RESUMO
The neural plate border (NPB) of vertebrate embryos is segregated from the neural plate (NP) and epidermal regions, and comprises an intermingled group of progenitors with multiple fate potential. Recent studies have shown that, during the gastrula stage, TFAP2A acts as a pioneer factor in remodeling the epigenetic landscape required to activate components of the NPB induction program. Here, we show that chick Tfap2a has two highly conserved binding sites for miR-137, and both display a reciprocal expression pattern at the NPB and NP, respectively. In addition, ectopic miR-137 expression reduced TFAP2A, whereas its functional inhibition expanded their territorial distribution overlapping with PAX7. Furthermore, we demonstrate that loss of the de novo DNA methyltransferase DNMT3A expanded miR-137 expression to the NPB. Bisulfite sequencing revealed a markedly elevated presence of non-canonical CpH methylation within the miR-137 promoter region when comparing NPB and NP samples. Our findings show that miR-137 contributes to the robustness of NPB territorial restriction in vertebrate development.
Assuntos
Metilação de DNA , Regulação da Expressão Gênica no Desenvolvimento , MicroRNAs , Placa Neural , Fator de Transcrição AP-2 , Animais , MicroRNAs/genética , MicroRNAs/metabolismo , Embrião de Galinha , Metilação de DNA/genética , Placa Neural/metabolismo , Placa Neural/embriologia , Fator de Transcrição AP-2/metabolismo , Fator de Transcrição AP-2/genética , DNA (Citosina-5-)-Metiltransferases/metabolismo , DNA (Citosina-5-)-Metiltransferases/genética , DNA Metiltransferase 3A/metabolismo , Regiões Promotoras Genéticas/genética , Sítios de LigaçãoRESUMO
Neurulation involves a complex coordination of cellular movements that are in great part based on the modulation of the actin cytoskeleton. MARCKS, an F-actin-binding protein and the major substrate for PKC, is necessary for gastrulation and neurulation morphogenetic movements in mice, frogs, and fish. We previously showed that this protein accumulates at the apical region of the closing neural plate in chick embryos, and here further explore its role in this process and how it is regulated by PKC phosphorylation. PKC activation by PMA caused extensive neural tube closure defects in cultured chick embryos, together with MARCKS phosphorylation and redistribution to the cytoplasm. This was concomitant with an evident disruption of neural plate cell polarity and extensive apical cell extrusion. This effect was not due to actomyosin hypercontractility, but it was reproduced upon MARCKS knockdown. Interestingly, the overexpression of a nonphosphorylatable form of MARCKS was able to revert the cellular defects observed in the neural plate after PKC activation. Altogether, these results suggest that MARCKS function during neurulation would be to maintain neuroepithelial polarity through the stabilization of subapical F-actin, a function that appears to be counteracted by PKC activation.
Assuntos
Substrato Quinase C Rico em Alanina Miristoilada/metabolismo , Substrato Quinase C Rico em Alanina Miristoilada/fisiologia , Neurulação/fisiologia , Citoesqueleto de Actina/metabolismo , Actinas/metabolismo , Animais , Proteínas de Transporte/metabolismo , Polaridade Celular/fisiologia , Embrião de Galinha , Galinhas/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas de Membrana/metabolismo , Proteínas dos Microfilamentos/metabolismo , Placa Neural/metabolismo , Neurulação/genética , Fosforilação , Proteína Quinase C/metabolismo , Proteína Quinase C/fisiologia , Transdução de SinaisRESUMO
Collective cell migration is essential in many fundamental aspects of normal development, like morphogenesis, organ formation, wound healing, and immune responses, as well as in the etiology of severe pathologies, like cancer metastasis. In spite of the huge amount of data accumulated on cell migration, such a complex process involves many molecular actors, some of which still remain to be functionally characterized. One of these signals is the heterotrimeric G-protein pathway that has been studied mainly in gastrulation movements. Recently we have reported that Ric-8A, a GEF for Gα proteins, plays an important role in neural crest migration in Xenopus development. Xenopus neural crest cells, a highly migratory embryonic cell population induced at the border of the neural plate that migrates extensively in order to differentiate in other tissues during development, have become a good model to understand the dynamics that regulate cell migration. In this review, we aim to provide sufficient evidence supporting how useful Xenopus model with its different tools, such as explants and transplants, paired with improved in vivo imaging techniques, will allow us to tackle the multiple signaling mechanisms involved in neural crest cell migration.
Assuntos
Movimento Celular/genética , Proteínas Heterotriméricas de Ligação ao GTP/genética , Morfogênese/genética , Xenopus laevis/genética , Animais , Regulação da Expressão Gênica no Desenvolvimento , Proteínas Heterotriméricas de Ligação ao GTP/metabolismo , Crista Neural/crescimento & desenvolvimento , Crista Neural/metabolismo , Placa Neural/crescimento & desenvolvimento , Placa Neural/metabolismo , Transdução de Sinais/genética , Xenopus laevis/crescimento & desenvolvimentoRESUMO
One of the earliest manifestations of neural induction is onset of expression of the neural marker Sox2, mediated by the activation of the enhancers N1 and N2. By using loss and gain of function, we find that Sox2 expression requires the activity of JmjD2A and the Msk1 kinase, which can respectively demethylate the repressive H3K9me3 mark and phosphorylate the activating H3S10 (H3S10ph) mark. Bimolecular fluorescence complementation reveals that the adaptor protein 14-3-3, known to bind to H3S10ph, interacts with JMJD2A and may be involved in its recruitment to regulatory regions of the Sox2 gene. Chromatin immunoprecipitation reveals dynamic binding of JMJD2A to the Sox2 promoter and N-1 enhancer at the time of neural plate induction. Finally, we show a clear temporal antagonism on the occupancy of H3K9me3 and H3S10ph modifications at the promoter of the Sox2 locus before and after the neural plate induction. Taken together, our results propose a series of epigenetic events necessary for the early activation of the Sox2 gene in neural progenitor cells.
Assuntos
Histona Desmetilases com o Domínio Jumonji/metabolismo , Fatores de Transcrição SOXB1/genética , Fatores de Transcrição SOXB1/metabolismo , Proteínas 14-3-3/metabolismo , Animais , Embrião de Galinha , Desenvolvimento Embrionário/genética , Elementos Facilitadores Genéticos/genética , Epigenômica , Regulação da Expressão Gênica no Desenvolvimento/genética , Placa Neural/metabolismo , Células-Tronco Neurais/metabolismo , Fosforilação , Regiões Promotoras Genéticas , Proteínas Quinases S6 Ribossômicas 90-kDa/metabolismo , Fatores de Transcrição/metabolismoRESUMO
BACKGROUND: During the initial stages zebrafish neurulation, neural plate cells undergo highly coordinated movements before they assemble into a multicellular solid neural rod. We have previously identified that the underlying mesoderm is critical to ensure such coordination and generate correct neural tube organization. However, how intertissue coordination is achieved in vivo during zebrafish neural tube morphogenesis is unknown. RESULTS: In this work, we use quantitative live imaging to study the coordinated movements of neural ectoderm and mesoderm during dorsal tissue convergence. We show the extracellular matrix components laminin and fibronectin that lie between mesoderm and neural plate act to couple the movements of neural plate and mesoderm during early stages of neurulation and to maintain the close apposition of these two tissues. CONCLUSIONS: Our study highlights the importance of the extracellular matrix proteins laminin and libronectin in coupling the movements and spatial proximity of mesoderm and neuroectoderm during the morphogenetic movements of neurulation. Developmental Dynamics 245:580-589, 2016. © 2016 Wiley Periodicals, Inc.
Assuntos
Matriz Extracelular/fisiologia , Mesoderma/metabolismo , Placa Neural/metabolismo , Neurulação , Animais , Embrião não Mamífero , Fibronectinas/fisiologia , Laminina/fisiologia , Morfogênese , Tubo Neural , Peixe-ZebraRESUMO
In vertebrates, the embryonic dorsal midline is a crucial signalling centre that patterns the surrounding tissues during development. Members of the FoxA subfamily of transcription factors are expressed in the structures that compose this centre. Foxa2 is essential for dorsal midline development in mammals, since knock-out mouse embryos lack a definitive node, notochord and floor plate. The related gene foxA4 is only present in amphibians. Expression begins in the blastula -chordin and -noggin expressing centre (BCNE) and is later restricted to the dorsal midline derivatives of the Spemann's organiser. It was suggested that the early functions of mammalian foxa2 are carried out by foxA4 in frogs, but functional experiments were needed to test this hypothesis. Here, we show that some important dorsal midline functions of mammalian foxa2 are exerted by foxA4 in Xenopus. We provide new evidence that the latter prevents the respecification of dorsal midline precursors towards contiguous fates, inhibiting prechordal and paraxial mesoderm development in favour of the notochord. In addition, we show that foxA4 is required for the correct regionalisation and maintenance of the central nervous system. FoxA4 participates in constraining the prospective rostral forebrain territory during neural specification and is necessary for the correct segregation of the most anterior ectodermal derivatives, such as the cement gland and the pituitary anlagen. Moreover, the early expression of foxA4 in the BCNE (which contains precursors of the whole forebrain and most of the midbrain and hindbrain) is directly required to restrict anterior neural development.