RESUMEN
Newly synthesised glycoproteins enter the rough endoplasmic reticulum through a translocation pore. The translocon associated protein (TRAP) complex is located close to the pore. In a patient with a homozygous start codon variant in TRAPγ (SSR3), absence of TRAPγ causes disruption of the TRAP complex, impairs protein translocation into the endoplasmic reticulum and affects transport, for example, into the brush-border membrane. Furthermore, we observed an unbalanced non-occupancy of N-glycosylation sites. The major clinical features are intrauterine growth retardation, facial dysmorphism, congenital diarrhoea, failure to thrive, pulmonary disease and severe psychomotor disability.
Asunto(s)
Retículo Endoplásmico Rugoso/genética , Retardo del Crecimiento Fetal/genética , Glicoproteínas/genética , Fosfatasa Ácida Tartratorresistente/genética , Niño , Preescolar , Diarrea/genética , Diarrea/patología , Insuficiencia de Crecimiento/genética , Insuficiencia de Crecimiento/patología , Femenino , Retardo del Crecimiento Fetal/patología , Glicoproteínas/biosíntesis , Glicosilación , Humanos , Lactante , Recién Nacido , Enfermedades Pulmonares/genética , Enfermedades Pulmonares/patología , Masculino , Trastornos Psicomotores/genética , Trastornos Psicomotores/patología , Fosfatasa Ácida Tartratorresistente/deficienciaRESUMEN
Importin 13 (Imp13) is a bidirectional nuclear transporter of proteins involved in a range of important cellular processes, with an N-terminally truncated inhibitory isoform (tImp13) specifically expressed in testis. To gain insight into tImp13 function, we performed a yeast-2-hybrid screen from a human testis cDNA library, identifying for the first time a suite of interactors with roles in diverse cellular process. We validated the interaction of tImp13 with Eukaryotic translation initiation factor 4γ2 (EIF4G2) and High mobility group containing protein 20A (HMG20A), benchmarking that with glucocorticoid receptor (GR), a known Imp13 interactor expressed in testis. Coimmunoprecipitation assays indicated association of both tImp13 and Imp13 with EIF4G2, HMG20A and GR. Quantitative confocal microscopic analysis revealed the ability of tImp13 to inhibit the nuclear localisation of EIF4G2, HMG20A and GR, as well as that of Imp13 to act as a nuclear exporter for both EIF4G2 and HMG20A, and as a nuclear importer for GR. The physiological relevance of these results was highlighted by the cytoplasmic localisation of EIF4G2, HMG20A and GR in pachytene spermatocytes/round spermatids in the murine testis where tImp13 is present at high levels, in contrast to the nuclear localisation of HMG20A and GR in spermatogonia, where tImp13 is largely absent. Interestingly, Imp13, EIF4G2, HMG20A and GR were found together in the acrosome vesicle of murine epididymal spermatozoa. Collectively, our findings show, for the first time, that tImp13 may have a functional role in the mature spermatozoa, in addition to that in the meiotic germ cells of the testis.
Asunto(s)
Núcleo Celular/metabolismo , Regulación del Desarrollo de la Expresión Génica , Carioferinas/metabolismo , Espermátides/metabolismo , Espermatocitos/metabolismo , Espermatogénesis/genética , Animales , Factor 4G Eucariótico de Iniciación/genética , Factor 4G Eucariótico de Iniciación/metabolismo , Biblioteca de Genes , Proteínas del Grupo de Alta Movilidad/genética , Proteínas del Grupo de Alta Movilidad/metabolismo , Humanos , Carioferinas/genética , Masculino , Ratones , Unión Proteica , Mapeo de Interacción de Proteínas , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Transporte de Proteínas , Receptores de Glucocorticoides/genética , Receptores de Glucocorticoides/metabolismo , Transducción de Señal , Espermátides/crecimiento & desarrollo , Espermátides/ultraestructura , Espermatocitos/crecimiento & desarrollo , Espermatocitos/ultraestructura , Testículo/citología , Testículo/crecimiento & desarrollo , Testículo/metabolismo , Técnicas del Sistema de Dos HíbridosRESUMEN
The Spalt-like 4 (Sall4) zinc finger protein is a critical transcription factor for pluripotency in embryonic stem cells (ESCs). It is also involved in the formation of a variety of organs, in mice, and humans. We report the essential roles of Sall4 in mouse primordial germ cell (PGC) specification. PGC specification is accompanied by the activation of the stem cell program and repression of the somatic cell program in progenitor cells. Conditional inactivation of Sall4 during PGC specification led to a reduction in the number of PGCs in embryonic gonads. Sall4(del/del) PGCs failed to translocate from the mesoderm to the endoderm and underwent apoptosis. In Sall4(del/del) PGC progenitors, somatic cell program genes (Hoxa1 and Hoxb1) were derepressed, while activation of the stem cell program was not impaired. We demonstrated that in differentiated ESCs, Sall4 bound to these somatic cell program gene loci, which are reportedly occupied by Prdm1 in embryonic carcinoma cells. Given that Sall4 and Prdm1 are known to associate with the histone deacetylase repressor complex, our findings suggest that Sall4 suppresses the somatic cell program possibly by recruiting the repressor complex in conjunction with Prdm1; therefore, it is essential for PGC specification.
Asunto(s)
Proteínas de Unión al ADN/metabolismo , Células Germinativas/citología , Factores de Transcripción/metabolismo , Animales , Diferenciación Celular/fisiología , Proteínas de Unión al ADN/genética , Femenino , Perfilación de la Expresión Génica , Células Germinativas/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Factor 1 de Unión al Dominio 1 de Regulación Positiva , Células Madre , Factores de Transcripción/genéticaRESUMEN
Sex determination is essential for the sexual reproduction to generate the next generation by the formation of functional male or female gametes. In mammals, primary sex determination is commenced by the presence or absence of the Y chromosome, which controls the fate of the gonadal primordium. The somatic precursor of gonads, the genital ridge is formed at the mid-gestation stage and gives rise to one of two organs, a testis or an ovary. The fate of the genital ridge, which is governed by the differentiation of somatic cells into Sertoli cells in the testes or granulosa cells in the ovaries, further determines the sex of an individual and their germ cells. Mutation studies in human patients with disorders of sex development and mouse models have revealed factors that are involved in mammalian sex determination. In most of mammals, a single genetic trigger, the Y-linked gene Sry (sex determination region on Y chromosome), regulates testicular differentiation. Despite identification of Sry in 1990, precise mechanisms underlying the sex determination of bipotential genital ridges are still largely unknown. Here, we review the recent progress that has provided new insights into the mechanisms underlying genital ridge formation as well as the regulation of Sry expression and its functions in male sex determination of mice.
Asunto(s)
Regulación del Desarrollo de la Expresión Génica , Genitales Masculinos/metabolismo , Procesos de Determinación del Sexo , Proteína de la Región Y Determinante del Sexo/genética , Animales , Genitales Masculinos/embriología , Humanos , Masculino , Ratones , Modelos Genéticos , Factor de Transcripción SOX9/genéticaRESUMEN
A well-established experimental paradigm to analyze gene function in development is to elucidate the impact of gain and loss of gene activity on cell differentiation, tissue modelling, organogenesis, and morphogenesis. This chapter describes the experimental protocols to study gene function by means of electroporation and lipofection to manipulate genetic activity in whole embryos and fetal organs in vitro. These techniques allow for more precise control of the timing, with reference to developmental age or stage, and the cell/tissue-specificity of the changes in gene activity. They provide an alternative strategy that can expedite the analysis of gene function before resorting to the conventional means of transgenesis and gene targeting in the whole organism.
Asunto(s)
Embrión de Mamíferos/metabolismo , Desarrollo Embrionario/genética , Morfogénesis/genética , Organogénesis/genética , Animales , Electroporación , Embrión de Mamíferos/citología , Regulación del Desarrollo de la Expresión Génica , Marcación de Gen , Ratones , Biología Molecular/métodosRESUMEN
The Y-linked gene Sry regulates mammalian sex determination in bipotential embryonic gonads. Here, we report that the transcription factors Six1 and Six4 are required for male gonadal differentiation. Loss of Six1 and Six4 together, but neither alone, resulted in a male-to-female sex-reversal phenotype in XY mutant gonads accompanied by a failure in Sry activation. Decreased gonadal precursor cell formation at the onset of Sry expression and a gonadal size reduction in both sexes were also found in mutant embryos. Forced Sry transgene expression in XY mutant gonads rescued testicular development but not the initial disruption to precursor growth. Furthermore, we identified two downstream targets of Six1/Six4 in gonadal development, Fog2 (Zfpm2) and Nr5a1 (Ad4BP/Sf1). These two distinct Six1/Six4-regulated pathways are considered to be crucial for gonadal development. The regulation of Fog2 induces Sry expression in male sex determination, and the regulation of Nr5a1 in gonadal precursor formation determines gonadal size.
Asunto(s)
Proteínas de Homeodominio/metabolismo , Procesos de Determinación del Sexo/genética , Testículo/embriología , Testículo/metabolismo , Transactivadores/metabolismo , Animales , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Femenino , Regulación del Desarrollo de la Expresión Génica , Proteínas de Homeodominio/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Modelos Biológicos , Tamaño de los Órganos/genética , Organogénesis , Proteína de la Región Y Determinante del Sexo/genética , Proteína de la Región Y Determinante del Sexo/metabolismo , Cordón Espermático/embriología , Cordón Espermático/metabolismo , Factor Esteroidogénico 1/genética , Factor Esteroidogénico 1/metabolismo , Testículo/citología , Transactivadores/genética , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Regulación hacia Arriba/genéticaRESUMEN
Dullard/Ctdnep1 is a member of the serine/threonine phosphatase family of the C-terminal domain of eukaryotic RNA polymerase II. Embryos lacking Dullard activity fail to form primordial germ cells (PGCs). In the mouse, the formation of PGCs is influenced by BMP4 and WNT3 activity. Although Dullard is reputed to negatively regulate BMP receptor function, in this study we found mutations in Dullard had no detectable effect on BMP4 and p-Smad activity. Furthermore Dullard mutations did not influence the dosage-dependent inductive effect of Bmp4 in PGC formation. However, Dullard may function as a positive regulator of WNT signalling. Combined loss of one copy each of Dullard and Wnt3 had a synergistic effect on the reduction of PGC numbers in the compound heterozygous embryo. In addition, loss of Dullard function was accompanied by down-regulation of WNT/ß-catenin signalling activity and a reduction in the level of Dishevelled 2 (Dvl2). Therefore, Dullard may play a role in the fine-tuning of WNT signalling activity by modulating the expression of ligands/antagonists and the availability of Dvl2 protein during specification of the germ cell lineage.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/genética , Regulación del Desarrollo de la Expresión Génica , Células Germinativas/metabolismo , Fosfoproteínas Fosfatasas/genética , Fosfoproteínas/genética , Transducción de Señal/genética , Proteína Wnt3/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Animales , Proteína Morfogenética Ósea 4/genética , Proteína Morfogenética Ósea 4/metabolismo , Linaje de la Célula/genética , Proteínas Dishevelled , Embrión de Mamíferos , Células Germinativas/citología , Heterocigoto , Homocigoto , Ratones , Morfogénesis/genética , Mutación , Fosfoproteínas Fosfatasas/deficiencia , Fosfoproteínas/metabolismo , Proteínas Smad/genética , Proteínas Smad/metabolismo , Proteína Wnt3/metabolismo , beta Catenina/genética , beta Catenina/metabolismoRESUMEN
WNT signaling activity is involved in the regulation of many cellular functions, including proliferation, migration, cell fate specification, maintenance of pluripotency and induction of tumorigenicity. Here we summarize recent progress towards understanding the regulation of canonical WNT/ß-catenin signaling activity through feedback regulatory loops involving the ligands, agonists and antagonists, the availability of intracellular pools of active ß-catenin and the cross-regulation of the WNT activity by ß-catenin independent pathway. We also review recent findings on the role of WNT/ß-catenin signaling in tissue lineage differentiation during embryogenesis and the maintenance and self renewal of embryo-derived stem cells in vitro.
Asunto(s)
Diferenciación Celular , Embrión de Mamíferos/citología , Regulación del Desarrollo de la Expresión Génica , Proteínas Wnt/metabolismo , Vía de Señalización Wnt , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Movimiento Celular , Embrión de Mamíferos/metabolismo , Desarrollo Embrionario , Células Madre Embrionarias/citología , Receptores Frizzled/genética , Receptores Frizzled/metabolismo , Estratos Germinativos/citología , Estratos Germinativos/embriología , Glipicanos/metabolismo , Corazón/embriología , Corazón/crecimiento & desarrollo , Proteoglicanos de Heparán Sulfato/metabolismo , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Ratones , Fenotipo , Factores de Transcripción SOXB1/genética , Factores de Transcripción SOXB1/metabolismo , Proteínas Wnt/agonistas , Proteínas Wnt/antagonistas & inhibidores , Proteínas Wnt/genética , beta Catenina/genética , beta Catenina/metabolismoRESUMEN
The translocon-associated protein (TRAP, also termed the signal sequence receptor) complex is required for the efficient translocation of secretory and membrane proteins in the endoplasmic reticulum, and is also involved in the endoplasmic reticulum stress-mediated unfolded protein response pathway. To investigate the roles of Trap-γ, a TRAP complex subunit, we generated Trap-γ knockout mice and found that mutant pups died soon after birth because of retarded embryonic organ growth, especially in the lung. The mutant placentae showed severe vascular network malformation in the labyrinth and significant reductions in blood space areas, which had an adverse effect on intrauterine embryonic growth. Placental malformation was already found by the mid-gestation-stage mutant placenta, with poor vascular endothelial cell proliferation in the chorionic plate region and increased apoptotic cell death in the labyrinth. Thus, Trap-γ appears to be required for vascular network formation in murine placental development.
Asunto(s)
Proteínas de Unión al Calcio/metabolismo , Glicoproteínas de Membrana/metabolismo , Placenta/irrigación sanguínea , Subunidades de Proteína/metabolismo , Receptores Citoplasmáticos y Nucleares/metabolismo , Receptores de Péptidos/metabolismo , Animales , Apoptosis , Peso Corporal , Proteínas de Unión al Calcio/genética , Proliferación Celular , Embrión de Mamíferos/anatomía & histología , Embrión de Mamíferos/fisiología , Células Madre Embrionarias/citología , Células Madre Embrionarias/fisiología , Células Endoteliales/citología , Células Endoteliales/fisiología , Femenino , Marcación de Gen , Glicoproteínas de Membrana/genética , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos CBA , Ratones Noqueados , Mutagénesis Sitio-Dirigida , Tamaño de los Órganos , Placenta/anomalías , Placenta/patología , Placentación , Embarazo , Subunidades de Proteína/genética , Receptores Citoplasmáticos y Nucleares/genética , Receptores de Péptidos/genéticaRESUMEN
Mouse embryos lacking Lhx1 (Lim1) activity display defective gastrulation and are deficient of primordial germ cells (PGCs) (Tsang et al. [2001] International Journal of Developmental Biology 45:549-555). To dissect the specific role of Lhx1 in germ cell development, we studied embryos with conditional inactivation of Lhx1 activity in epiblast derivatives, which, in contrast to completely null embryos, develop normally through gastrulation before manifesting a head truncation phenotype. Initially, PGCs are localized properly to the definitive endoderm of the posterior gut in the conditional mutant embryos, but they depart from the embryonic gut prematurely. The early exit of PGCs from the gut is accompanied by the failure to maintain a strong expression of Ifitm1 in the mesoderm enveloping the gut, which may mediate the repulsive activity that facilitates the retention of PGCs in the hindgut during early organogenesis. Lhx1 therefore may influence the localization of PGCs by modulating Ifitm1-mediated repulsive activity.
Asunto(s)
Embrión de Mamíferos/citología , Embrión de Mamíferos/metabolismo , Células Germinativas/citología , Proteínas de Homeodominio/metabolismo , Animales , Endodermo/citología , Femenino , Regulación del Desarrollo de la Expresión Génica , Estratos Germinativos/citología , Estratos Germinativos/metabolismo , Proteínas de Homeodominio/genética , Hibridación in Situ , Proteínas con Homeodominio LIM , Masculino , Ratones , Ratones Noqueados , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factores de TranscripciónRESUMEN
The kidney develops through reciprocal interactions between two precursor tissues: the metanephric mesenchyme and the ureteric bud. We previously demonstrated that the zinc finger protein Sall1 is essential for ureteric bud attraction toward the mesenchyme. Here, we show that Kif26b, a kinesin family gene, is a downstream target of Sall1 and that disruption of this gene causes kidney agenesis because of impaired ureteric bud attraction. In the Kif26b-null metanephros, compact adhesion between mesenchymal cells adjacent to the ureteric buds and the polarized distribution of integrin alpha8 were impaired, resulting in failed maintenance of Gdnf, a critical ureteric bud attractant. Overexpression of Kif26b in vitro caused increased cell adhesion through interactions with nonmuscle myosin. Thus, Kif26b is essential for kidney development because it regulates the adhesion of mesenchymal cells in contact with ureteric buds.
Asunto(s)
Adhesión Celular/fisiología , Riñón/embriología , Cinesinas/metabolismo , Mesodermo/fisiología , Animales , Southern Blotting , Clonación Molecular , Cartilla de ADN/genética , Femenino , Inmunohistoquímica , Hibridación in Situ , Riñón/citología , Cinesinas/genética , Espectrometría de Masas , Ratones , Ratones Endogámicos C57BL , TransfecciónRESUMEN
Importin-mediated transport of cargoes is known to be a key mechanism for nucleo-cytoplasmic trafficking of molecules. Ipo13, which is a member of Importin-beta gene family, encodes two transcripts by utilizing different transcription start sites. In the mouse, the full-length transcript (L-Ipo13) is expressed in the primordial germ cells in the embryo and is later expressed predominantly at the pachytene phase of meiosis in both male and female germ cells. The shorter transcript (TS-Ipo13) is only expressed in the germ cells in the adult testis. Activity of L-Ipo13, but not TS-Ipo13, mediates the nuclear accumulation of ubiquitin-conjugating enzyme 9 (UBC9), a cargo of human IPO13. This finding is consistent with the progressive accumulation of UBC9 in the nucleus of the meiotic germ cells after the onset of L-Ipo13 expression. Following siRNA knockdown of IPO13 activity in the fetal ovary, fewer germ cells are found to progress to the late-pachytene stage of meiosis and nuclear accumulation of UBC9 is reduced. Our findings strongly implicate a stage-specific role of IPO13 in nuclear-cytoplasmic translocation of cargoes that accompanies meiotic differentiation of the mouse germ cells.
Asunto(s)
Regulación del Desarrollo de la Expresión Génica , Células Germinativas/metabolismo , Carioferinas/fisiología , Meiosis , Secuencia de Aminoácidos , Animales , Núcleo Celular/metabolismo , Clonación Molecular , Citoplasma/metabolismo , Femenino , Carioferinas/química , Carioferinas/metabolismo , Ratones , Datos de Secuencia Molecular , Homología de Secuencia de Aminoácido , Enzimas Ubiquitina-Conjugadoras/metabolismoRESUMEN
The mouse embryo is built by assembling the progenitors of various tissue types into a body plan. Early postimplantation development involves the establishment of anatomical asymmetries and regionalized gene expression in the conceptus, the specification of tissue lineages, and the coordination of cell movement for correct positioning of the lineage progenitors before and at gastrulation. Recent findings reveal that Wnt and Tgfbeta signalling function is instrumental in delineating the anterior-posterior embryonic axis by defining the site of primitive streak formation and by directing the movement of the visceral endoderm. These signalling activities are also required for the specification of anterior and posterior fates of the epiblast cells and for the induction and navigation of the primordial germ cells.
Asunto(s)
Tipificación del Cuerpo/fisiología , Gástrula/fisiología , Activinas/fisiología , Animales , Linaje de la Célula , Movimiento Celular , Endodermo/citología , Endodermo/fisiología , Células Germinativas/fisiología , Proteínas de la Membrana/fisiología , Ratones , Proteína Nodal , Transducción de Señal , Factor de Crecimiento Transformador beta/fisiología , Proteínas Wnt/fisiologíaRESUMEN
The low density lipoprotein receptor-related protein 4 gene (Lrp4) was identified by subtractive screening of cDNAs of the migratory primordial germ cells (PGCs) of E8.5-9.5 embryo and E3.5 blastocysts. Lrp4 is expressed in PGCs in the hindgut and the dorsal mesentery of E9.5 embryos, and in germ cells in the genital ridges of male and female E10.5-13.5 embryos. Lrp4 is also expressed in spermatogonia of the neonatal and adult testes and in the immature oocytes and follicular cells of the adult ovary. The absence of Lrp4 expression in the blastocyst, embryonic stem cells and embryonic germ cells suggests the Lrp4 is a molecular marker that distinguishes the germ cells from embryo-derived pluripotent stem cells.
Asunto(s)
Células Germinativas/metabolismo , Receptores de LDL/genética , Secuencia de Aminoácidos , Animales , Movimiento Celular , ADN Complementario/aislamiento & purificación , Femenino , Componentes del Gen , Gónadas/embriología , Gónadas/metabolismo , Proteínas Relacionadas con Receptor de LDL , Masculino , Ratones , Datos de Secuencia Molecular , Células Madre Multipotentes/metabolismo , Distribución TisularRESUMEN
The family of interferon-induced transmembrane protein (Ifitm/mil/fragilis) genes encodes cell surface proteins that may modulate cell adhesion and influence cell differentiation. Mouse Ifitm1 and -3, which are expressed in primordial germ cells (PGCs), are implicated to have roles in germ cell development, but the specific functions have been unclear. Our results show that Ifitm1 activity is required for PGC transit from the mesoderm into the endoderm, and that it appears to act via a repulsive mechanism, such that PGCs avoid Ifitm1-expressing tissues. In contrast, Ifitm3, which is expressed in migratory PGCs, is sufficient to confer autonomous PGC-like homing properties to somatic cells. These guidance activities are mediated by the N-terminal extracellular domain of the specific IFITM, which cannot be substituted by that of another family member. Complex homo- and/or heterotypic intercellular interactions among various IFITMs in PGCs and neighboring cells may underpin coordinated germ cell guidance in mice.
Asunto(s)
Movimiento Celular/fisiología , Embrión de Mamíferos/metabolismo , Endodermo/metabolismo , Células Germinativas/fisiología , Proteínas de la Membrana/fisiología , Mesodermo/metabolismo , Animales , Antígenos de Diferenciación , Proteínas Cromosómicas no Histona , Sondas de ADN , Regulación del Desarrollo de la Expresión Génica , Hibridación in Situ , Ratones , Factor 3 de Transcripción de Unión a Octámeros/genética , Factor 3 de Transcripción de Unión a Octámeros/metabolismo , ARN Interferente Pequeño/farmacología , Proteínas de Unión al ARN/fisiología , Proteínas Recombinantes de Fusión/fisiología , Proteínas Represoras/genética , Proteínas Represoras/metabolismoRESUMEN
Mouse interferon-induced transmembrane protein (IFITM) gene, Ifitm3 (previously known as mil-1 and fragilis), is expressed in primordial germ cells (PGCs), in their precursors, and in germ cells of the fetal gonads (Saitou et al. [2002] Nature 418:293-300; Tanaka and Matsui [2002] Mech Dev 119S:S261-S267). By examining the expression of green fluorescent protein transgene under the control of DNA sequences flanking exon 1, we have identified domains that direct Ifitm3 transcription in PGCs and their precursors in gastrula stage and 13.5 days post coitum embryos. Germ cell-specific expression is achieved by the activity of a consensus element unique to the Ifitm genes, which may act to suppress Ifitm3 expression in somatic tissues. The lack of any influence of the interferon-stimulable response elements on transgene expression in the germ-line suggests that interferon-mediated response is not critical for activating Ifitm3.
Asunto(s)
Regulación del Desarrollo de la Expresión Génica , Células Germinativas/citología , Interferones/metabolismo , Proteínas de la Membrana/biosíntesis , Proteínas de la Membrana/fisiología , Proteínas de Unión al ARN/biosíntesis , Proteínas de Unión al ARN/fisiología , Regiones no Traducidas 3' , Animales , Secuencia de Bases , ADN/genética , Exones , Femenino , Gástrula , Genes Reporteros , Proteínas Fluorescentes Verdes/metabolismo , Inmunohistoquímica , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos DBA , Microscopía Fluorescente , Modelos Genéticos , Datos de Secuencia Molecular , Plásmidos/metabolismo , Estructura Terciaria de Proteína , Homología de Secuencia de Ácido Nucleico , Factores Sexuales , Factores de Tiempo , TransgenesRESUMEN
In all multicellular organisms, germ cells originating from a fertilized egg have the highly specialized role of transmitting genetic information to the next generation. In many animal species, the establishment of the germ cell lineage is regulated by the maternally inherited germplasm. In mammals, however, germline determination is not based on the unequal distribution of maternal determinants. In the processes of mammalian germ cell formation and subsequent differentiation, the molecular basis of the acquisition of germ cell status is not well understood. Since migrating primordial germ cells (PGCs) are lineage-restricted to the germline, they have already acquired a germ cell specific fate distinct from that of pluri/multi-potent stem cells. However, there have been no molecules known to be expressed in migrating PGCs but not in the inner cell mass of blastocysts. Such molecules should be involved in early germ cell development, and they should make good markers for following the process of PGC formation. To identify such molecules, we performed a subtracted cDNA screening with migrating PGCs and blastocysts in mice, and isolated 11 clones preferentially expressed in PGCs. Here, we report the identification of two genes with similarity to human interferon-induced transmembrane protein (Ifitm) genes, and expression patterns of these genes in forming and in differentiating PGCs. During germ cell formation, mouse Ifitm like (mil)-1 was expressed in putative PGC ancestors in embryos at 6.5-7.5 days post coitum. In migrating PGCs, mil-1 expression was continuously observed and mil-2 expression was first detected during germ cell differentiation.
Asunto(s)
Regulación del Desarrollo de la Expresión Génica , Inductores de Interferón , Animales , Diferenciación Celular , Embrión de Mamíferos/metabolismo , Células Germinativas/metabolismo , Humanos , Interferones , RatonesRESUMEN
In all multicellular organisms, germ cells originating from a fertilized egg have the highly specialized role of transmitting genetic information to the next generation. In many animal species, the establishment of the germ cell lineage is regulated by the maternally inherited germplasm. In mammals, however, germline determination is not based on the unequal distribution of maternal determinants. In the processes of mammalian germ cell formation and subsequent differentiation, the molecular basis of the acquisition of germ cell status is not well understood. Since migrating primordial germ cells (PGCs) are lineage-restricted to the germline, they have already acquired a germ cell specific fate distinct from that of pluri/multi-potent stem cells. However, there have been no molecules known to be expressed in migrating PGCs but not in the inner cell mass of blastocysts. Such molecules should be involved in early germ cell development, and they should make good markers for following the process of PGC formation. To identify such molecules, we performed a subtracted cDNA screening with migrating PGCs and blastocysts in mice, and isolated 11 clones preferentially expressed in PGCs. Here, we report the identification of two genes with similarity to human interferon-induced transmembrane protein (Ifitm) genes, and expression patterns of these genes in forming and in differentiating PGCs. During germ cell formation, mouse Ifitm like (mil)-1 was expressed in putative PGC ancestors in embryos at 6.5-7.5 days post coitum. In migrating PGCs, mil-1 expression was continuously observed and mil-2 expression was first detected during germ cell differentiation.