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1.
Hum Mol Genet ; 31(24): 4217-4227, 2022 12 16.
Artigo em Inglês | MEDLINE | ID: mdl-35899771

RESUMO

Ets1 deletion in some mouse strains causes septal defects and has been implicated in human congenital heart defects in Jacobsen syndrome, in which one copy of the Ets1 gene is missing. Here, we demonstrate that loss of Ets1 in mice results in a decrease in neural crest (NC) cells migrating into the proximal outflow tract cushions during early heart development, with subsequent malalignment of the cushions relative to the muscular ventricular septum, resembling double outlet right ventricle (DORV) defects in humans. Consistent with this, we find that cultured cardiac NC cells from Ets1 mutant mice or derived from iPS cells from Jacobsen patients exhibit decreased migration speed and impaired cell-to-cell interactions. Together, our studies demonstrate a critical role for ETS1 for cell migration in cardiac NC cells that are required for proper formation of the proximal outflow tracts. These data provide further insights into the molecular and cellular basis for development of the outflow tracts, and how perturbation of NC cells can lead to DORV.


Assuntos
Cardiopatias Congênitas , Crista Neural , Proteína Proto-Oncogênica c-ets-1 , Animais , Humanos , Camundongos , Movimento Celular/genética , Coração , Organogênese , Proteína Proto-Oncogênica c-ets-1/genética
2.
Dev Dyn ; 247(9): 1083-1092, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-30079980

RESUMO

BACKGROUND: Neural crest is a vertebrate specific cell population. Induced at lateral borders of the neural plate, neural crest cells (NCCs) subsequently undergo epithelial-to-mesenchymal transition (EMT) to detach from the neuroepithelium before migrating into various locations in the embryo. Despite the wealth of knowledge of transcription factors involved in this process, little is known about the effectors that directly regulate neural crest EMT and migration. RESULTS: Here, we examined the activity of matrix metalloproteinase MMP14 in NCCs and found that MMP14 is expressed in both premigratory and migrating NCCs. Overexpression of MMP14 led to premature migration of NCCs, while down-regulation of MMP14 resulted in reduced neural crest migration. Transplantation experiment further showed that MMP14 is required in NCCs, whereas MMP2, which can be activated by MMP14, is required in the surrounding mesenchyme. in vitro explant culture showed that MMP14 is required for neural crest EMT but not for spreading. This is possibly mediated by the changes in cadherin levels, as decreasing MMP14 level led to increased cadherin expression and increasing MMP14 level led to reduced cadherin expression. CONCLUSIONS: The results demonstrate that MMP14 is critical for neural crest EMT and migration, partially through regulating the levels of cadherins. Developmental Dynamics 247:1083-1092, 2018. © 2018 Wiley Periodicals, Inc.


Assuntos
Movimento Celular , Transição Epitelial-Mesenquimal , Metaloproteinase 14 da Matriz/fisiologia , Crista Neural/citologia , Xenopus/embriologia , Animais , Caderinas/metabolismo , Embrião não Mamífero/citologia , Desenvolvimento Embrionário , Metaloproteinase 14 da Matriz/metabolismo , Crista Neural/metabolismo , Crânio/citologia
3.
Nature ; 449(7165): 1068-72, 2007 Oct 25.
Artigo em Inglês | MEDLINE | ID: mdl-17914355

RESUMO

Post-translational histone modifications have important regulatory roles in chromatin structure and function. One example of such modifications is histone ubiquitination, which occurs predominately on histone H2A and H2B. Although the recent identification of the ubiquitin ligase for histone H2A has revealed important roles for H2A ubiquitination in Hox gene silencing as well as in X-chromosome inactivation, the enzyme(s) involved in H2A deubiquitination and the function of H2A deubiquitination are not known. Here we report the identification and functional characterization of the major deubiquitinase for histone H2A, Ubp-M (also called USP16). Ubp-M prefers nucleosomal substrates in vitro, and specifically deubiquitinates histone H2A but not H2B in vitro and in vivo. Notably, knockdown of Ubp-M in HeLa cells results in slow cell growth rates owing to defects in the mitotic phase of the cell cycle. Further studies reveal that H2A deubiquitination by Ubp-M is a prerequisite for subsequent phosphorylation of Ser 10 of H3 and chromosome segregation when cells enter mitosis. Furthermore, we demonstrate that Ubp-M regulates Hox gene expression through H2A deubiquitination and that blocking the function of Ubp-M results in defective posterior development in Xenopus laevis. This study identifies the major deubiquitinase for histone H2A and demonstrates that H2A deubiquitination is critically involved in cell cycle progression and gene expression.


Assuntos
Ciclo Celular/fisiologia , Endopeptidases/metabolismo , Regulação da Expressão Gênica , Histonas/metabolismo , Ubiquitina Tiolesterase/metabolismo , Ubiquitinação , Proteínas de Xenopus/metabolismo , Animais , Divisão Celular , Endopeptidases/deficiência , Endopeptidases/genética , Genes Homeobox/genética , Células HeLa , Histonas/química , Proteínas de Homeodomínio/genética , Humanos , Fosfosserina/metabolismo , Especificidade por Substrato , Fatores de Transcrição/genética , Ubiquitina Tiolesterase/deficiência , Ubiquitina Tiolesterase/genética , Proteínas de Xenopus/deficiência , Proteínas de Xenopus/genética , Xenopus laevis/embriologia , Xenopus laevis/genética
4.
J Cardiovasc Dev Dis ; 10(2)2023 Jan 29.
Artigo em Inglês | MEDLINE | ID: mdl-36826547

RESUMO

A frog is a classical model organism used to uncover processes and regulations of early vertebrate development, including heart development. Recently, we showed that a frog also represents a useful model to study a rare human congenital heart disease, hypoplastic left heart syndrome. In this review, we first summarized the cellular events and molecular regulations of vertebrate heart development, and the benefit of using a frog model to study congenital heart diseases. Next, we described the challenges in elucidating the etiology of hypoplastic left heart syndrome and discussed how a frog model may contribute to our understanding of the molecular and cellular bases of the disease. We concluded that a frog model offers its unique advantage in uncovering the cellular mechanisms of hypoplastic left heart syndrome; however, combining multiple model organisms, including frogs, is needed to gain a comprehensive understanding of the disease.

5.
Front Cell Dev Biol ; 11: 1106595, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36923257

RESUMO

The septin cytoskeleton has been demonstrated to interact with other cytoskeletal components to regulate various cellular processes, including cell migration. However, the mechanisms of how septin regulates cell migration are not fully understood. In this study, we use the highly migratory neural crest cells of frog embryos to examine the role of septin filaments in cell migration. We found that septin filaments are required for the proper migration of neural crest cells by controlling both the speed and the direction of cell migration. We further determined that septin filaments regulate these features of cell migration by interacting with actin stress fibers. In neural crest cells, septin filaments co-align with actin stress fibers, and the loss of septin filaments leads to impaired stability and contractility of actin stress fibers. In addition, we showed that a partial loss of septin filaments leads to drastic changes in the orientations of newly formed actin stress fibers, suggesting that septin filaments help maintain the persistent orientation of actin stress fibers during directed cell migration. Lastly, our study revealed that these activities of septin filaments depend on Cdc42ep1, which colocalizes with septin filaments in the center of neural crest cells. Cdc42ep1 interacts with septin filaments in a reciprocal manner, with septin filaments recruiting Cdc42ep1 to the cell center and Cdc42ep1 supporting the formation of septin filaments.

6.
bioRxiv ; 2023 Mar 31.
Artigo em Inglês | MEDLINE | ID: mdl-37034645

RESUMO

Rho-GTPases are central regulators within a complex signaling network that controls the cytoskeletal organization and cell movement. This network includes multiple GTPases, such as the most studied Rac1, Cdc42, and RhoA, and their numerous effectors that provide mutual regulation and feedback loops. Here we investigate the temporal and spatial relationship between Rac1 and Cdc42 during membrane ruffling using a simulation model which couples GTPase signaling with cell morphodynamics to capture the GTPase behavior observed with FRET-based biosensors. We show that membrane velocity is regulated by the kinetic rate of GTPase activation rather than the concentration of active GTPase. Our model captures both uniform and polarized ruffling. We also show that cell-type specific time delays between Rac1 and Cdc42 activation can be reproduced with a single signaling motif, in which the delay is controlled by feedback from Cdc42 to Rac1. The resolution of our simulation output matches those of the time-lapsed recordings of cell dynamics and GTPase activity. This approach allows us to validate simulation results with quantitative precision using the same pipeline for the analysis of simulated and experimental data.

7.
Cells ; 12(12)2023 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-37371108

RESUMO

Rho-GTPases are central regulators within a complex signaling network that controls cytoskeletal organization and cell movement. The network includes multiple GTPases, such as the most studied Rac1, Cdc42, and RhoA, along with their numerous effectors that provide mutual regulation through feedback loops. Here we investigate the temporal and spatial relationship between Rac1 and Cdc42 during membrane ruffling, using a simulation model that couples GTPase signaling with cell morphodynamics and captures the GTPase behavior observed with FRET-based biosensors. We show that membrane velocity is regulated by the kinetic rate of GTPase activation rather than the concentration of active GTPase. Our model captures both uniform and polarized ruffling. We also show that cell-type specific time delays between Rac1 and Cdc42 activation can be reproduced with a single signaling motif, in which the delay is controlled by feedback from Cdc42 to Rac1. The resolution of our simulation output matches those of time-lapsed recordings of cell dynamics and GTPase activity. Our data-driven modeling approach allows us to validate simulation results with quantitative precision using the same pipeline for the analysis of simulated and experimental data.


Assuntos
Membrana Celular , Movimento Celular , Proteínas rac1 de Ligação ao GTP , Proteínas rho de Ligação ao GTP , Membrana Celular/metabolismo , Membrana Celular/fisiologia , Movimento Celular/genética , Movimento Celular/fisiologia , Proteínas rac1 de Ligação ao GTP/metabolismo , Proteínas rho de Ligação ao GTP/metabolismo , Transdução de Sinais
8.
Sci Adv ; 9(35): eadg9245, 2023 09.
Artigo em Inglês | MEDLINE | ID: mdl-37647399

RESUMO

Fluorescence microscopy is one of the most indispensable and informative driving forces for biological research, but the extent of observable biological phenomena is essentially determined by the content and quality of the acquired images. To address the different noise sources that can degrade these images, we introduce an algorithm for multiscale image restoration through optimally sparse representation (MIRO). MIRO is a deterministic framework that models the acquisition process and uses pixelwise noise correction to improve image quality. Our study demonstrates that this approach yields a remarkable restoration of the fluorescence signal for a wide range of microscopy systems, regardless of the detector used (e.g., electron-multiplying charge-coupled device, scientific complementary metal-oxide semiconductor, or photomultiplier tube). MIRO improves current imaging capabilities, enabling fast, low-light optical microscopy, accurate image analysis, and robust machine intelligence when integrated with deep neural networks. This expands the range of biological knowledge that can be obtained from fluorescence microscopy.


Assuntos
Algoritmos , Elétrons , Microscopia de Fluorescência , Processamento de Imagem Assistida por Computador , Redes Neurais de Computação
9.
Methods Mol Biol ; 2438: 517-526, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35147961

RESUMO

The neural crest is a highly migratory cell population that evolved in vertebrates. Born at the lateral borders of the neural plate, neural crest cells migrate long distances along defined paths and contribute to the development of many tissue and structures. Neural crest has become an important model for studying directional cell migration. Frog Xenopus laevis is particularly feasible in these studies. Both in vivo and in vitro analyses are performed to study frog neural crest cell migration. While in vivo analysis can provide direct knowledge of how neural crest cells interact with neighboring tissues during their migration, in vitro analysis can produce high-resolution results on cell morphological changes and cell motility. Here we provide a detailed protocol for performing quantitative analysis of Xenopus laevis neural crest cell migration in vitro.


Assuntos
Crista Neural , Placa Neural , Animais , Diferenciação Celular , Movimento Celular , Xenopus laevis
10.
Sci Rep ; 11(1): 19512, 2021 09 30.
Artigo em Inglês | MEDLINE | ID: mdl-34593939

RESUMO

The Rho family GTPases are molecular switches that regulate cytoskeletal dynamics and cell movement through a complex spatiotemporal organization of their activity. In Patiria miniata (starfish) oocytes under in vitro experimental conditions (with overexpressed Ect2, induced expression of Δ90 cyclin B, and roscovitine treatment), such activity generates multiple co-existing regions of coherent propagation of actin waves. Here we use computational modeling to investigate the development and properties of such wave domains. The model reveals that the formation of wave domains requires a balance between the activation and inhibition in the Rho signaling motif. Intriguingly, the development of the wave domains is preceded by a stage of low-activity quasi-static patterns, which may not be readily observed in experiments. Spatiotemporal patterns of this stage and the different paths of their destabilization define the behavior of the system in the later high-activity (observable) stage. Accounting for a strong intrinsic noise allowed us to achieve good quantitative agreement between simulated dynamics in different parameter regimes of the model and different wave dynamics in Patiria miniata and wild type Xenopus laevis (frog) data. For quantitative comparison of simulated and experimental results, we developed an automated method of wave domain detection, which revealed a sharp reversal in the process of pattern formation in starfish oocytes. Overall, our findings provide an insight into spatiotemporal regulation of complex and diverse but still computationally reproducible cell-level actin dynamics.


Assuntos
Modelos Moleculares , Domínios e Motivos de Interação entre Proteínas , Proteínas rho de Ligação ao GTP/química , Proteínas rho de Ligação ao GTP/metabolismo , Algoritmos , Animais , Ativação Enzimática , Oócitos/metabolismo , Estrelas-do-Mar , Relação Estrutura-Atividade , Imagem com Lapso de Tempo
11.
Dev Biol ; 335(1): 132-42, 2009 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-19712673

RESUMO

The neural crest is a highly migratory cell population, unique to vertebrates, that forms much of the craniofacial skeleton and peripheral nervous system. In exploring the cell biological basis underlying this behavior, we have identified an unconventional myosin, myosin-X (Myo10) that is required for neural crest migration. Myo10 is highly expressed in both premigratory and migrating cranial neural crest (CNC) cells in Xenopus embryos. Disrupting Myo10 expression using antisense morpholino oligonucleotides leads to impaired neural crest migration and subsequent cartilage formation, but only a slight delay in induction. In vivo grafting experiments reveal that Myo10-depleted CNC cells migrate a shorter distance and fail to segregate into distinct migratory streams. Finally, in vitro cultures and cell dissociation-reaggregation assays suggest that Myo10 may be critical for cell protrusion and cell-cell adhesion. These results demonstrate an essential role for Myo10 in normal cranial neural crest migration and suggest a link to cell-cell interactions and formation of processes.


Assuntos
Movimento Celular/fisiologia , Miosinas/metabolismo , Crista Neural/citologia , Crânio , Proteínas de Xenopus/metabolismo , Xenopus laevis , Animais , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Adesão Celular/fisiologia , Extensões da Superfície Celular/metabolismo , Indução Embrionária , Técnicas de Silenciamento de Genes , Miosinas/genética , Oligonucleotídeos Antissenso/genética , Oligonucleotídeos Antissenso/metabolismo , Crânio/citologia , Crânio/embriologia , Transplantes , Proteína 1 Relacionada a Twist/genética , Proteína 1 Relacionada a Twist/metabolismo , Proteínas de Xenopus/genética , Xenopus laevis/anatomia & histologia , Xenopus laevis/embriologia , Xenopus laevis/metabolismo
12.
Cell Stem Cell ; 27(4): 574-589.e8, 2020 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-32810435

RESUMO

Hypoplastic left heart syndrome (HLHS) is a complex congenital heart disease characterized by abnormalities in the left ventricle, associated valves, and ascending aorta. Studies have shown intrinsic myocardial defects but do not sufficiently explain developmental defects in the endocardial-derived cardiac valve, septum, and vasculature. Here, we identify a developmentally impaired endocardial population in HLHS through single-cell RNA profiling of hiPSC-derived endocardium and human fetal heart tissue with an underdeveloped left ventricle. Intrinsic endocardial defects contribute to abnormal endothelial-to-mesenchymal transition, NOTCH signaling, and extracellular matrix organization, key factors in valve formation. Endocardial abnormalities cause reduced cardiomyocyte proliferation and maturation by disrupting fibronectin-integrin signaling, consistent with recently described de novo HLHS mutations associated with abnormal endocardial gene and fibronectin regulation. Together, these results reveal a critical role for endocardium in HLHS etiology and provide a rationale for considering endocardial function in regenerative strategies.


Assuntos
Síndrome do Coração Esquerdo Hipoplásico , Células-Tronco Pluripotentes Induzidas , Endocárdio , Humanos , Miocárdio , Transdução de Sinais
13.
Front Physiol ; 10: 542, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31133876

RESUMO

Somitogenesis is a critical process during vertebrate development that establishes the segmented body plan and gives rise to the vertebra, skeletal muscles, and dermis. While segmentation clock and wave front mechanisms have been elucidated to control the size and time of somite formation, regulation of the segmentation process that physically separates somites is not understood in detail. Here, we identified a cytoskeletal player, Cdc42 effector protein 3 (Cdc42ep3, CEP3) that is required for somite segmentation in Xenopus embryos. CEP3 is specifically expressed in somite tissue during somite segmentation. Loss-of-function experiments showed that CEP3 is not required for the specification of paraxial mesoderm, nor the differentiation of muscle cells, but is required for the segmentation process. Live imaging analysis further revealed that CEP3 is required for cell shape changes and alignment during somitogenesis. When CEP3 was knocked down, somitic cells did not elongate efficiently along the mediolateral axis and failed to undertake the 90° rotation. As a result, cells remained in a continuous sheet without an apparent segmentation cleft. CEP3 likely interacts with Cdc42 during this process, and both increased and decreased Cdc42 activity led to defective somite segmentation. Segmentation defects caused by Cdc42 knockdown can be partially rescued by the overexpression of CEP3. Conversely, loss of CEP3 resulted in the maintenance of high levels of Cdc42 activity at the cell membrane, which is normally reduced during and after somite segmentation. These results suggest that there is a feedback regulation between Cdc42 and CEP3 during somite segmentation and the activity of Cdc42 needs to be fine-tuned to control the coordinated cell shape changes and movement required for somite segmentation.

14.
J Cardiovasc Dev Dis ; 6(1)2019 Feb 23.
Artigo em Inglês | MEDLINE | ID: mdl-30813450

RESUMO

Hypoplastic left heart syndrome occurs in up to 3% of all infants born with congenital heart disease and is a leading cause of death in this population. Although there is strong evidence for a genetic component, a specific genetic cause is only known in a small subset of patients, consistent with a multifactorial etiology for the syndrome. There is controversy surrounding the mechanisms underlying the syndrome, which is likely due, in part, to the phenotypic variability of the disease. The most commonly held view is that the "decreased" growth of the left ventricle is due to a decreased flow during a critical period of ventricular development. Research has also been hindered by what has been, up until now, a lack of genetically engineered animal models that faithfully reproduce the human disease. There is a growing body of evidence, nonetheless, indicating that the hypoplasia of the left ventricle is due to a primary defect in ventricular development. In this review, we discuss the evidence demonstrating that, at least for a subset of cases, the chamber hypoplasia is the consequence of hyperplasia of the contained cardiomyocytes. In this regard, hypoplastic left heart syndrome could be viewed as a neonatal form of cardiomyopathy. We also discuss the role of the endocardium in the development of the ventricular hypoplasia, which may provide a mechanistic basis for how impaired flow to the developing ventricle leads to the anatomical changes seen in the syndrome.

15.
Mech Dev ; 124(9-10): 657-67, 2007.
Artigo em Inglês | MEDLINE | ID: mdl-17716876

RESUMO

ErbB signaling regulates cell adhesion and movements during Xenopus gastrulation, but the downstream pathways involved have not been elucidated. In this study, we show that phosphatidylinositol-3 kinase (PI3K) and Erk mitogen-activated protein kinase (MAPK) mediate ErbB signaling to regulate gastrulation. Both PI3K and MAPK function sequentially in mesoderm specification and movements, and ErbB signaling is important only for the late phase activation of these pathways to control cell behaviors. Activation of either PI3K or Erk MAPK rescues gastrulation defects in ErbB4 morphant embryos, and restores convergent extension in the trunk mesoderm as well as coherent cell migration in the head mesoderm. The two signals preferentially regulate different aspects of cell behaviors, with PI3K more efficient in rescuing cell adhesion and spreading and MAPK more effective in stimulating the formation of filopodia. PI3K and MAPK also weakly activate each other, and together they modulate gastrulation movements. Our results reveal that PI3K and Erk MAPK, which have previously been considered as mesodermal inducing signals, also act downstream of ErbB signaling to participate in regulation of gastrulation morphogenesis.


Assuntos
MAP Quinases Reguladas por Sinal Extracelular/fisiologia , Gástrula/enzimologia , Gastrulação/fisiologia , Proteínas Oncogênicas v-erbB/metabolismo , Fosfatidilinositol 3-Quinases/fisiologia , Transdução de Sinais/fisiologia , Animais , Mesoderma/enzimologia , Proteínas Oncogênicas v-erbB/fisiologia , Xenopus laevis/embriologia
16.
Wiley Interdiscip Rev Dev Biol ; 7(5): e322, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-29722151

RESUMO

Neural crest (NC) cells are a stem-like multipotent population of progenitor cells that are present in vertebrate embryos, traveling to various regions in the developing organism. Known as the "fourth germ layer," these cells originate in the ectoderm between the neural plate (NP), which will become the brain and spinal cord, and nonneural tissues that will become the skin and the sensory organs. NC cells can differentiate into more than 30 different derivatives in response to the appropriate signals including, but not limited to, craniofacial bone and cartilage, sensory nerves and ganglia, pigment cells, and connective tissue. The molecular and cellular mechanisms that control the induction and specification of NC cells include epigenetic control, multiple interactive and redundant transcriptional pathways, secreted signaling molecules, and adhesion molecules. NC cells are important not only because they transform into a wide variety of tissue types, but also because their ability to detach from their epithelial neighbors and migrate throughout developing embryos utilizes mechanisms similar to those used by metastatic cancer cells. In this review, we discuss the mechanisms required for the induction and specification of NC cells in various vertebrate species, focusing on the roles of early morphogenesis, cell adhesion, signaling from adjacent tissues, and the massive transcriptional network that controls the formation of these amazing cells. This article is categorized under: Nervous System Development > Vertebrates: General Principles Gene Expression and Transcriptional Hierarchies > Regulatory Mechanisms Gene Expression and Transcriptional Hierarchies > Gene Networks and Genomics Signaling Pathways > Cell Fate Signaling.

17.
J Mol Cell Biol ; 10(5): 376-387, 2018 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-29040749

RESUMO

The member of Rho family of small GTPases Cdc42 plays important and conserved roles in cell polarity and motility. The Cdc42ep family proteins have been identified to bind to Cdc42, yet how they interact with Cdc42 to regulate cell migration remains to be elucidated. In this study, we focus on Cdc42ep1, which is expressed predominantly in the highly migratory neural crest cells in frog embryos. Through morpholino-mediated knockdown, we show that Cdc42ep1 is required for the migration of cranial neural crest cells. Loss of Cdc42ep1 leads to rounder cell shapes and the formation of membrane blebs, consistent with the observed disruption in actin organization and focal adhesion alignment. As a result, Cdc42ep1 is critical for neural crest cells to apply traction forces at the correct place to migrate efficiently. We further show that Cdc42ep1 is localized to two areas in neural crest cells: in membrane protrusions together with Cdc42 and in perinuclear patches where Cdc42 is absent. Cdc42 directly interacts with Cdc42ep1 (through the CRIB domain) and changes in Cdc42 level shift the distribution of Cdc42ep1 between these two subcellular locations, controlling the formation of membrane protrusions and directionality of migration as a consequence. These results suggest that Cdc42ep1 elaborates Cdc42 activity in neural crest cells to promote their efficient migration.


Assuntos
Proteínas do Citoesqueleto/metabolismo , Proteínas de Membrana/metabolismo , Proteínas Monoméricas de Ligação ao GTP/metabolismo , Crista Neural/citologia , Proteínas de Xenopus/metabolismo , Xenopus laevis/embriologia , Citoesqueleto de Actina/metabolismo , Animais , Animais Geneticamente Modificados , Cartilagem/embriologia , Movimento Celular/fisiologia , Proteínas do Citoesqueleto/genética , Embrião não Mamífero , Matriz Extracelular/metabolismo , Proteínas de Membrana/genética , Proteínas Monoméricas de Ligação ao GTP/genética , Crista Neural/metabolismo , Proteínas de Xenopus/genética , Xenopus laevis/genética
18.
Cardiovasc Res ; 106(1): 67-75, 2015 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-25691536

RESUMO

AIMS: Ets1 is an important transcription factor that is expressed in both the cardiac neural crest (NC) and heart mesoderm of vertebrate embryos. Moreover, Ets1 deletion in humans results in congenital heart abnormalities. To clarify the functional contributions of Ets1 in cardiac NC vs. heart mesoderm, we performed tissue-targeted loss-of-function analysis to compare the relative roles of Ets1 in these two tissues during heart formation using Xenopus embryos as a model system. METHODS AND RESULTS: We confirmed by in situ hybridization analysis that Ets1 is expressed in NC and heart mesoderm during embryogenesis. Using a translation-blocking antisense morpholino to knockdown Ets1 protein selectively in the NC, we observed defects in NC delamination from the neural tube, collective cell migration, as well as segregation of NC streams in the cranial and cardiac regions. Many cardiac NC cells failed to reach their destination in the heart, resulting in defective aortic arch artery formation. A different set of defects was noted when Ets1 knockdown was targeted to heart mesoderm. The formation of the primitive heart tube was dramatically delayed and the endocardial tissue appeared depleted. As a result, the conformation of the heart was severely disrupted. In addition, the outflow tract septum was missing, and trabeculae formation in the ventricle was abolished. CONCLUSION: Our study shows that Ets1 is required in both the cardiac NC and heart mesoderm, albeit for different aspects of heart formation. Our results reinforce the suggestion that proper interaction between these tissues is critical for normal heart development.


Assuntos
Desenvolvimento Embrionário/fisiologia , Coração/embriologia , Mesoderma/embriologia , Crista Neural/embriologia , Proteína Proto-Oncogênica c-ets-1/fisiologia , Fatores de Transcrição/fisiologia , Xenopus laevis/embriologia , Animais , Aorta Torácica/embriologia , Cartilagem/embriologia , Adesão Celular/fisiologia , Movimento Celular/fisiologia , Células Cultivadas , Desenvolvimento Embrionário/genética , Regulação da Expressão Gênica no Desenvolvimento/genética , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Técnicas de Silenciamento de Genes , Mesoderma/citologia , Modelos Animais , Crista Neural/citologia , Proteína Proto-Oncogênica c-ets-1/genética , Crânio/embriologia , Fatores de Transcrição/deficiência , Fatores de Transcrição/genética
19.
Stem Cell Reports ; 5(4): 499-507, 2015 Oct 13.
Artigo em Inglês | MEDLINE | ID: mdl-26441305

RESUMO

Premigratory neural crest cells comprise a transient, embryonic population that arises within the CNS, but subsequently migrates away and differentiates into many derivatives. Previously, premigratory neural crest could not be maintained in a multipotent, adhesive state without spontaneous differentiation. Here, we report conditions that enable maintenance of neuroepithelial "crestospheres" that self-renew and retain multipotency for weeks. Moreover, under differentiation conditions, these cells can form multiple derivatives in vitro and in vivo after transplantation into chick embryos. Similarly, human embryonic stem cells directed to a neural crest fate can be maintained as crestospheres and subsequently differentiated into several derivatives. By devising conditions that maintain the premigratory state in vitro, these results demonstrate that neuroepithelial neural crest precursors are capable of long-term self-renewal. This approach will help uncover mechanisms underlying their developmental potential, differentiation and, together with the induced pluripotent stem cell techniques, the pathology of human neurocristopathies.


Assuntos
Diferenciação Celular , Células-Tronco Multipotentes/citologia , Crista Neural/citologia , Animais , Movimento Celular , Embrião de Galinha , Galinhas , Humanos , Células-Tronco Pluripotentes Induzidas/citologia
20.
PLoS One ; 9(7): e103024, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25051358

RESUMO

Neural crest cells are highly motile, yet a limited number of genes governing neural crest migration have been identified by conventional studies. To test the hypothesis that cell migration genes are likely to be conserved over large evolutionary distances and from diverse tissues, we searched for vertebrate homologs of genes important for migration of various cell types in the invertebrate nematode and examined their expression during vertebrate neural crest cell migration. Our systematic analysis utilized a combination of comparative genomic scanning, functional pathway analysis and gene expression profiling to uncover previously unidentified genes expressed by premigratory, emigrating and/or migrating neural crest cells. The results demonstrate that similar gene sets are expressed in migratory cell types across distant animals and different germ layers. Bioinformatics analysis of these factors revealed relationships between these genes within signaling pathways that may be important during neural crest cell migration.


Assuntos
Movimento Celular/genética , Biologia Computacional/métodos , Genes de Helmintos/genética , Nematoides/genética , Crista Neural/metabolismo , Vertebrados/genética , Animais , Biomarcadores/metabolismo , Caenorhabditis elegans/citologia , Caenorhabditis elegans/embriologia , Caenorhabditis elegans/genética , Embrião de Galinha , Embrião não Mamífero/embriologia , Embrião não Mamífero/metabolismo , Evolução Molecular , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica no Desenvolvimento , Genômica/métodos , Hibridização In Situ , Nematoides/citologia , Nematoides/embriologia , Crista Neural/citologia , Crista Neural/embriologia , Transdução de Sinais/genética , Vertebrados/embriologia
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