RESUMEN
Hemogen is a hematopoietic tissue-specific gene that regulates the proliferation and differentiation of hematopoietic cells; however, the mechanism underlying its function in erythropoiesis is unknown. We found that depletion of hemogen in human CD34+ erythroid progenitor cells and HUDEP2 cells significantly reduced the expression of genes associated with heme and hemoglobin synthesis, supporting a positive role for hemogen in erythroid maturation. In human K562 cells, hemogen antagonized the occupancy of corepressors nucleosome remodeling and histone deacetylase (NuRD) complex and facilitated LDB1 complex-mediated chromatin looping. Hemogen recruited SWI/SNF complex ATPase BRG1 as a coactivator to regulate nucleosome accessibility and H3K27ac enrichment for promoter and enhancer activity. To determine whether hemogen/BRG1 cooperativity is conserved in mammalian systems, we generated hemogen-knockout/knockin mice and investigated hemogen/BRG1 function in murine erythropoiesis. Loss of hemogen in embryonic days 12.5 to 16.5 fetal liver cells impeded erythroid differentiation through reducing the production of mature erythroblasts. Chromatin immunoprecipitation sequencing in wild-type and hemogen-knockout animals revealed that BRG1 is largely dependent on hemogen to regulate chromatin accessibility at erythroid gene promoters and enhancers. In summary, the hemogen/BRG1 interaction in mammals is essential for fetal erythroid maturation and hemoglobin production through its active role in promoter and enhancer activity and chromatin organization.
Asunto(s)
ADN Helicasas , Eritropoyesis , Proteínas Nucleares , Nucleosomas , Factores de Transcripción , Animales , Cromatina/genética , ADN Helicasas/genética , ADN Helicasas/metabolismo , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Elementos de Facilitación Genéticos , Eritropoyesis/genética , Hemoglobinas/genética , Proteínas con Dominio LIM/genética , Proteínas con Dominio LIM/metabolismo , Ratones , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Regiones Promotoras Genéticas , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
Germ-cell transcription factors control gene networks that regulate oocyte differentiation and primordial follicle formation during early, postnatal mouse oogenesis. Taking advantage of gene-edited mice lacking transcription factors expressed in female germ cells, we analyzed global gene expression profiles in perinatal ovaries from wildtype, FiglaNull, Lhx8Null and Sohlh1Null mice. Figla deficiency dysregulates expression of meiosis-related genes (e.g. Sycp3, Rad51, Ybx2) and a variety of genes (e.g. Nobox, Lhx8, Taf4b, Sohlh1, Sohlh2, Gdf9) associated with oocyte growth and differentiation. The absence of FIGLA significantly impedes meiotic progression, causes DNA damage and results in oocyte apoptosis. Moreover, we find that FIGLA and other transcriptional regulator proteins (e.g. NOBOX, LHX8, SOHLH1, SOHLH2) are co-expressed in the same subset of germ cells in perinatal ovaries and Figla ablation dramatically disrupts KIT, NOBOX, LHX8, SOHLH1 and SOHLH2 abundance. In addition, not only do FIGLA, LHX8 and SOHLH1 cross-regulate each other, they also cooperate by direct interaction with each during early oocyte development and share downstream gene targets. Thus, our findings substantiate a major role for FIGLA, LHX8 and SOHLH1 as multifunctional regulators of networks necessary for oocyte maintenance and differentiation during early folliculogenesis.
Asunto(s)
Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Redes Reguladoras de Genes , Proteínas con Homeodominio LIM/metabolismo , Oocitos/metabolismo , Oogénesis/genética , Factores de Transcripción/metabolismo , Animales , Apoptosis , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Proliferación Celular/genética , Daño del ADN , Femenino , Regulación de la Expresión Génica , Células HEK293 , Humanos , Proteínas con Homeodominio LIM/genética , Meiosis/genética , Ratones , Oocitos/citología , Ovario/metabolismo , Factores de Transcripción/genéticaRESUMEN
PIWI-interacting RNAs (piRNAs) are small non-coding RNAs essential for animal germ cell development. Despite intense investigation of post-transcriptional processing, chromatin regulators for piRNA biogenesis in mammals remain largely unexplored. Here we document that BTBD18 is a pachytene nuclear protein in mouse testes that occupies a subset of pachytene piRNA-producing loci. Ablation of Btbd18 in mice disrupts piRNA biogenesis, prevents spermiogenesis, and results in male sterility. Transcriptome profiling, chromatin accessibility, and RNA polymerase II occupancy demonstrate that BTBD18 facilitates expression of pachytene piRNA precursors by promoting transcription elongation. Thus, our study identifies BTBD18 as a specific controller for transcription activation through RNA polymerase II elongation at a subset of genomic piRNA loci.
Asunto(s)
Sitios Genéticos , Proteínas Nucleares/metabolismo , ARN Interferente Pequeño/metabolismo , Elongación de la Transcripción Genética , Animales , Apoptosis/genética , Eliminación de Gen , Células Germinativas/citología , Células Germinativas/metabolismo , Masculino , Meiosis/genética , Ratones , Precursores del ARN/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Espermatogénesis/genética , Testículo/citologíaRESUMEN
Monospermic fertilization is mediated by the extracellular zona pellucida composed of ZP1, ZP2 and ZP3. Sperm bind to the N-terminus of ZP2 which is cleaved after fertilization by ovastacin (encoded by Astl) exocytosed from egg cortical granules to prevent sperm binding. AstlNull mice lack the post-fertilization block to sperm binding and the ability to rescue this phenotype with AstlmCherry transgenic mice confirms the role of ovastacin in providing a definitive block to polyspermy. During oogenesis, endogenous ovastacin traffics through the endomembrane system prior to storage in peripherally located cortical granules. Deletion mutants of ovastacinmCherry expressed in growing oocytes define a unique 7 amino acid motif near its N-terminus that is necessary and sufficient for cortical granule localization. Deletion of the 7 amino acids by CRISPR/Cas9 at the endogenous locus (AstlΔ) prevents cortical granule localization of ovastacin. The misdirected enzyme is present within the endomembrane system and ZP2 is prematurely cleaved. Sperm bind poorly to the zona pellucida of AstlΔ/Δ mice with partially cleaved ZP2 and female mice are sub-fertile.
Asunto(s)
Gránulos Citoplasmáticos/metabolismo , Fertilización , Metaloproteasas/metabolismo , Oocitos/metabolismo , Señales de Clasificación de Proteína , Glicoproteínas de la Zona Pelúcida/metabolismo , Animales , Femenino , Metaloproteasas/química , Metaloproteasas/genética , Ratones , Transporte de Proteínas , ProteolisisRESUMEN
Although many genes that specify neocortical projection neuron subtypes have been identified, the downstream effectors that control differentiation of those subtypes remain largely unknown. Here, we demonstrate that the LIM domain-binding proteins Ldb1 and Ldb2 exhibit dynamic and inversely correlated expression patterns during cerebral cortical development. Ldb1-deficient brains display severe defects in proliferation and changes in regionalization, phenotypes resembling those of Lhx mutants. Ldb2-deficient brains, on the other hand, exhibit striking phenotypes affecting layer 5 pyramidal neurons: Immature neurons have an impaired capacity to segregate into mature callosal and subcerebral projection neurons. The analysis of Ldb2 single-mutant mice reveals a compensatory role of Ldb1 for Ldb2 during corticospinal motor neuron (CSMN) differentiation. Animals lacking both Ldb1 and Ldb2 uncover the requirement for Ldb2 during CSMN differentiation, manifested as incomplete CSMN differentiation, and ultimately leading to a failure of the corticospinal tract.
Asunto(s)
Diferenciación Celular , Proteínas de Unión al ADN/deficiencia , Regulación del Desarrollo de la Expresión Génica/fisiología , Proteínas con Dominio LIM/deficiencia , Neuronas Motoras/metabolismo , Tractos Piramidales/metabolismo , Factores de Transcripción/deficiencia , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Animales , Diferenciación Celular/fisiología , Ratones Transgénicos , Neurogénesis/fisiología , Factores de Transcripción/metabolismoRESUMEN
LIM domain-binding protein 1 (Ldb1) is a nuclear cofactor that interacts with LIM homeodomain proteins to form multiprotein complexes that are important for transcription regulation. Ldb1 has been shown to play essential roles in various processes during mouse embryogenesis. To determine the role of Ldb1 in mid- and hindbrain development, we have generated a conditional mutant with a specific deletion of the Ldb1 in the Engrailed-1-expressing region of the developing mid- and hindbrain. Our study showed that the deletion impaired the expression of signaling molecules, such as fibroblast growth factor 8 (FGF8) and Wnt1, in the isthmic organizer and the expression of Shh in the ventral midbrain. The midbrain and the cerebellum were severely reduced in size, and the midbrain dopaminergic (mDA) neurons were missing in the mutant. These defects are identical to the phenotype that has been observed previously in mice with a deletion of the LIM homeodomain gene Lmx1b. Our results thus provide genetic evidence supporting that Ldb1 and Lmx1b function cooperatively to regulate mid- and hindbrain development. In addition, we found that mouse embryonic stem cells lacking Ldb1 failed to generate several types of differentiated neurons, including the mDA neurons, serotonergic neurons, cholinergic neurons, and olfactory bulb neurons, indicating an essential cell-autonomous role for Ldb1 in the development of these neurons.
Asunto(s)
Proteínas de Unión al ADN/metabolismo , Neuronas Dopaminérgicas/citología , Neuronas Dopaminérgicas/metabolismo , Proteínas con Dominio LIM/metabolismo , Mesencéfalo/citología , Organizadores Embrionarios/embriología , Organizadores Embrionarios/metabolismo , Animales , Biomarcadores/metabolismo , Diferenciación Celular , Cerebelo/embriología , Embrión de Mamíferos/metabolismo , Regulación del Desarrollo de la Expresión Génica , Mesencéfalo/embriología , Ratones , Células Madre Embrionarias de Ratones/citología , Células Madre Embrionarias de Ratones/metabolismo , Mutación/genéticaRESUMEN
The mamillary body (MM) is a group of hypothalamic nuclei related to memory and spatial navigation that interconnects hippocampal, thalamic, and tegmental regions. Here we demonstrate that Lhx5, a LIM-HD domain transcription factor expressed early in the developing posterior hypothalamus, is required for the generation of the MM and its derived tracts. The MM markers Foxb1, Sim2, and Lhx1 are absent in Lhx5 knock-out mice from early embryonic stages, suggesting abnormal specification of this region. This was supported by the absence of Nkx2.1 and expansion of Shh in the prospective mamillary area. Interestingly, we also found an ectopic domain expressing Lhx2 and Lhx9 along the anterio-posterior hypothalamic axis. Our results suggest that Lhx5 controls early aspects of hypothalamic development by regulating gene expression and cellular specification in the prospective MM.
RESUMEN
Cleft palate is a common birth defect in humans. Therefore, understanding the molecular genetics of palate development is important from both scientific and medical perspectives. Lhx6 and Lhx8 encode LIM homeodomain transcription factors, and inactivation of both genes in mice resulted in profound craniofacial defects including cleft secondary palate. The initial outgrowth of the palate was severely impaired in the mutant embryos, due to decreased cell proliferation. Through genome-wide transcriptional profiling, we discovered that p57(Kip2) (Cdkn1c), encoding a cell cycle inhibitor, was up-regulated in the prospective palate of Lhx6(-/-);Lhx8(-/-) mutants. p57(Kip2) has been linked to Beckwith-Wiedemann syndrome and IMAGe syndrome in humans, which are developmental disorders with increased incidents of palate defects among the patients. To determine the molecular mechanism underlying the regulation of p57(Kip2) by the Lhx genes, we combined chromatin immunoprecipitation, in silico search for transcription factor-binding motifs, and in vitro reporter assays with putative cis-regulatory elements. The results of these experiments indicated that LHX6 and LHX8 regulated p57(Kip2) via both direct and indirect mechanisms, with the latter mediated by Forkhead box (FOX) family transcription factors. Together, our findings uncovered a novel connection between the initiation of palate development and a cell cycle inhibitor via LHX. We propose a model in which Lhx6 and Lhx8 negatively regulate p57(Kip2) expression in the prospective palate area to allow adequate levels of cell proliferation and thereby promote normal palate development. This is the first report elucidating a molecular genetic pathway downstream of Lhx in palate development.
Asunto(s)
Inhibidor p57 de las Quinasas Dependientes de la Ciclina/genética , Regulación del Desarrollo de la Expresión Génica , Proteínas con Homeodominio LIM/genética , Proteínas del Tejido Nervioso/genética , Hueso Paladar/embriología , Hueso Paladar/metabolismo , Factores de Transcripción/genética , Animales , Proliferación Celular , Inhibidor p57 de las Quinasas Dependientes de la Ciclina/metabolismo , Factores de Transcripción Forkhead/genética , Factores de Transcripción Forkhead/metabolismo , Expresión Génica , Perfilación de la Expresión Génica , Estudio de Asociación del Genoma Completo , Genotipo , Humanos , Proteínas con Homeodominio LIM/metabolismo , Maxilar/embriología , Maxilar/metabolismo , Ratones , Mutación , Proteínas del Tejido Nervioso/metabolismo , Organogénesis/genética , Hueso Paladar/patología , Unión Proteica , Proteínas Represoras/genética , Proteínas Represoras/metabolismo , Factores de Transcripción/metabolismo , Activación TranscripcionalRESUMEN
BACKGROUND: LIM domain binding protein 1 (LDB1) is a transcriptional co-factor, which interacts with multiple transcription factors and other proteins containing LIM domains. Complete inactivation of Ldb1 in mice resulted in early embryonic lethality with severe patterning defects during gastrulation. Tissue-specific deletions using a conditional knockout allele revealed additional roles of Ldb1 in the development of the central nervous system, hematopoietic system, and limbs. The goal of the current study was to determine the importance of Ldb1 function during craniofacial development in mouse embryos. RESULTS: We generated tissue-specific Ldb1 mutants using Wnt1-Cre, which causes deletion of a floxed allele in the neural crest; neural crest-derived cells contribute to most of the mesenchyme of the developing face. All examined Wnt1-Cre;Ldb1(fl/-) mutants suffered from cleft secondary palate. Therefore, we performed a series of experiments to investigate how Ldb1 regulated palate development. First, we examined the expression of Ldb1 during normal development, and found that Ldb1 was expressed broadly in the palatal mesenchyme during early stages of palate development. Second, we compared the morphology of the developing palate in control and Ldb1 mutant embryos using sections. We found that the mutant palatal shelves had abnormally blunt appearance, and failed to elevate above the tongue at the posterior domain. An in vitro head culture experiment indicated that the elevation defect was not due to interference by the tongue. Finally, in the Ldb1 mutant palatal shelves, cell proliferation was abnormal in the anterior, and the expression of Wnt5a, Pax9 and Osr2, which regulate palatal shelf elevation, was also altered. CONCLUSIONS: The function of Ldb1 in the neural crest-derived palatal mesenchyme is essential for normal morphogenesis of the secondary palate.
Asunto(s)
Fisura del Paladar/genética , Proteínas de Unión al ADN/genética , Proteínas con Dominio LIM/genética , Cresta Neural/metabolismo , Hueso Paladar/metabolismo , Animales , Apoptosis/genética , Proliferación Celular , Células Cultivadas , Fisura del Paladar/embriología , Fisura del Paladar/metabolismo , Proteínas de Unión al ADN/metabolismo , Femenino , Técnica del Anticuerpo Fluorescente , Regulación del Desarrollo de la Expresión Génica , Hibridación in Situ , Proteínas con Dominio LIM/metabolismo , Mesodermo/embriología , Mesodermo/metabolismo , Mesodermo/patología , Ratones , Ratones Noqueados , Ratones Transgénicos , Cresta Neural/embriología , Cresta Neural/patología , Factor de Transcripción PAX9 , Factores de Transcripción Paired Box/genética , Factores de Transcripción Paired Box/metabolismo , Hueso Paladar/embriología , Hueso Paladar/patología , Embarazo , Factores de Tiempo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Proteínas Wnt/genética , Proteínas Wnt/metabolismo , Proteína Wnt-5a , Proteína Wnt1/genética , Proteína Wnt1/metabolismoRESUMEN
The progenitor zones of the embryonic mouse ventral telencephalon give rise to GABAergic and cholinergic neurons. We have shown previously that two LIM-homeodomain (LIM-HD) transcription factors, Lhx6 and Lhx8, that are downstream of Nkx2.1, are critical for the development of telencephalic GABAergic and cholinergic neurons. Here we investigate the role of Ldb1, a nuclear protein that binds directly to all LIM-HD factors, in the development of these ventral telencephalon derived neurons. We show that Ldb1 is expressed in the Nkx2.1 cell lineage during embryonic development and in mature neurons. Conditional deletion of Ldb1 causes defects in the expression of a series of genes in the ventral telencephalon and severe impairment in the tangential migration of cortical interneurons from the ventral telencephalon. Similar to the phenotypes observed in Lhx6 or Lhx8 mutant mice, the Ldb1 conditional mutants show a reduction in the number of both GABAergic and cholinergic neurons in the telencephalon. Furthermore, our analysis reveals defects in the development of the parvalbumin-positive neurons in the globus pallidus and striatum of the Ldb1 mutants. These results provide evidence that Ldb1 plays an essential role as a transcription co-regulator of Lhx6 and Lhx8 in the control of mammalian telencephalon development.
Asunto(s)
Neuronas Colinérgicas/metabolismo , Proteínas de Unión al ADN/metabolismo , Neuronas GABAérgicas/metabolismo , Proteínas con Dominio LIM/metabolismo , Proteínas Nucleares/metabolismo , Telencéfalo/embriología , Factores de Transcripción/metabolismo , Animales , Diferenciación Celular/genética , Linaje de la Célula/genética , Proteínas de Unión al ADN/genética , Desarrollo Embrionario/genética , Regulación del Desarrollo de la Expresión Génica , Globo Pálido/embriología , Proteínas Hedgehog/biosíntesis , Proteínas Hedgehog/metabolismo , Proteínas con Dominio LIM/genética , Proteínas con Homeodominio LIM/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Proteínas del Tejido Nervioso/metabolismo , Telencéfalo/crecimiento & desarrollo , Factor Nuclear Tiroideo 1RESUMEN
The accessory olfactory bulb (AOB) is a critical olfactory structure that has been implicated in mediating social behavior. It receives input from the vomeronasal organ and projects to targets in the amygdaloid complex. Its anterior and posterior components (aAOB and pAOB) display molecular, connectional and functional segregation in processing reproductive and defensive and aggressive behaviors, respectively. We observed a dichotomy in the development of the projection neurons of the aAOB and pAOB in mice. We found that they had distinct sites of origin and that different regulatory molecules were required for their specification and migration. aAOB neurons arose locally in the rostral telencephalon, similar to main olfactory bulb neurons. In contrast, pAOB neurons arose caudally, from the neuroepithelium of the diencephalic-telencephalic boundary, from which they migrated rostrally to reach their destination. This unusual origin and migration is conserved in Xenopus, providing an insight into the origin of a key component of this system in evolution.
Asunto(s)
Vías Aferentes/fisiología , Evolución Biológica , Movimiento Celular/fisiología , Regulación del Desarrollo de la Expresión Génica/fisiología , Neuronas/fisiología , Bulbo Olfatorio , Factores de Edad , Animales , Animales Recién Nacidos , Bromodesoxiuridina/metabolismo , Quinasa 5 Dependiente de la Ciclina/genética , Quinasa 5 Dependiente de la Ciclina/metabolismo , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Diencéfalo/citología , Diencéfalo/fisiología , Electroporación/métodos , Embrión de Mamíferos , Femenino , Proteínas Luminiscentes/genética , Masculino , Ratones , Ratones Transgénicos , Microinyecciones/métodos , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Proteínas Nucleares , Bulbo Olfatorio/citología , Bulbo Olfatorio/embriología , Bulbo Olfatorio/crecimiento & desarrollo , Oocitos , Técnicas de Cultivo de Órganos , Embarazo , Telencéfalo/citología , Telencéfalo/fisiología , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Proteína Tumoral p73 , Proteínas Supresoras de Tumor , Órgano Vomeronasal/citología , Órgano Vomeronasal/embriología , Órgano Vomeronasal/crecimiento & desarrollo , XenopusRESUMEN
BACKGROUND: Mice lacking the activities of Dlx1 and Dlx2 (Dlx1/2-/-) exhibit cleft palate, one of the most common human congenital defects, but the etiology behind this phenotype has been unknown. Therefore, we analyzed the morphological, cellular, and molecular changes caused by inactivation of Dlx1 and Dlx2 as related to palate development. RESULTS: Dlx1/2-/- mutants exhibited lack of vertical growth in the posterior palate during the earliest stage of palatogenesis. We attributed this growth deficiency to reduced cell proliferation. Expression of a cell cycle regulator Ccnd1 was specifically down-regulated in the same region. Previous studies established that the epithelial-mesenchymal signaling loop involving Shh, Bmp4, and Fgf10 is important for cell proliferation and tissue growth during palate development. This signaling loop was disrupted in Dlx1/2-/- palate. Interestingly, however, the decreases in Ccnd1 expression and mitosis in Dlx1/2-/- mutants were independent of this signaling loop. Finally, Dlx1/2 activity was required for normal expression of several transcription factor genes whose mutation results in palate defects. CONCLUSIONS: The functions of Dlx1 and Dlx2 are crucial for the initial formation of the posterior palatal shelves, and that the Dlx genes lie upstream of multiple signaling molecules and transcription factors important for later stages of palatogenesis.
Asunto(s)
Fisura del Paladar/metabolismo , Proteínas de Homeodominio/metabolismo , Hueso Paladar/embriología , Hueso Paladar/metabolismo , Factores de Transcripción/metabolismo , Animales , Proliferación Celular , Ciclina D1/genética , Ciclina D1/metabolismo , Regulación del Desarrollo de la Expresión Génica/genética , Regulación del Desarrollo de la Expresión Génica/fisiología , Proteínas de Homeodominio/genética , Ratones , Ratones Noqueados , Factores de Transcripción/genéticaRESUMEN
Lhx6 and Lhx8 transcription factor coexpression in early-born MGE neurons is required to induce neuronal Shh expression. We provide evidence that these transcription factors regulate expression of a Shh enhancer in MGE neurons. Lhx6 and Lhx8 are also required to prevent Nkx2-1 expression in a subset of pallial interneurons. Shh function in early-born MGE neurons was determined by genetically eliminating Shh expression in the MGE mantle zone (MZ). This mutant had reduced SHH signaling in the overlying progenitor zone, which led to reduced Lhx6, Lhx8, and Nkx2-1 expression in the rostrodorsal MGE and a preferential reduction of late-born somatostatin(+) and parvalbumin(+) cortical interneurons. Thus, Lhx6 and Lhx8 regulate MGE development through autonomous and nonautonomous mechanisms, the latter by promoting Shh expression in MGE neurons, which in turn feeds forward to promote the developmental program of the rostrodorsal MGE.
Asunto(s)
Regulación del Desarrollo de la Expresión Génica/fisiología , Cuerpos Geniculados/citología , Proteínas Hedgehog/metabolismo , Proteínas de Homeodominio/metabolismo , Interneuronas/fisiología , Proteínas del Tejido Nervioso/metabolismo , Células-Madre Neurales/fisiología , Factores de Edad , Animales , Animales Recién Nacidos , Diferenciación Celular , Proliferación Celular , Supervivencia Celular/genética , Células Cultivadas , Ensayo de Cambio de Movilidad Electroforética , Embrión de Mamíferos , Regulación del Desarrollo de la Expresión Génica/genética , Cuerpos Geniculados/embriología , Cuerpos Geniculados/crecimiento & desarrollo , Proteínas Hedgehog/genética , Proteínas de Homeodominio/genética , Proteínas con Homeodominio LIM , Ratones , Ratones Transgénicos , Mutación/genética , Proteínas del Tejido Nervioso/genética , Neuronas/fisiología , Proteínas Nucleares/metabolismo , Factor Nuclear Tiroideo 1 , Factores de Transcripción/metabolismoRESUMEN
The nuclear adaptor Ldb1 functions as a core component of multiprotein transcription complexes that regulate differentiation in diverse cell types. In the hematopoietic lineage, Ldb1 forms a complex with the non-DNA-binding adaptor Lmo2 and the transcription factors E2A, Scl and GATA-1 (or GATA-2). Here we demonstrate a critical and continuous requirement for Ldb1 in the maintenance of both fetal and adult mouse hematopoietic stem cells (HSCs). Deletion of Ldb1 in hematopoietic progenitors resulted in the downregulation of many transcripts required for HSC maintenance. Genome-wide profiling by chromatin immunoprecipitation followed by sequencing (ChIP-Seq) identified Ldb1 complex-binding sites at highly conserved regions in the promoters of genes involved in HSC maintenance. Our results identify a central role for Ldb1 in regulating the transcriptional program responsible for the maintenance of HSCs.
Asunto(s)
Células Madre Adultas/metabolismo , Proteínas de Unión al ADN/metabolismo , Células Madre Embrionarias/metabolismo , Desarrollo Fetal , Células Madre Hematopoyéticas/metabolismo , Traslado Adoptivo , Células Madre Adultas/citología , Células Madre Adultas/inmunología , Células Madre Adultas/trasplante , Animales , Diferenciación Celular/genética , Diferenciación Celular/inmunología , Supervivencia Celular/genética , Supervivencia Celular/inmunología , Células Cultivadas , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/inmunología , Células Madre Embrionarias/citología , Células Madre Embrionarias/inmunología , Células Madre Embrionarias/trasplante , Femenino , Desarrollo Fetal/genética , Desarrollo Fetal/inmunología , Regulación del Desarrollo de la Expresión Génica/inmunología , Células Madre Hematopoyéticas/citología , Células Madre Hematopoyéticas/inmunología , Secuenciación de Nucleótidos de Alto Rendimiento , Proteínas con Dominio LIM , Ratones , Ratones Noqueados , Ratones Transgénicos , Embarazo , Unión Proteica , Elementos Reguladores de la Transcripción/genética , Elementos Reguladores de la Transcripción/inmunología , Activación Transcripcional/genética , Activación Transcripcional/inmunologíaRESUMEN
Cajal-Retzius (C-R) cells play important roles in the lamination of the mammalian cortex via reelin secretion. The genetic mechanisms underlying the development of these neurons have just begun to be unraveled. Here, we show that two closely related LIM-homeobox genes Lhx1 and Lhx5 are expressed in reelin+ cells in various regions in the mouse telencephalon at or adjacent to sites where the C-R cells are generated, including the cortical hem, the mantle region of the septal/retrobulbar area, and the ventral pallium. Whereas Lhx5 is expressed in all of these reelin-expressing domains, Lhx1 is preferentially expressed in the septal area and in a continuous domain spanning from lateral olfactory region to caudomedial territories. Genetic ablation of Lhx5 results in decreased reelin+ and p73+ cells in the neocortical anlage, in the cortical hem, and in the septal, olfactory, and caudomedial telencephalic regions. The overall reduction in number of C-R cells in Lhx5 mutants is accompanied by formation of ectopic reelin+ cell clusters at the caudal telencephalon. Based on differential expression of molecular markers and by fluorescent cell tracing in cultured embryos, we located the origin of reelin+ ectopic cell clusters at the caudomedial telencephalic region. We also confirmed the existence of a normal migration stream of reelin+ cells from the caudomedial area to telencephalic olfactory territories in wild-type embryos. These results reveal a complex role for Lhx5 in regulating the development and normal distribution of C-R cells in the developing forebrain.
Asunto(s)
Moléculas de Adhesión Celular Neuronal/metabolismo , Corteza Cerebral/metabolismo , Proteínas de la Matriz Extracelular/metabolismo , Proteínas de Homeodominio/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Neuronas/metabolismo , Serina Endopeptidasas/metabolismo , Factores de Transcripción/metabolismo , Animales , Moléculas de Adhesión Celular Neuronal/genética , Movimiento Celular , Técnicas de Cultivo de Embriones , Proteínas de la Matriz Extracelular/genética , Proteínas de Homeodominio/genética , Inmunohistoquímica , Hibridación in Situ , Proteínas con Homeodominio LIM , Ratones , Ratones Noqueados , Proteínas del Tejido Nervioso/genética , Proteína Reelina , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Serina Endopeptidasas/genética , Factores de Transcripción/genéticaRESUMEN
The mammalian pituitary gland originates from two separate germinal tissues during embryonic development. The anterior and intermediate lobes of the pituitary are derived from Rathke's pouch, a pocket formed by an invagination of the oral ectoderm. The posterior lobe is derived from the infundibulum, which is formed by evagination of the neuroectoderm in the ventral diencephalon. Previous studies have shown that development of Rathke's pouch and the generation of distinct populations of hormone-producing endocrine cell lineages in the anterior/intermediate pituitary lobes is regulated by a number of transcription factors expressed in the pouch and by inductive signals from the ventral diencephalon/infundibulum. However, little is known about factors that regulate the development of the posterior pituitary lobe. In this study, we show that the LIM-homeobox gene Lhx2 is extensively expressed in the developing ventral diencephalon, including the infundibulum and the posterior lobe of the pituitary. Deletion of Lhx2 gene results in persistent cell proliferation, a complete failure of evagination of the neuroectoderm in the ventral diencephalon, and defects in the formation of the distinct morphological features of the infundibulum and the posterior pituitary lobe. Rathke's pouch is formed and endocrine cell lineages are generated in the anterior/intermediate pituitary lobes of the Lhx2 mutant. However, the shape and organization of the pouch and the anterior/intermediate pituitary lobes are severely altered due to the defects in development of the infundibulum and the posterior lobe. Our study thus reveals an essential role for Lhx2 in the regulation of posterior pituitary development and suggests a mechanism whereby development of the posterior lobe may affect the development of the anterior and intermediate lobes of the pituitary gland.
Asunto(s)
Proteínas de Homeodominio/metabolismo , Hipófisis/embriología , Hipófisis/metabolismo , Factores de Transcripción/metabolismo , Animales , Apoptosis , Tipificación del Cuerpo/genética , Linaje de la Célula , Proliferación Celular , Diencéfalo/embriología , Diencéfalo/metabolismo , Diencéfalo/patología , Embrión de Mamíferos/metabolismo , Embrión de Mamíferos/patología , Regulación del Desarrollo de la Expresión Génica , Genes Homeobox , Proteínas de Homeodominio/genética , Proteínas con Homeodominio LIM , Ratones , Modelos Biológicos , Mutación/genética , Hipófisis/patología , Neurohipófisis/embriología , Neurohipófisis/metabolismo , Neurohipófisis/patología , ARN Mensajero/genética , ARN Mensajero/metabolismo , Transducción de Señal/genética , Factores de Transcripción/genéticaRESUMEN
Vertebrate limb development is controlled by three signaling centers that regulate limb patterning and growth along the proximodistal (PD), anteroposterior (AP) and dorsoventral (DV) limb axes. Coordination of limb development along these three axes is achieved by interactions and feedback loops involving the secreted signaling molecules that mediate the activities of these signaling centers. However, it is unknown how these signaling interactions are processed in the responding cells. We have found that distinct LIM homeodomain transcription factors, encoded by the LIM homeobox (LIM-HD) genes Lhx2, Lhx9 and Lmx1b integrate the signaling events that link limb patterning and outgrowth along all three axes. Simultaneous loss of Lhx2 and Lhx9 function resulted in patterning and growth defects along the AP and the PD limb axes. Similar, but more severe, phenotypes were observed when the activities of all three factors, Lmx1b, Lhx2 and Lhx9, were significantly reduced by removing their obligatory co-factor Ldb1. This reveals that the dorsal limb-specific factor Lmx1b can partially compensate for the function of Lhx2 and Lhx9 in regulating AP and PD limb patterning and outgrowth. We further showed that Lhx2 and Lhx9 can fully substitute for each other, and that Lmx1b is partially redundant, in controlling the production of output signals in mesenchymal cells in response to Fgf8 and Shh signaling. Our results indicate that several distinct LIM-HD transcription factors in conjunction with their Ldb1 co-factor serve as common central integrators of distinct signaling interactions and feedback loops to coordinate limb patterning and outgrowth along the PD, AP and DV axes after limb bud formation.
Asunto(s)
Tipificación del Cuerpo , Extremidades/embriología , Regulación del Desarrollo de la Expresión Génica , Proteínas de Homeodominio/metabolismo , Transducción de Señal , Factores de Transcripción/metabolismo , Animales , Proliferación Celular , Factor 10 de Crecimiento de Fibroblastos/genética , Factor 10 de Crecimiento de Fibroblastos/metabolismo , Proteínas Hedgehog/genética , Proteínas Hedgehog/metabolismo , Proteínas de Homeodominio/genética , Péptidos y Proteínas de Señalización Intercelular/genética , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Ratones , Factores de Transcripción/genéticaRESUMEN
Here we analyze the role of the Lhx6 lim-homeobox transcription factor in regulating the development of subsets of neocortical, hippocampal, and striatal interneurons. An Lhx6 loss-of-function allele, which expresses placental alkaline phosphatase (PLAP), allowed analysis of the development and fate of Lhx6-expressing interneurons in mice lacking this homeobox transcription factor. There are Lhx6+;Dlx+ and Lhx6-;Dlx+ subtypes of tangentially migrating interneurons. Most interneurons in Lhx6(PLAP/PLAP) mutants migrate to the cortex, although less efficiently, and exhibit defects in populating the marginal zone and superficial parts of the neocortical plate. By contrast, migration to superficial parts of the hippocampus is not seriously affected. Furthermore, whereas parvalbumin+ and somatostatin+ interneurons do not differentiate, NPY+ interneurons are present; we suggest that these NPY+ interneurons are derived from the Lhx6-;Dlx+ subtype. Striatal interneurons show deficits distinct from pallial interneurons, including a reduction in the NPY+ subtype. We provide evidence that Lhx6 mediates these effects through promoting expression of receptors that regulate interneuron migration (ErbB4, CXCR4, and CXCR7), and through promoting the expression of transcription factors either known (Arx) or implicated (bMaf, Cux2, and NPAS1) in controlling interneuron development.
Asunto(s)
Diferenciación Celular/fisiología , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Interneuronas/citología , Ratones , Mutación , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Telencéfalo/citología , Telencéfalo/metabolismo , Animales , Animales Recién Nacidos , Movimiento Celular , Corteza Cerebral/citología , Cuerpo Estriado/citología , Cuerpo Estriado/metabolismo , Embrión de Mamíferos , Hipocampo/citología , Hipocampo/metabolismo , Interneuronas/metabolismo , Interneuronas/fisiología , Proteínas con Homeodominio LIM , Ratones/embriología , Ratones/crecimiento & desarrollo , Ratones Endogámicos , Neocórtex/citología , Neocórtex/metabolismo , Factores de TranscripciónRESUMEN
Purkinje cells are one of the major types of neurons that form the neural circuitry in the cerebellum essential for fine control of movement and posture. During development, Purkinje cells also are critically involved in the regulation of proliferation of progenitors of granule cells, the other major type of neurons in the cerebellum. The process that controls differentiation of Purkinje cells from their early precursors is poorly understood. Here we report that two closely related LIM-homeobox genes, Lhx1 and Lhx5, were expressed in the developing Purkinje cells soon after they exited the cell cycle and migrated out of the cerebellar ventricular zone. Double-mutant mice lacking function of both Lhx1 and Lhx5 showed a severe reduction in the number of Purkinje cells. In addition, targeted inactivation of Ldb1, which encodes a cofactor for all LIM-homeodomain proteins, resulted in a similar phenotype. Our studies thus provide evidence that these transcription regulators are essential for controlling Purkinje cell differentiation in the developing mammalian cerebellum.
Asunto(s)
Cerebelo/embriología , Proteínas de Unión al ADN/fisiología , Proteínas de Homeodominio/fisiología , Proteínas del Tejido Nervioso/fisiología , Células de Purkinje/citología , Factores de Transcripción/fisiología , Animales , Diferenciación Celular , Femenino , Proteínas con Dominio LIM , Proteínas con Homeodominio LIM , Ratones , Ratones Endogámicos C57BL , EmbarazoRESUMEN
Patients with nail-patella syndrome, which among other symptoms also includes podocyte-associated renal failure, suffer from mutations in the LMX1B gene. The disease severity among patients is quite variable and has given rise to speculations on the presence of modifier genes. Promising candidates for modifier proteins are the proteins interacting with LMX1B, such as LDB1 and E47. Since human kidney samples from patients are difficult to obtain, conventional Lmx1b knock-out mice have been extremely valuable to study the role of Lmx1b in podocyte differentiation. In contrast to findings in these mice, however, in which a downregulation of the Col4a3, Col4a4 and Nphs2 genes has been described, no such changes have been detected in kidney biopsies from patients. We now report on our results on the characterization of constitutive podocyte-specific Lmx1b, Ldb1 and E2a knock-out mice. Constitutive podocyte-specific Lmx1b knock-out mice survive for approximately 2 weeks after birth and do not present with a downregulation of the Col4a3, Col4a4 and Nphs2 genes, therefore they mimic the human disease more closely. The podocyte-specific Ldb1 knock-out mice survive longer, but then also succumb to renal failure, whereas the E2a knock-out mice show no renal symptoms for at least 6 months after birth. We conclude that LDB1, but not E2A is a promising candidate as a modifier gene in patients with nail-patella syndrome.