RESUMO
The neural fate is generally considered to be the intrinsic direction of embryonic stem (ES) cell differentiation. However, little is known about the intracellular mechanism that leads undifferentiated cells to adopt the neural fate in the absence of extrinsic inductive signals. Here we show that the zinc-finger nuclear protein Zfp521 is essential and sufficient for driving the intrinsic neural differentiation of mouse ES cells. In the absence of the neural differentiation inhibitor BMP4, strong Zfp521 expression is intrinsically induced in differentiating ES cells. Forced expression of Zfp521 enables the neural conversion of ES cells even in the presence of BMP4. Conversely, in differentiation culture, Zfp521-depleted ES cells do not undergo neural conversion but tend to halt at the epiblast state. Zfp521 directly activates early neural genes by working with the co-activator p300. Thus, the transition of ES cell differentiation from the epiblast state into neuroectodermal progenitors specifically depends on the cell-intrinsic expression and activator function of Zfp521.
Assuntos
Diferenciação Celular , Células-Tronco Embrionárias/citologia , Células-Tronco Neurais/citologia , Fatores de Transcrição/metabolismo , Animais , Proteína Morfogenética Óssea 4/deficiência , Proteína Morfogenética Óssea 4/genética , Proteína Morfogenética Óssea 4/metabolismo , Caderinas/metabolismo , Linhagem da Célula , Células Cultivadas , Embrião de Mamíferos/citologia , Embrião de Mamíferos/embriologia , Embrião de Mamíferos/metabolismo , Células-Tronco Embrionárias/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/genética , Camadas Germinativas/citologia , Camadas Germinativas/embriologia , Camadas Germinativas/metabolismo , Células HEK293 , Humanos , Camundongos , Modelos Biológicos , Placa Neural/citologia , Placa Neural/embriologia , Placa Neural/metabolismo , Células-Tronco Neurais/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , Fatores de Transcrição SOXB1/metabolismo , Fatores de Transcrição/deficiência , Fatores de Transcrição/genética , Ativação Transcricional , Xenopus , Fatores de Transcrição de p300-CBP/metabolismoRESUMO
During embryogenesis, bone morphogenetic protein (BMP) signaling needs to be finely tuned in a locally restricted manner. Here, we report a cell-intrinsic mode of BMP response control executed by the membrane protein Jiraiya. In the Xenopus embryo, zygotic Jiraiya, expressed exclusively in the neuroectoderm, is essential and sufficient for limiting dorsal neural development, which is dependent on BMP signals. In animal cap assays, Jiraiya selectively and cell-autonomously inhibits BMP signaling, while Jiraiya's knockdown enhances the signaling. In the cell, Jiraiya selectively forms a complex with type II BMP receptor (BMPRII) and downregulates the cell surface localization of functional BMPRII. This functional interaction with Jiraiya depends on the unique tail domain of BMPRII, and, in particular, the conserved EVNNNG motif, the function of which has been unknown. Thus, Jiraiya represents a cell-intrinsic cutoff mechanism for dynamic responsiveness to BMP signals via subtype-selective receptor control.
Assuntos
Padronização Corporal , Receptores de Proteínas Morfogenéticas Ósseas Tipo II/metabolismo , Proteínas Morfogenéticas Ósseas/metabolismo , Proteínas de Membrana/metabolismo , Placa Neural/embriologia , Transdução de Sinais , Proteínas de Xenopus/metabolismo , Xenopus/embriologia , Motivos de Aminoácidos , Sequência de Aminoácidos , Animais , Padronização Corporal/genética , Receptores de Proteínas Morfogenéticas Ósseas Tipo II/química , Membrana Celular/metabolismo , Sequência Conservada , Embrião não Mamífero/citologia , Embrião não Mamífero/metabolismo , Retículo Endoplasmático/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Proteínas de Membrana/genética , Dados de Sequência Molecular , Sistema Nervoso/citologia , Sistema Nervoso/embriologia , Sistema Nervoso/metabolismo , Placa Neural/metabolismo , Especificidade de Órgãos , Ligação Proteica , Estrutura Terciária de Proteína , Transporte Proteico , Transdução de Sinais/genética , Relação Estrutura-Atividade , Xenopus/genética , Xenopus/metabolismo , Proteínas de Xenopus/genéticaRESUMO
During early embryogenesis, the neural plate is specified along the anterior-posterior (AP) axis by the action of graded patterning signals. In particular, the attenuation of canonical Wnt signals plays a central role in the determination of the anterior brain region. Here, we show that the extracellular matrix (ECM) protein Del1, expressed in the anterior neural plate, is essential for forebrain development in the Xenopus embryo. Overexpression of Del1 expands the forebrain domain and promotes the formation of head structures, such as the eye, in a Chordin-induced secondary axis. Conversely, the inhibition of Del1 function by a morpholino oligonucleotide (MO) represses forebrain development. Del1 also augments the expression of forebrain markers in neuralized animal cap cells, whereas Del1-MO suppresses them. We previously reported that Del1 interferes with BMP signaling in the dorsal-ventral patterning of the gastrula marginal zone. By contrast, we demonstrate here that Del1 function in AP neural patterning is mediated mainly by the inhibition of canonical Wnt signaling. Wnt-induced posteriorization of the neural plate is counteracted by Del1, and the Del1-MO phenotype (posteriorization) is reversed by Dkk1. Topflash reporter assays show that Del1 suppresses luciferase activities induced by Wnt1 and beta-catenin. This inhibitory effect of Del1 on canonical Wnt signaling, but not on BMP signaling, requires the Ror2 pathway, which is implicated in non-canonical Wnt signaling. These findings indicate that the ECM protein Del1 promotes forebrain development by creating a local environment that attenuates the cellular response to posteriorizing Wnt signals via a unique pathway.
Assuntos
Proteínas de Membrana/metabolismo , Placa Neural/metabolismo , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo , Transdução de Sinais , Proteínas de Xenopus/metabolismo , Xenopus laevis/embriologia , Xenopus laevis/metabolismo , Transporte Ativo do Núcleo Celular , Animais , Padronização Corporal , Cabeça/embriologia , Proteínas de Membrana/genética , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/genética , Proteínas Wnt/metabolismo , Proteínas de Xenopus/genética , beta Catenina/metabolismoRESUMO
During gastrulation of the amphibian embryo, specification of the three germ layers, endo-, ecto-, and mesoderm, is regulated by maternal and zygotic mechanisms. Although it is known that mesoderm specification requires the cooperation between TGF-beta signaling and p53 activity and requires maternal factors, essential zygotic factors have been elusive. Here, we report that the Zn-finger protein XFDL156 is an ectodermal, zygotic factor that suppresses mesodermal differentiation. XFDL156 overexpression suppresses mesodermal markers, and its depletion induces aberrant mesodermal differentiation in the presumptive ectoderm. Furthermore, we find that XFDL156 and its mammalian homologs interact with the C-terminal regulatory region of p53, thereby inhibiting p53 target gene induction and mesodermal differentiation. Thus, XFDL156 actively restricts mesodermal differentiation in the presumptive ectoderm by controlling the spatiotemporal responsiveness to p53.
Assuntos
Proteínas de Transporte/metabolismo , Diferenciação Celular , Mesoderma/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Proteínas de Xenopus/metabolismo , Xenopus laevis/embriologia , Animais , Linhagem Celular , Linhagem Celular Tumoral , Ectoderma/citologia , Ectoderma/metabolismo , Embrião não Mamífero/metabolismo , Humanos , Mesoderma/citologia , Estrutura Terciária de Proteína , Fator de Crescimento Transformador beta/metabolismo , Proteína Supressora de Tumor p53/química , Xenopus laevis/metabolismoRESUMO
Using a large-scale in situ hybridization screening system, we found that mRNA coding for ME1, a basic helix-loop-helix (bHLH) transcription factor, was abundantly expressed in Paneth cells of adult small intestinal crypts. Other functionally related E-protein mRNAs, ME2, and E2A, however, could not be detected in the cells. ME1 mRNA was first detected in the jejunum and ileum two weeks after birth when the number of Paneth cells starts to increase. ME1 is the first identified bHLH transcription factor expressed in the Paneth cells and may be used as a molecular marker and a key molecule for analyzing transcriptional regulation in the Paneth cell.
Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Mucosa Intestinal/fisiologia , Camundongos/genética , Celulas de Paneth/fisiologia , RNA Mensageiro/genética , Animais , Sequência de Bases , Clonagem Molecular , DNA Complementar/genética , Regulação da Expressão Gênica , Hibridização In Situ , Jejuno/crescimento & desenvolvimento , Jejuno/fisiologia , Camundongos/fisiologiaRESUMO
In Xenopus, an asymmetric distribution of Wnt activity that follows cortical rotation in the fertilized egg leads to the dorsal-ventral (DV) axis establishment. However, how a clear DV polarity develops from the initial difference in Wnt activity still remains elusive. We report here that the Teashirt-class Zn-finger factor XTsh3 plays an essential role in dorsal determination by enhancing canonical Wnt signaling. Knockdown of the XTsh3 function causes ventralization in the Xenopus embryo. Both in vivo and in vitro studies show that XTsh3 substantially enhances Wnt signaling activity in a beta-catenin-dependent manner. XTsh3 cooperatively promotes the formation of a secondary axis on the ventral side when combined with weak Wnt activity, whereas XTsh3 alone has little axis-inducing ability. Furthermore, Wnt1 requires XTsh3 for its dorsalizing activity in vivo. Immunostaining and protein analyses indicate that XTsh3 is a nuclear protein that physically associates with beta-catenin and efficiently increases the level of beta-catenin in the nucleus. We discuss the role of XTsh3 as an essential amplifying factor of canonical Wnt signaling in embryonic dorsal determination.