thymic aplasia or hypoplasia occurs despite uncompromised gene induction in the pharyngeal endoderm. In addition, when hypoplastic thymic tissue is found, it is ectopically located, but functional in thymopoiesis. Our data indicate that thymic phenotypes produced by neural crest deficits result from aberrant formation of pharyngeal pouches and impaired migration of thymic primordia because the mesenchymal content in the branchial arches is below a threshold level.


Asunto(s)
Aorta Torácica/embriología , Proteínas Morfogenéticas Óseas/metabolismo , Cresta Neural/embriología , Animales , Animales Modificados Genéticamente , Linaje de la Célula , Movimiento Celular , ADN Complementario/metabolismo , Síndrome de DiGeorge/genética , Regulación hacia Abajo , Biblioteca de Genes , Humanos , Hibridación in Situ , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Modelos Biológicos , Mutación , Cresta Neural/metabolismo , Fenotipo , Transducción de Señal , Timo/embriología , Timo/metabolismo , Factores de Tiempo , Xenopus
14.
Mech Dev ; 120(12): 1423-32, 2003 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-14654215

RESUMEN

We report that gene silencing via intracytoplasmic microinjections of morpholino-modified antisense oligonucleotides is an effective and reproducible method to study both maternal and zygotic gene functions during early and late stages of mouse preimplantation development. The zygotic expression of the beta-geo transgene in the ROSA26 mouse strain could be inhibited until at least the early blastula stages. Thus morpholino-triggered gene inactivation appears to be a useful method to study the functional role of genes in preimplantation development. Using this approach, we have investigated a potential role of maternal expression of Cdh1, the gene encoding the cell-adhesion molecule E-cadherin. Inhibition of translation of maternal E-cadherin mRNA causes a developmental arrest at the two-cell stage. BrUTP incorporation assays indicated that this developmental defect cannot be explained by a general failure in transcriptional activity. This defect is reversible since E-cadherin mRNA can rescue the affected embryos, suggesting that a functional adhesion complex, present at the junction between blastomeres, is a prerequisite for the normal development of the mouse preimplantation embryo. Our study thus reveals a previously unanticipated role of maternal E-cadherin during early stages of mouse development.


Asunto(s)
Cadherinas/genética , Cadherinas/metabolismo , Regulación hacia Abajo/genética , Desarrollo Embrionario y Fetal , Oligonucleótidos Antisentido/genética , Animales , Adhesión Celular , Embrión de Mamíferos/embriología , Embrión de Mamíferos/metabolismo , Femenino , Ratones , Oocitos/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Transcripción Genética/genética , Transgenes/genética
15.
PLoS One ; 7(4): e35322, 2012.
Artículo en Inglés | MEDLINE | ID: mdl-22530006

RESUMEN

Establishment of totipotency after somatic cell nuclear transfer (NT) requires not only reprogramming of gene expression, but also conversion of the cell cycle from quiescence to the precisely timed sequence of embryonic cleavage. Inadequate adaptation of the somatic nucleus to the embryonic cell cycle regime may lay the foundation for NT embryo failure and their reported lower cell counts. We combined bright field and fluorescence imaging of histone H(2b)-GFP expressing mouse embryos, to record cell divisions up to the blastocyst stage. This allowed us to quantitatively analyze cleavage kinetics of cloned embryos and revealed an extended and inconstant duration of the second and third cell cycles compared to fertilized controls generated by intracytoplasmic sperm injection (ICSI). Compared to fertilized embryos, slow and fast cleaving NT embryos presented similar rates of errors in M phase, but were considerably less tolerant to mitotic errors and underwent cleavage arrest. Although NT embryos vary substantially in their speed of cell cycle progression, transcriptome analysis did not detect systematic differences between fast and slow NT embryos. Profiling of amino acid turnover during pre-implantation development revealed that NT embryos consume lower amounts of amino acids, in particular arginine, than fertilized embryos until morula stage. An increased arginine supplementation enhanced development to blastocyst and increased embryo cell numbers. We conclude that a cell cycle delay, which is independent of pluripotency marker reactivation, and metabolic restraints reduce cell counts of NT embryos and impede their development.


Asunto(s)
Ciclo Celular/fisiología , Reprogramación Celular/fisiología , Aminoácidos/metabolismo , Animales , Arginina/metabolismo , Blastocisto/metabolismo , Análisis por Conglomerados , Daño del ADN , Transferencia de Embrión , Embrión de Mamíferos/citología , Embrión de Mamíferos/fisiología , Desarrollo Embrionario/fisiología , Femenino , Regulación del Desarrollo de la Expresión Génica , Cinética , Masculino , Ratones , Ratones Endogámicos C3H , Ratones Endogámicos C57BL , Técnicas de Transferencia Nuclear , Imagen de Lapso de Tiempo , Transgenes
16.
Genesis ; 45(7): 456-9, 2007 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-17607693

RESUMEN

The trophectoderm (TE) of blastocysts, the first epithelium established in mammalian development, (1) plays signaling, supportive, and patterning functions during preimplantation development, (2) ensures embryo implantation into the uterine wall, and (3) gives rise to extraembryonic tissues essential for embryo patterning and growth after implantation. We show that mouse TE, itself permissive to lentiviral (LV) infection, represents a robust nonpermeable physical barrier to the virus particles, thereby shielding the cells of the inner cell mass from viral infection. This LV feature will allow modulations of gene expression in a lineage-specific manner, thus having significant applications in mouse functional genetics.


Asunto(s)
Linaje de la Célula , Técnicas de Transferencia de Gen , Vectores Genéticos , Lentivirus , Animales , Femenino , Genes Reporteros , Masculino , Ratones , Ratones Endogámicos C57BL
17.
Development ; 134(1): 31-41, 2007 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-17138661

RESUMEN

During mammalian embryogenesis the trophectoderm represents the first epithelial structure formed. The cell adhesion molecule E-cadherin is ultimately necessary for the transition from compacted morula to the formation of the blastocyst to ensure correct establishment of adhesion junctions in the trophectoderm. Here, we analyzed to what extent E-cadherin confers unique adhesion and signaling properties in trophectoderm formation in vivo. Using a gene replacement approach, we introduced N-cadherin cDNA into the E-cadherin genomic locus. We show that the expression of N-cadherin driven from the E-cadherin locus reflects the expression pattern of endogenous E-cadherin. Heterozygous mice co-expressing E- and N-cadherin are vital and show normal embryonic development. Interestingly, N-cadherin homozygous mutant embryos phenocopy E-cadherin-null mutant embryos. Upon removal of the maternal E-cadherin, we demonstrate that N-cadherin is able to provide sufficient cellular adhesion to mediate morula compaction, but is insufficient for the subsequent formation of a fully polarized functional trophectoderm. When ES cells were isolated from N-cadherin homozygous mutant embryos and teratomas were produced, these ES cells differentiated into a large variety of tissue-like structures. Importantly, different epithelial-like structures expressing N-cadherin were formed, including respiratory epithelia, squamous epithelia with signs of keratinization and secretory epithelia with goblet cells. Thus, N-cadherin can maintain epithelia in differentiating ES cells, but not during the formation of the trophectoderm. Our results point to a specific and unique function for E-cadherin during mouse preimplantation development.


Asunto(s)
Blastocisto/fisiología , Cadherinas/genética , Cadherinas/metabolismo , Ectodermo/fisiología , Embrión de Mamíferos/metabolismo , Animales , Blastocisto/citología , Cadherinas/análisis , Adhesión Celular , Diferenciación Celular , Linaje de la Célula , Células Cultivadas , Cruzamientos Genéticos , Ectodermo/citología , Embrión de Mamíferos/citología , Embrión de Mamíferos/embriología , Células Madre Embrionarias/citología , Células Madre Embrionarias/metabolismo , Eliminación de Gen , Regulación del Desarrollo de la Expresión Génica , Inmunohistoquímica , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Microscopía por Video , beta-Galactosidasa/metabolismo
18.
J Immunol ; 178(4): 2241-8, 2007 Feb 15.
Artículo en Inglés | MEDLINE | ID: mdl-17277129

RESUMEN

Endoplasmic reticulum-associated aminopeptidase 1 (ERAP1) is involved in the final processing of endogenous peptides presented by MHC class I molecules to CTLs. We generated ERAP1-deficient mice and analyzed cytotoxic responses upon infection with three viruses, including lymphocytic choriomeningitis virus, which causes vigorous T cell activation and is controlled by CTLs. Despite pronounced effects on the presentation of selected epitopes, the in vivo cytotoxic response was altered for only one of several epitopes tested. Moreover, control of lymphocytic choriomeningitis virus was not impaired in the knockout mice. Thus, we conclude that lack of ERAP1 has little influence on antiviral immunohierarchies and antiviral immunity in the infections studied. We also focused on the role of ERAP1 in cross-presentation. We demonstrate that ERAP1 is required for efficient cross-presentation of cell-associated Ag and of OVA/anti-OVA immunocomplexes. Surprisingly, however, ERAP1 deficiency has no effect on cross-presentation of soluble OVA, suggesting that for soluble exogenous proteins, final processing may not take place in an environment containing active ERAP1.


Asunto(s)
Aminopeptidasas/inmunología , Presentación de Antígeno/inmunología , Infecciones por Arenaviridae/inmunología , Retículo Endoplásmico/inmunología , Virus de la Coriomeningitis Linfocítica/inmunología , Linfocitos T Citotóxicos/inmunología , Aminopeptidasas/deficiencia , Animales , Presentación de Antígeno/genética , Complejo Antígeno-Anticuerpo/genética , Complejo Antígeno-Anticuerpo/inmunología , Infecciones por Arenaviridae/genética , Reacciones Cruzadas/genética , Reacciones Cruzadas/inmunología , Retículo Endoplásmico/genética , Epítopos de Linfocito T/genética , Epítopos de Linfocito T/inmunología , Antígenos de Histocompatibilidad Clase I/genética , Antígenos de Histocompatibilidad Clase I/inmunología , Activación de Linfocitos/genética , Activación de Linfocitos/inmunología , Virus de la Coriomeningitis Linfocítica/genética , Ratones , Ratones Noqueados , Antígenos de Histocompatibilidad Menor
19.
Cell ; 125(5): 971-86, 2006 Jun 02.
Artículo en Inglés | MEDLINE | ID: mdl-16751105

RESUMEN

Vertebrate skeletogenesis involves two processes, skeletal patterning and osteoblast differentiation. Here, we show that Satb2, encoding a nuclear matrix protein, is expressed in branchial arches and in cells of the osteoblast lineage. Satb2-/- mice exhibit both craniofacial abnormalities that resemble those observed in humans carrying a translocation in SATB2 and defects in osteoblast differentiation and function. Multiple osteoblast-specific genes were identified as targets positively regulated by SATB2. In addition, SATB2 was found to repress the expression of several Hox genes including Hoxa2, an inhibitor of bone formation and regulator of branchial arch patterning. Molecular analysis revealed that SATB2 directly interacts with and enhances the activity of both Runx2 and ATF4, transcription factors that regulate osteoblast differentiation. This synergy was genetically confirmed by bone formation defects in Satb2/Runx2 and Satb2/Atf4 double heterozygous mice. Thus, SATB2 acts as a molecular node in a transcriptional network regulating skeletal development and osteoblast differentiation.


Asunto(s)
Anomalías Craneofaciales/genética , Huesos Faciales/anomalías , Proteínas de Unión a la Región de Fijación a la Matriz/genética , Osteoblastos/metabolismo , Osteogénesis/genética , Cráneo/anomalías , Factores de Transcripción/genética , Factor de Transcripción Activador 4/genética , Animales , Tipificación del Cuerpo/genética , Región Branquial/anomalías , Región Branquial/citología , Región Branquial/metabolismo , Diferenciación Celular/genética , Subunidad alfa 1 del Factor de Unión al Sitio Principal/genética , Anomalías Craneofaciales/metabolismo , Anomalías Craneofaciales/fisiopatología , Huesos Faciales/metabolismo , Regulación del Desarrollo de la Expresión Génica/genética , Genes Homeobox/genética , Proteínas de Homeodominio/genética , Ratones , Ratones Noqueados , Mutación/genética , Osteoblastos/citología , Proteínas Represoras/genética
20.
Dev Biol ; 291(2): 382-97, 2006 Mar 15.
Artículo en Inglés | MEDLINE | ID: mdl-16458883

RESUMEN

SOX9 is an evolutionary conserved transcription factor that is expressed in a variety of tissues, with essential functions in cartilage, testis, heart, glial cell, inner ear and neural crest development. By comparing human and pufferfish genomic sequences, we previously identified eight highly conserved sequence elements between 290 kb 5' and 450 kb 3' to human SOX9. In this study, we assayed the regulatory potential of elements E1 to E7 in transgenic mice using a lacZ reporter gene driven by a 529 bp minimal mouse Sox9 promoter. We found that three of these elements and the Sox9 promoter control distinct subsets of the tissue-specific expression pattern of Sox9. E3, located 251 kb 5' to SOX9, directs lacZ expression to cranial neural crest cells and to the inner ear. E1 is located 28 kb 5' to SOX9 and controls expression in the node, notochord, gut, bronchial epithelium and pancreas. Transgene expression in the neuroectoderm is mediated by E7, located 95 kb 3' to SOX9, which regulates expression in the telencephalon and midbrain, and by the Sox9 minimal promoter which controls expression in the ventral spinal cord and hindbrain. We show that E3-directed reporter gene expression in neural crest cells of the first but not of the second and third pharyngeal arch is dependent on beta-catenin, revealing a complex regulation of Sox9 in cranial neural crest cells. Moreover, we identify and discuss highly conserved transcription factor binding sites within enhancer E3 that are in good agreement with current models for neural crest and inner ear development. Finally, we identify enhancer E1 as a cis-regulatory element conserved between vertebrates and invertebrates, indicating that some cis-regulatory sequences that control developmental genes in vertebrates might be phylogenetically ancient.


Asunto(s)
Oído Interno/embriología , Elementos de Facilitación Genéticos/fisiología , Regulación del Desarrollo de la Expresión Génica , Proteínas del Grupo de Alta Movilidad/genética , Cresta Neural/embriología , Factores de Transcripción/genética , Animales , Secuencia de Bases , Sitios de Unión , Encéfalo/embriología , Encéfalo/metabolismo , Secuencia Conservada , Oído Interno/metabolismo , Humanos , Mucosa Intestinal/metabolismo , Ratones , Datos de Secuencia Molecular , Cresta Neural/metabolismo , Notocorda/metabolismo , Regiones Promotoras Genéticas , Factor de Transcripción SOX9 , beta Catenina/fisiología
SELECCIÓN DE REFERENCIAS
DETALLE DE LA BÚSQUEDA