RESUMO
During early mouse development the homeobox gene Hesx1 is expressed in prospective forebrain tissue, but later becomes restricted to Rathke's pouch, the primordium of the anterior pituitary gland. Mice lacking Hesx1 exhibit variable anterior CNS defects and pituitary dysplasia. Mutants have a reduced prosencephalon, anopthalmia or micropthalmia, defective olfactory development and bifurcations in Rathke's pouch. Neonates exhibit abnormalities in the corpus callosum, the anterior and hippocampal commissures, and the septum pellucidum. A comparable and equally variable phenotype in humans is septo-optic dysplasia (SOD). We have cloned human HESX1 and screened for mutations in affected individuals. Two siblings with SOD were homozygous for an Arg53Cys missense mutation within the HESX1 homeodomain which destroyed its ability to bind target DNA. These data suggest an important role for Hesx1/HESX1 in forebrain, midline and pituitary development in mouse and human.
Assuntos
Anormalidades Múltiplas/genética , Sequências Hélice-Alça-Hélice/genética , Proteínas de Homeodomínio/genética , Mutação , Hipófise/anormalidades , Septo Pelúcido/anormalidades , Anormalidades Múltiplas/patologia , Alelos , Sequência de Aminoácidos , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos , DNA/metabolismo , Desenvolvimento Embrionário e Fetal/genética , Feminino , Genótipo , Proteínas de Homeodomínio/fisiologia , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Dados de Sequência Molecular , Fases de Leitura Aberta , Nervo Óptico/embriologia , Nervo Óptico/patologia , Linhagem , Hipófise/embriologia , Proteínas Repressoras , Septo Pelúcido/embriologia , Fatores de Transcrição HES-1RESUMO
Expressed-sequence tag (EST) maps are an adjunct to sequence-based analytical methods of gene detection and localization for those species for which such data are available, and provide anchors for high-density homology and orthology mapping in species for which large-scale sequencing has yet to be done. Species for which radiation hybrid-based transcript maps have been established include human, rat, mouse, dog, cat and zebrafish. We have established a comprehensive first-generation-placement radiation hybrid map of the mouse consisting of 5,904 mapped markers (3,993 ESTs and 1,911 sequence-tagged sites (STSs)). The mapped ESTs, which often originate from small-EST clusters, are enriched for genes expressed during early mouse embryogenesis and are probably different from those localized in humans. We have confirmed by in situ hybridization that even singleton ESTs, which are usually not retained for mapping studies, may represent bona fide transcribed sequences. Our studies on mouse chromosomes 12 and 14 orthologous to human chromosome 14 show the power of our radiation hybrid map as a predictive tool for orthology mapping in humans.
Assuntos
Genoma , Células Híbridas/efeitos da radiação , RNA Mensageiro/genética , Animais , Mapeamento Cromossômico , Etiquetas de Sequências Expressas , Hibridização In Situ , CamundongosRESUMO
Recently, genes with similar expression patterns in the early gastrulae of several different vertebrate species have been identified. The remarkable conservation of these expression patterns suggests that fundamental similarities exist within the vertebrates at remarkably early stages. It has yet to be established exactly how these genes are activated in the correct spatial patterns and what their functions might be.
Assuntos
Gástrula/fisiologia , Vertebrados/embriologia , Animais , Indução Embrionária/fisiologia , Regulação da Expressão Gênica , Mesoderma , Transdução de Sinais , Vertebrados/genéticaRESUMO
Screens for genetic mutations have been instrumental in identifying genes needed to execute particular biological processes. They have also helped to resolve the function of individual genes. Now the notion of large-scale mutagenesis screens in mouse, an experimental model for humans, is becoming a reality.
Assuntos
Mutagênese , Animais , Testes Genéticos , Humanos , Camundongos , Modelos Genéticos , FenótipoRESUMO
BACKGROUND: After implantation, the basic body plan of the mammalian embryo is established during gastrulation when the epithelial founder tissue of the fetus, the epiblast, gives rise to new tissues by ingression through the primitive streak. Formation of the primitive streak defines the caudal aspect of the embryo and thus the anteroposterior axis. Further patterning of this axis has been attributed to signals produced by tissues arising from the primitive streak, and in particular the mesendoderm located along the midline of the embryo is thought to be responsible for the correct anteroposterior subdivision of the neurectoderm as it begins to form the central nervous system (CNS). RESULTS: In situ hybridization studies show that the onset of expression of the homeobox-containing gene Hesx1 coincides with the formation of the primitive streak, but occurs on the opposite side of the embryo, in a small domain of anterior endoderm. Lineage tracing using a lipophilic fluorescent label shows that the first endoderm cells to express Hesx1 are not destined to contribute to the future embryo, but instead belong to the primitive endoderm lineage and will be displaced by definitive endoderm arising from the primitive streak during gastrulation. Approximately 24 hours after Hesx1 transcripts are first detected in the endoderm, they start to appear in adjacent ectoderm that gives rise to the most anterior component of the developing CNS, the prosencephalon, which continues to express Hesx1. Eventually, Hesx1 transcripts are detectable only in Rathke's pouch as the pituitary starts to develop. Removal of endoderm cells expressing Hesx1 during the earlier stages of gastrulation either prevents or severely curtails the later expression of Hesx1 in ectoderm and neurectoderm, but does not affect gene expression in more caudal regions of the developing CNS. CONCLUSIONS: As overt anterior pattern is present in the visceral embryonic endoderm prior to formation of any axial mesendoderm, a mechanism for bestowing anterior pattern must exist which is independent of primitive streak descendants. Furthermore, correct molecular patterning of the most rostral neurectoderm appears to depend on the presence of this anterior visceral embryonic endoderm during the early stages of gastrulation. We propose that primitive endoderm is responsible for the initial induction of rostral identity in the embryo, and in particular for the correct definition of the future prosencephalic neurectoderm. Subsequently, this identity will be reinforced and maintained by axial mesendoderm when it displaces the visceral embryonic endoderm during the course of gastrulation.
Assuntos
Endoderma/metabolismo , Expressão Gênica , Proteínas de Homeodomínio/genética , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , Proteínas Repressoras , Fatores de Transcrição HES-1RESUMO
BACKGROUND: Signals from anterior endodermal cells that express the homeobox gene Hex initiate development of the most rostral tissues of the mouse embryo. The dorsal/anterior endoderm of the Xenopus gastrula, which expresses Hex and the putative head-inducing gene cerberus, is proposed to be equivalent to the mouse anterior endoderm. Here, we report the origin and signalling properties of this population of cells in the early Xenopus embryo. RESULTS: Xenopus anterior endoderm was found to derive in part from cells at the centre of the blastocoel floor that express XHex, the Xenopus cognate of Hex. Like their counterparts in the mouse embryo, these Hex-expressing blastomeres moved to the dorsal side of the Xenopus embryo as gastrulation commenced, and populated deep endodermal adjacent to Spemann's organiser. Experiments involving the induction of secondary axes confirmed that XHex expression was associated with anterior development. Ventral misexpression of XHex induced ectopic cerberus expression and conferred anterior signalling properties to the endoderm. Unlike the effect of misexpressing cerberus, these signals could not neuralise overlying ectoderm. CONCLUSIONS: XHex expression reveals the unexpected origin of an anterior signalling centre in Xenopus, which arises in part from the centre of the blastula and localises to the deep endoderm adjacent to Spemann's organiser. Signals originating from these endodermal cells impart an anterior identity to the overlying ectoderm, but are insufficient for neural induction. The anterior movement of Hex-expressing cells in both Xenopus and mouse embryos suggests that this process is a conserved feature of vertebrate development.
Assuntos
Blastocisto/fisiologia , Genes Homeobox , Proteínas de Homeodomínio/fisiologia , Xenopus laevis/embriologia , Animais , Blastocisto/citologia , Blastômeros/citologia , Indução Embrionária/fisiologia , Endoderma/citologia , Gástrula/fisiologia , Gástrula/ultraestrutura , Cabeça/embriologia , Proteínas de Homeodomínio/genética , Peptídeos e Proteínas de Sinalização Intercelular , Camundongos , Microinjeções , Morfogênese/genética , Especificidade de Órgãos , Proteínas/genética , Proteínas/fisiologia , RNA Mensageiro/administração & dosagem , RNA Mensageiro/genética , Especificidade da Espécie , Fatores de Transcrição , Proteínas de Xenopus , Xenopus laevis/genéticaRESUMO
Non-isotopic whole-mount in situ hybridization of mRNA is a novel technique that has greatly facilitated the precise three-dimensional localization of transcripts from genes whose expression is important during development. This methodology has recently been applied to the study of the mouse embryo and offers particular advantages over conventional procedures.
Assuntos
Embrião de Mamíferos/metabolismo , Expressão Gênica , Hibridização In Situ/métodos , RNA Mensageiro/biossíntese , Animais , Camundongos , Inclusão do Tecido , Transcrição GênicaRESUMO
The anteroposterior axis of the vertebrate embryo becomes explicit during gastrulation, the process that converts a relatively featureless embryonic precursor population into new tissues assembled into a recognisable body pattern. Vertebrate embryos arrive at gastrulation in very different states in terms of their size, cell number and reliance on factors inherited from the unfertilized egg. However, they emerge from gastrulation looking very similar, and there is now extensive molecular genetic evidence to indicate that the bare essentials of the gastrulation process have been well conserved during evolution. Here, we review recent findings in the mouse that suggest that anterior identity is, in fact, established before gastrulation starts. They suggest that it is first manifest in extraembryonic tissue and that this tissue is essential for the embryo to develop normal anterior structures, such as the forebrain. We also argue that this precocious anterior pattern could have a counterpart in other non-mammalian vertebrates.
Assuntos
Padronização Corporal , Animais , Movimento Celular/genética , Ectoderma , Regulação da Expressão Gênica no Desenvolvimento , CamundongosRESUMO
A prospective fate map of the late gastrulation mouse primitive streak has been charted in 8.5 dpc mouse embryos developed in culture, using the lineage marker DiI to label groups of cells. As at earlier stages, the fate of cells in the 8.5 dpc primitive streak is regionalised such that successively more caudal regions of the streak give rise to more lateral mesoderm. While most labelled cells over a 24 or 48 h culture period exit from the primitive streak, some are consistently found to remain within it. The most conspicuous resident population is present in the node. To determine when ingression of ectoderm through the streak ceases, ectoderm cells of the streak and posterior neuropore of 8.5-10.0 dpc embryos were labelled. Involution of surface cells to form mesoderm continues until closure of the posterior neuropore but is not seen thereafter.
Assuntos
Gástrula/citologia , Marcadores de Afinidade , Animais , Carbocianinas , Linhagem da Célula , Ectoderma/citologia , Endoderma/citologia , Corantes Fluorescentes , Camundongos , MorfogêneseRESUMO
The midline has a theoretical role in the development of left-right asymmetry, and this is supported by both genetic analyses and experimental manipulation of midline structures in vertebrates. The mouse brachyury (T) gene encodes a transcription factor which is expressed in the developing notochord and is required for its development. T/T mice lack a mature notochord and have a dorsalised neural tube. We have examined the hearts of T/T mice and have found consistent morphological abnormalities, resulting in ventrally displaced ventricular loops, and a 50% incidence of inverted heart situs. Three TGF-beta related genes, lefty-1, lefty-2 and nodal, are expressed asymmetrically in mouse embryos, and are implicated in the development of situs. We find that nodal, which is normally expressed around the node and in left lateral plate mesoderm in early somite embryos, is completely absent at this stage in T/T embryos. In contrast, lefty-1 and lefty-2, which are normally expressed in the left half of prospective floorplate and left lateral plate mesoderm, respectively, are both expressed in T/T embryos only in a broad patch of ventral cells in, and just rostral to, the node region. These results implicate the node as a source of instructive signals driving expression of nodal and lefty-2 in the left lateral plate mesoderm, and being required for normal looping and situs of the heart.
Assuntos
Proteínas de Ligação a DNA/genética , Desenvolvimento Embrionário e Fetal/genética , Regulação da Expressão Gênica no Desenvolvimento , Cardiopatias Congênitas/genética , Coração/embriologia , Proteínas com Domínio T , Fatores de Transcrição/genética , Animais , Proteínas Fetais/genética , Fatores de Determinação Direita-Esquerda , Camundongos , Camundongos Mutantes , Miocárdio/patologia , Proteína Nodal , Fator de Crescimento Transformador beta/genéticaRESUMO
Msg1 and Mrg1 are founding members of a gene family which exhibit distinct patterns of gene expression during mouse embryogenesis. Sequence analysis reveals that these genes are unlike any other gene identified to date, but they share two near-identical sequence domains. The Msg1 and Mrg1 expression profiles during early development are distinct from each other. Msg1 is predominantly expressed in nascent mesoderm, the heart tube, limb bud and sclerotome. Intriguingly, Msg1 expression is restricted, within these developing mesodermal sites, to posterior domains. Mrg1 is expressed prior to gastrulation in the anterior visceral endoderm. Expression is maintained in the endoderm once gastrulation has begun and commences in the rostralmost embryonic mesoderm which underlies the anterior visceral endoderm. Mrg1 expression persists in this rostral mesoderm as it is translocated caudalwards during the invagination of the foregut and the formation of the heart. Later Mrg1 expression predominates in the septum transversum caudal to the heart. This expression pattern suggests that the septum transversum originates from the rostralmost embryonic mesoderm which first expressed Mrg1 at the late primitive streak stage.
Assuntos
Proteínas de Ligação a DNA/genética , Desenvolvimento Embrionário e Fetal/genética , Regulação da Expressão Gênica no Desenvolvimento , Família Multigênica/genética , Proteínas Nucleares/genética , Proteínas Repressoras , Transativadores/genética , Sequência de Aminoácidos , Animais , Proteínas Reguladoras de Apoptose , Sequência de Bases , Coração/embriologia , Humanos , Mesoderma/metabolismo , Camundongos , Dados de Sequência Molecular , Fatores de TranscriçãoRESUMO
An increasing amount of evidence suggests that in mouse there are two signalling centres required for the formation of a complete neural axis: the anterior visceral endoderm (AVE), and the node and its derivatives. Embryological and genetic studies suggest that the AVE has a head-inducing activity. In contrast, the node appears to act first as a head inducer in synergy with the AVE initiating anterior neural patterning at early stages of mouse development, and later, node derivatives are necessary for maintenance and embellishment of anterior neural character. Hex and Hesx1 are homeobox genes that are expressed in relevant tissues involved in anterior patterning. The analysis of the Hex and Hesx1 mutant mice has revealed that the lack of these genes has little or no effect on the early steps of anterior neural induction. However, both genes are required subsequently for the proper expansion of the forebrain region. We suggest that disturbance in the specification of an Fgf8 signalling centre in the anterior neural ridge may account for the anterior defects observed in these mutants.
Assuntos
Genes Homeobox , Proteínas de Homeodomínio/genética , Prosencéfalo/embriologia , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos , Padronização Corporal/genética , Indução Embrionária , Endoderma/citologia , Fator 8 de Crescimento de Fibroblasto , Fatores de Crescimento de Fibroblastos/fisiologia , Regulação da Expressão Gênica no Desenvolvimento , Camundongos , Camundongos Knockout , Proteínas Repressoras , Transdução de Sinais , Fatores de Transcrição HES-1 , Fatores de TranscriçãoRESUMO
The mouse T (Brachyury) deletion causes defective mesoderm formation and notochord morphogenesis, and abnormalities in the caudal neural tube and somites. To investigate the effect of the wild type T gene on concurrently expressed genes, we have compared expression of a panel of such genes in homozygous T mutants with that in wild type and heterozygous T/+ control embryos. Two classes of genes were used in this study: those implicated in primitive streak or mesoderm formation, and those which are differentially expressed in regions of the neural tube and somites. Results of wholemount in situ analysis show that the mRNA levels of Evx-1, Wnt-3a and Wnt-5a decrease in T/T embryos late in gastrulation, although earlier expression patterns are similar to control embryos. In contrast, BMP-4 and Msx-1 expression patterns remain similar throughout the period studied. Pax-3 and Pax-6, which are expressed in specific dorsoventral domains of the neural tube, both have ventrally extended expression domains in caudal T/T neural tube. This is consistent with a missing ventral signal provided by the notochord. However, the expression of Msx-1 in the most dorsal domain of the neural tube is unaltered in T/T embryos. Pax-1 and Pax-3, which are expressed in the sclerotome and dermamyotome respectively, are expressed correctly in anterior T/T somites, although the Pax-3 expression domain is widened ventromedially. This extension into ventromedial somite domains is more pronounced caudally, supporting a function for the notochord in ventralizing somites.
Assuntos
Embrião de Mamíferos/fisiologia , Desenvolvimento Embrionário e Fetal/genética , Deleção de Genes , Expressão Gênica , Proteínas de Homeodomínio , Mesoderma/fisiologia , Fatores de Transcrição , Anormalidades Múltiplas/genética , Animais , Sequência de Bases , Proteínas Morfogenéticas Ósseas , Primers do DNA , Proteínas de Ligação a DNA/biossíntese , Embrião de Mamíferos/citologia , Feminino , Genes Homeobox , Substâncias de Crescimento/biossíntese , Homozigoto , Fator de Transcrição MSX1 , Masculino , Camundongos , Camundongos Mutantes , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Gravidez , Biossíntese de Proteínas , RNA Mensageiro/análise , RNA Mensageiro/biossínteseRESUMO
The homeobox gene Hesx1, which encodes a pituitary transcription factor, is first expressed at gastrulation in the mouse embryo. Hesx1 expression begins in prospective forebrain tissue but later becomes restricted to Rathke's pouch, the primordium of the anterior pituitary gland. Transgenic mice lacking Hesx1 exhibit a phenotype comprising variable anterior CNS defects, such as a reduced prosencephalon, abnormalities in the corpus callosum and septum pellucidum, anophthalmia or microphthalmia, defective olfactory development and bifurcations in Rathke's pouch with pituitary dysplasia. A comparable and highly variable phenotype in humans is septo-optic dysplasia. We have cloned and sequenced the human homologue HESX1 and screened for mutations in affected individuals using single-stranded conformational polymorphism analysis. Two siblings with septo-optic dysplasia were homozygous for a missense mutation within the HESX1 homeobox. This mutation resulted in the substitution of a highly conserved arginine residue (Arg53) by cysteine and led to a loss of in vitro DNA binding. Hence, a vital role for Hesx1/HESX1 in forebrain and pituitary development in mice and humans is suggested.
Assuntos
Genes Homeobox , Sequências Hélice-Alça-Hélice/genética , Proteínas de Homeodomínio/genética , Septo Pelúcido/anormalidades , Animais , Arginina/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos , Cisteína/genética , Genótipo , Humanos , Mutação de Sentido Incorreto , Fenótipo , Adeno-Hipófise/fisiologia , Prosencéfalo/fisiologia , Proteínas Repressoras , Fatores de Transcrição HES-1 , Transcrição GênicaRESUMO
This paper describes a technique for transplanting early postimplantation mouse embryos from their own implantation site to a decidua in another pregnant mouse. Evidence is provided that this procedure is compatible with their continued development. At a low frequency both 6th and 7th day embryos can re-establish themselves and continue apparently normal development and placentation for at least 6-8 days.