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Andrew J Copp - One of the best experts on this subject based on the ideXlab platform.
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biomechanical coupling facilitates spinal Neural tube closure in mouse embryos
Proceedings of the National Academy of Sciences of the United States of America, 2017Co-Authors: Gabriel L Galea, Nicholas D E Greene, Gauden Galea, Matteo A Mole, Ana Rolo, Dawn Savery, Dale Moulding, Lucy H Culshaw, Evanthia Nikolopoulou, Andrew J CoppAbstract:Neural tube (NT) formation in the spinal region of the mammalian embryo involves a wave of “zippering” that passes down the elongating spinal axis, uniting the Neural Fold tips in the dorsal midline. Failure of this closure process leads to open spina bifida, a common cause of severe neurologic disability in humans. Here, we combined a tissue-level strain-mapping workflow with laser ablation of live-imaged mouse embryos to investigate the biomechanics of mammalian spinal closure. Ablation of the zippering point at the embryonic dorsal midline causes far-reaching, rapid separation of the elevating Neural Folds. Strain analysis revealed tissue expansion around the zippering point after ablation, but predominant tissue constriction in the caudal and ventral Neural plate zone. This zone is biomechanically coupled to the zippering point by a supracellular F-actin network, which includes an actin cable running along the Neural Fold tips. Pharmacologic inhibition of F-actin or laser ablation of the cable causes Neural Fold separation. At the most advanced somite stages, when completion of spinal closure is imminent, the cable forms a continuous ring around the neuropore, and simultaneously, a new caudal-to-rostral zippering point arises. Laser ablation of this new closure initiation point causes Neural Fold separation, demonstrating its biomechanical activity. Failure of spinal closure in pre-spina bifida Zic2Ku mutant embryos is associated with altered tissue biomechanics, as indicated by greater neuropore widening after ablation. Thus, this study identifies biomechanical coupling of the entire region of active spinal neurulation in the mouse embryo as a prerequisite for successful NT closure.
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Rho GTPases in mammalian spinal Neural tube closure.
Small GTPases, 2016Co-Authors: Ana Rolo, Nicholas D E Greene, Sarah Escuin, Andrew J CoppAbstract:Neural tube closure is an important morphogenetic event that involves dramatic reshaping of both Neural and non-Neural tissues. Rho GTPases are key cytoskeletal regulators involved in cell motility and in several developmental processes, and are thus expected to play pivotal roles in neurulation. Here, we discuss 2 recent studies that shed light on the roles of distinct Rho GTPases in different tissues during neurulation. RhoA plays an essential role in regulating actomyosin dynamics in the Neural epithelium of the elevating Neural Folds, while Rac1 is required for the formation of cell protrusions in the non-Neural surface ectoderm during Neural Fold fusion.
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ephrina epha receptor interactions in mouse spinal neurulation implications for Neural Fold fusion
The International Journal of Developmental Biology, 2009Co-Authors: Noraishah M Abdulaziz, Mark Turmaine, Nicholas D E Greene, Andrew J CoppAbstract:The molecular basis of Neural Fold adhesion and fusion is a poorly understood aspect of neurulation. Cell surface glycosyl phosphatidylinositol (GPI)-anchored proteins have been implicated in Neural Fold adhesion, with ephrinAs particularly attractive candidates in view of the cranial Neural tube defects observed in mice lacking ephrinA5 or the EphA7 receptor. Here, we demonstrate that ephrinsA1, A3 and A4, as well as several EphA receptors, are expressed in the closing mouse spinal Neural tube. Most ephrinAs and EphA receptors were found to be expressed in multiple tissues in the caudal region, whereas EphA2 receptor was expressed specifically at the apices of the Neural Folds just prior to onset of Neural tube fusion. Using mouse whole embryo culture, we found that cleavage of GPI-anchored molecules from the embryonic cell surface resulted in delay of spinal Neural tube closure. Injection of EphA1 and EphA3 fusion proteins intraamniotically into cultured embryos was used to specifically disrupt ephrinA-EphA receptor interactions, and led to inhibition of spinal Neural tube closure, without adverse effects on growth or developmental progression. These treatments did not disturb Neural plate bending or Neural Fold elevation, both of which are critical for spinal Neural tube closure. Our findings demonstrate that ephrinA-EphA receptor interactions are required for closure of the mouse spinal Neural tube, and support the hypothesis that ephrinA-EphA receptor interactions may participate in the molecular recognition events that culminate in adhesion and fusion of the tips of the Neural Folds during spinal neurulation.
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Failure of Neural tube closure in the loop-tail (Lp) mutant mouse: analysis of the embryonic mechanism
Brain research. Developmental brain research, 1997Co-Authors: Dianne Gerrelli, Andrew J CoppAbstract:Abstract Loop-tail ( Lp ) is unique among mouse mutants in failing to initiate Neural tube closure at the cervical/hindbrain boundary (so-called `Closure 1'), at the 5–7 somite stage. Lp/Lp embryos go on to develop a malformation that closely resembles cranio-rachischisis, the most severe Neural tube defect found in humans. We investigated several possible embryological mechanisms that may underlie this failure of Neural tube closure in Lp . The genotypes of Lp/Lp , Lp/+ and +/+ embryos from mixed litters were identified using the polymerase chain reaction to amplify a polymorphic microsatellite sequence that is very closely linked to Lp . At post-neurulation stages of development, Lp/Lp embryos have a shortened body axis, which could suggest a defect of axial elongation as the primary anomaly in Lp . However, we found that axial elongation is normal in Lp homozygotes prior to the stage of defective Closure 1, indicating that the shortened body axis of later embryos is a secondary effect of the neurulation anomaly, or an independent effect of the Lp mutation. Some workers have reported cell proliferation rates to be abnormal in later stage Lp/Lp embryos. We observed variations in [ 3 H]thymidine labelling index, and mitotic index, between embryonic tissues, and between embryos at different somite stages. However, Lp/Lp , Lp/+ and +/+ embryos had closely similar cell proliferation parameters, arguing against a mechanism based on faulty embryonic growth. Thirdly, we tested the hypothesis that the defect in loop-tail results from an inability of the Neural Folds to become apposed, specifically at the site of Closure 1. By tying a silk suture around the embryonic axis, at the future site of Closure 1, we were able to effect convergence of the Neural Folds at this site. Neural Fold closure failed to progress along the body axis in sutured Lp/Lp embryos, however, in contrast to operated Lp/+ and +/+ embryos which exhibited normal progression of Neural tube closure. The embryonic defect in loop-tail appears, therefore, to involve either a general inability of the spinal Neural Folds to become apposed along the spinal region, or a defect in the process of Neural Fold fusion.
Roberto Mayor - One of the best experts on this subject based on the ideXlab platform.
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the posteriorizing gene gbx2 is a direct target of wnt signalling and the earliest factor in Neural crest induction
Development, 2009Co-Authors: Bo Li, Sei Kuriyama, Mauricio Moreno, Roberto MayorAbstract:Wnt signalling is required for Neural crest (NC) induction; however, the direct targets of the Wnt pathway during NC induction remain unknown. We show here that the homeobox gene Gbx2 is essential in this process and is directly activated by Wnt/beta-catenin signalling. By ChIP and transgenesis analysis we show that the Gbx2 regulatory elements that drive expression in the NC respond directly to Wnt/beta-catenin signalling. Gbx2 has previously been implicated in posteriorization of the Neural plate. Here we unveil a new role for this gene in Neural Fold patterning. Loss-of-function experiments using antisense morpholinos against Gbx2 inhibit NC and expand the preplacodal domain, whereas Gbx2 overexpression leads to transformation of the preplacodal domain into NC cells. We show that the NC specifier activity of Gbx2 is dependent on the interaction with Zic1 and the inhibition of preplacodal genes such as Six1. In addition, we demonstrate that Gbx2 is upstream of the Neural Fold specifiers Pax3 and Msx1. Our results place Gbx2 as the earliest factor in the NC genetic cascade being directly regulated by the inductive molecules, and support the notion that posteriorization of the Neural Folds is an essential step in NC specification. We propose a new genetic cascade that operates in the distinction between anterior placodal and NC territories.
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Neural crests are actively precluded from the anterior Neural Fold by a novel inhibitory mechanism dependent on Dickkopf1 secreted by the prechordal mesoderm.
Developmental biology, 2007Co-Authors: Carlos Carmona-fontaine, Gustavo Acuña, Kristina Ellwanger, Christof Niehrs, Roberto MayorAbstract:It is known the interactions between the Neural plate and epidermis generate Neural crest (NC), but it is unknown why the NC develops only at the lateral border of the Neural plate and not in the anterior Fold. Using grafting experiments we show that there is a previously unidentified mechanism that precludes NC from the anterior region. We identify prechordal mesoderm as the tissue that inhibits NC in the anterior territory and show that the Wnt/β-catenin antagonist Dkk1, secreted by this tissue, is sufficient to mimic this NC inhibition. We show that Dkk1 is required for preventing the formation of NC in the anterior Neural Folds as loss-of-function experiments using a Dkk1 blocking antibody in Xenopus as well as the analysis of Dkk1-null mouse embryos transform the anterior Neural Fold into NC. This can be mimicked by Wnt/β-catenin signaling activation without affecting the anterior posterior patterning of the Neural plate, or placodal specification. Finally, we show that the NC cells induced at the anterior Neural Fold are able to migrate and differentiate as normal NC. These results demonstrate that anterior regions of the embryo lack NC because of a mechanism, conserved from fish to mammals, that suppresses Wnt/β-catenin signaling via Dkk1.
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Extracellular signals, cell interactions and transcription factors involved in the induction of the Neural crest cells
Biological research, 2002Co-Authors: Manuel J. Aybar, Alvaro Glavic, Roberto MayorAbstract:ABSTRACT The Neural crest is induced at the border between the Neural plate and the epidermis. A complex set of signals isrequired for the specification of the crest cells between the epidermis and the Neural plate. Here we discussevidence supporting a model for Neural crest induction in which different signals contribute in a sequential order.First, a gradient of bone morphogenic proteins (BMPs) is established in the ectoderm that results in segreggationinto Neural plate, Neural Folds and epidermis at increasing levels of BMP activity. Thus, the Neural Folds areinduced at a precise threshold concentration of BMP, but this Neural Fold has an anterior character. In a secondstep, these anterior Neural Folds are transformed into prospective Neural crest by posteriorizing signals due tofibroblast growth factor, Wnts and retinoic acid. Finally, the induced cells interact to complete Neural crestinduction by a process that requires Notch/Delta signaling. Once Neural crest formation has been induced by thiscombination of extracellular and intracellular signals, a cascade of transcription factors is activated in these cellsthat culminates in the ultimate steps of Neural crest differentiation.Key terms: Neural crest, induction, BMPs, Wnts, FGF, retinoic acid, Notch, delta
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Induction and development of Neural crest in Xenopus laevis.
Cell and tissue research, 2001Co-Authors: Roberto Mayor, Manuel J. AybarAbstract:Neural crest cells are a migratory embryonic cell population that form at the border between the Neural plate and the future epidermis. This border, the Neural plate border, corresponds to the Neural Fold. The Neural Fold surrounds the entire Neural plate, but only the lateral and posterior portions of the Fold give rise to Neural crest cells, while the anterior Neural Fold differentiates as forebrain. This review focuses on Neural crest development in Xenopus laevis embryos, and analyzes aspects of the induction of the Neural crest in Xenopus, summarizing available information relating to the expression of several genes in the Neural crest. Two models for Neural crest induction are discussed. In the first model, the Neural crest is induced by the interaction between the Neural plate and the epidermis. In the second, the specification of the Neural plate border arises as a consequence of a gradient of BMP activity. The role of posteriorizing signals on Neural crest specification is also discussed. Finally, we propose that the specification and differentiation of the Neural crest is controlled by a cascade of transcription factors, encoded and expressed from a hierarchy of genes. A set of extracellular signals establishes the positional information in the ectoderm, which activates Prepattern genes (Gli, Xiro, Zic, Dlx, etc.) across extended and overlapping domains. A local combination of these genes at the Neural plate border activates the cascade of Neural crest specification, while different sets of genes are activated at both sides of the Neural Folds (in the epidermis and the Neural plate). The genes activated in regions adjacent to the Neural plate border have an inhibitory effect on the Neural crest transcription program.
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Expression of Xenopus snail in mesoderm and prospective Neural Fold ectoderm.
Developmental dynamics : an official publication of the American Association of Anatomists, 1993Co-Authors: Linda J. Essex, Roberto Mayor, Michael G. SargentAbstract:Expression of the Xsna gene during Xenopus laevis embryogenesis has been analysed by in situ hybridisation. Like its homo- lope snail in Drosophila, Xsna is expressed zy- gotically in all early mesoderm. Expression starts during stage 9 in the dorsal marginal zone and spreads to the ventral side by stage 10. During gastrulation, each cell begins to express as it in- volutes so that cells newly expressing Xsna are added to the forming mesoderm mantle in an an- terior-to-posterior progression. Xsna expression is then down-regulated in a tissue-specific fashion that reveals the subdivision of the mesoderm be- fore its derivatives are overtly differentiated; e.g., the appearance of the notochord, myotomes, and pronephroi are preceded by the disappearance of Xsna mRNA, while undifferentiated mesoderm re- mains labelled, even into tadpole stages. Xsna is expressed in the suprablastoporal endoderm dur- ing gastrulation and in its derivatives, the pre- chordal and sub-notochordal endoderm, during neurulation. Relationships between Xbra, Xtwi, and Xsna expression are examined. Xsna is also expressed in the prospective Neural Fold ectoderm from stage 11 in a low arc above the dorsal marginal zone, precisely identifying a dis- tinct band of cells that surrounds the prospective Neural plate that we designate the Neural plate border. The anterior transverse Neural Fold, which becomes forebrain, ceases Xsna expression dur- ing neurulation. In the longitudinal Neural Folds, the deep and superficial ectoderm compartments labelled by Xsna expression are the prospective Neural crest and prospective roof of the Neural tube, respectively. Xsna expression persists in the Neural crest during migration and in some deriv- atives at least until metamorphosis but ceases in the roof of the Neural tube soon after neuru- lation. 0 1993 Wiley-Liss, Inc.
Raymond Habas - One of the best experts on this subject based on the ideXlab platform.
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MIM regulates vertebrate Neural tube closure
Development, 2011Co-Authors: Yuko Komiya, Courtney Mezzacappa, Deepak K. Khadka, Loren W. Runnels, Raymond HabasAbstract:Neural tube closure is a critical morphogenetic event that is regulated by dynamic changes in cell shape and behavior. Although previous studies have uncovered a central role for the non-canonical Wnt signaling pathway in Neural tube closure, the underlying mechanism remains poorly resolved. Here, we show that the missing in metastasis (MIM; Mtss1) protein, previously identified as a Hedgehog response gene and actin and membrane remodeling protein, specifically binds to Daam1 and couples non-canonical Wnt signaling to Neural tube closure. MIM binds to a conserved domain within Daam1, and this interaction is positively regulated by Wnt stimulation. Spatial expression of MIM is enriched in the anterior Neural plate and Neural Folds, and depletion of MIM specifically inhibits anterior Neural Fold closure without affecting convergent extension movements or mesoderm cell fate specification. Particularly, we find that MIM is required for Neural Fold elevation and apical constriction along with cell polarization and elongation in both the superficial and deep layers of the anterior Neural plate. The function of MIM during Neural tube closure requires both its membrane-remodeling domain and its actin-binding domain. Finally, we show that the effect of MIM on Neural tube closure is not due to modulation of Hedgehog signaling in the Xenopus embryo. Together, our studies define a morphogenetic pathway involving Daam1 and MIM that transduces non-canonical Wnt signaling for the cytoskeletal changes and membrane dynamics required for vertebrate Neural tube closure.
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profilin is an effector for daam1 in non canonical wnt signaling and is required for vertebrate gastrulation
Development, 2006Co-Authors: Akira Sato, Deepak K. Khadka, Loren W. Runnels, Wei Liu, Ritu Bharti, Igor B Dawid, Raymond HabasAbstract:Non-canonical Wnt signaling plays important roles during vertebrate embryogenesis and is required for cell motility during gastrulation. However, the molecular mechanisms of how Wnt signaling regulates modification of the actin cytoskeleton remain incompletely understood. We had previously identified the Formin homology protein Daam1 as an important link between Dishevelled and the Rho GTPase for cytoskeletal modulation. Here, we report that Profilin1 is an effector downstream of Daam1 required for cytoskeletal changes. Profilin1 interacted with the FH1 domain of Daam1 and was localized with Daam1 to actin stress fibers in response to Wnt signaling in mammalian cells. In addition, depletion of Profilin1 inhibited stress fiber formation induced by non-canonical Wnt signaling. Inhibition or depletion of Profilin1 in vivo specifically inhibited blastopore closure in Xenopus but did not affect convergent extension movements, tissue separation or Neural Fold closure. Our studies define a molecular pathway downstream of Daam1 that controls Wnt-mediated cytoskeletal reorganization for a specific morphogenetic process during vertebrate gastrulation.
Nicholas D E Greene - One of the best experts on this subject based on the ideXlab platform.
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biomechanical coupling facilitates spinal Neural tube closure in mouse embryos
Proceedings of the National Academy of Sciences of the United States of America, 2017Co-Authors: Gabriel L Galea, Nicholas D E Greene, Gauden Galea, Matteo A Mole, Ana Rolo, Dawn Savery, Dale Moulding, Lucy H Culshaw, Evanthia Nikolopoulou, Andrew J CoppAbstract:Neural tube (NT) formation in the spinal region of the mammalian embryo involves a wave of “zippering” that passes down the elongating spinal axis, uniting the Neural Fold tips in the dorsal midline. Failure of this closure process leads to open spina bifida, a common cause of severe neurologic disability in humans. Here, we combined a tissue-level strain-mapping workflow with laser ablation of live-imaged mouse embryos to investigate the biomechanics of mammalian spinal closure. Ablation of the zippering point at the embryonic dorsal midline causes far-reaching, rapid separation of the elevating Neural Folds. Strain analysis revealed tissue expansion around the zippering point after ablation, but predominant tissue constriction in the caudal and ventral Neural plate zone. This zone is biomechanically coupled to the zippering point by a supracellular F-actin network, which includes an actin cable running along the Neural Fold tips. Pharmacologic inhibition of F-actin or laser ablation of the cable causes Neural Fold separation. At the most advanced somite stages, when completion of spinal closure is imminent, the cable forms a continuous ring around the neuropore, and simultaneously, a new caudal-to-rostral zippering point arises. Laser ablation of this new closure initiation point causes Neural Fold separation, demonstrating its biomechanical activity. Failure of spinal closure in pre-spina bifida Zic2Ku mutant embryos is associated with altered tissue biomechanics, as indicated by greater neuropore widening after ablation. Thus, this study identifies biomechanical coupling of the entire region of active spinal neurulation in the mouse embryo as a prerequisite for successful NT closure.
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Rho GTPases in mammalian spinal Neural tube closure.
Small GTPases, 2016Co-Authors: Ana Rolo, Nicholas D E Greene, Sarah Escuin, Andrew J CoppAbstract:Neural tube closure is an important morphogenetic event that involves dramatic reshaping of both Neural and non-Neural tissues. Rho GTPases are key cytoskeletal regulators involved in cell motility and in several developmental processes, and are thus expected to play pivotal roles in neurulation. Here, we discuss 2 recent studies that shed light on the roles of distinct Rho GTPases in different tissues during neurulation. RhoA plays an essential role in regulating actomyosin dynamics in the Neural epithelium of the elevating Neural Folds, while Rac1 is required for the formation of cell protrusions in the non-Neural surface ectoderm during Neural Fold fusion.
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ephrina epha receptor interactions in mouse spinal neurulation implications for Neural Fold fusion
The International Journal of Developmental Biology, 2009Co-Authors: Noraishah M Abdulaziz, Mark Turmaine, Nicholas D E Greene, Andrew J CoppAbstract:The molecular basis of Neural Fold adhesion and fusion is a poorly understood aspect of neurulation. Cell surface glycosyl phosphatidylinositol (GPI)-anchored proteins have been implicated in Neural Fold adhesion, with ephrinAs particularly attractive candidates in view of the cranial Neural tube defects observed in mice lacking ephrinA5 or the EphA7 receptor. Here, we demonstrate that ephrinsA1, A3 and A4, as well as several EphA receptors, are expressed in the closing mouse spinal Neural tube. Most ephrinAs and EphA receptors were found to be expressed in multiple tissues in the caudal region, whereas EphA2 receptor was expressed specifically at the apices of the Neural Folds just prior to onset of Neural tube fusion. Using mouse whole embryo culture, we found that cleavage of GPI-anchored molecules from the embryonic cell surface resulted in delay of spinal Neural tube closure. Injection of EphA1 and EphA3 fusion proteins intraamniotically into cultured embryos was used to specifically disrupt ephrinA-EphA receptor interactions, and led to inhibition of spinal Neural tube closure, without adverse effects on growth or developmental progression. These treatments did not disturb Neural plate bending or Neural Fold elevation, both of which are critical for spinal Neural tube closure. Our findings demonstrate that ephrinA-EphA receptor interactions are required for closure of the mouse spinal Neural tube, and support the hypothesis that ephrinA-EphA receptor interactions may participate in the molecular recognition events that culminate in adhesion and fusion of the tips of the Neural Folds during spinal neurulation.
Marianne Bronner-fraser - One of the best experts on this subject based on the ideXlab platform.
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Competence, specification and commitment to an olfactory placode fate.
Development (Cambridge England), 2008Co-Authors: Sujata Bhattacharyya, Marianne Bronner-fraserAbstract:The nasal placode shares a common origin with other sensory placodes within a pre-placodal domain at the cranial Neural plate border. However, little is known about early events in nasal placode development as it segregates from prospective lens, Neural tube and epidermis. Here, Dlx3, Dlx5, Pax6 and the pan-neuronal marker Hu serve as molecular labels to follow the maturation of olfactory precursors over time. When competence to form olfactory placode was tested by grafting ectoderm from different axial levels to the anterior Neural Fold, we found that competence is initially broad for head, but not trunk, ectoderm and declines rapidly with time. Isolated olfactory precursors are specified by HH10, concomitant with their complete segregation from other placodal, epidermal and Neural progenitors. Heterotopic transplantation of olfactory progenitors reveals they are capable of autonomous differentiation only 12 hours later, shortly before overt placode invagination at HH14. Taken together, these results show that olfactory placode development is a step-wise process whereby signals from adjacent tissues specify competent ectoderm at or before HH10, followed by gradual commitment just prior to morphological differentiation.
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Timing and competence of Neural crest formation.
Developmental neuroscience, 2000Co-Authors: Martin L. Basch, Mark A. J. Selleck, Marianne Bronner-fraserAbstract:Neural crest cells can be induced by an interaction between Neural plate and ectoderm. To clarify the timing and nature of these inductive interactions, we have examined the time of competence of the Neural plate to become Neural crest as well as the time of Neural Fold specification. The Neural plate is competent to respond to inductive interactions with the nonNeural ectoderm for a limited period, rapidly losing its responsive ability after stage 10. In contrast, nonNeural ectoderm from numerous stages retains the ability to induce Neural crest cells from competent Neural plate. When Neural Folds are explanted to test their ability to produce Neural crest without further tissue interactions, we find that Folds derived from all rostrocaudal levels of the open Neural plate are already specified to express the Neural crest marker Slug. However, additional signals may be required for maintenance of Slug expression, since the transcript is later down-regulated in vitro in the absence of tissue interactions. Taken together, these results suggest that there are multiple stages of Neural crest induction. The earliest induction must have occurred by the end of gastrulation, since the newly formed Neural Fold population is already specified to form Neural crest. However, isolated Neural Folds eventually down-regulate Slug, suggesting a second phase that maintains Neural crest formation. Thus, induction of the Neural crest may involve multiple and sustained tissue interactions.
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Regulative response of the cranial Neural tube after Neural Fold ablation: spatiotemporal nature of Neural crest regeneration and up-regulation of Slug
Development (Cambridge England), 1995Co-Authors: John Sechrist, M. Angela Nieto, Roham T. Zamanian, Marianne Bronner-fraserAbstract:After unilateral ablation of the avian cranial Neural Folds, the remaining neuroepithelial cells are able to replace the missing Neural crest population (Scherson et al., 1993). Here, we characterize the cellular and molecular nature of this regulative response by defining: (1) the time and location of Neural crest cell production by the neuroepithelium; (2) rostrocaudal axial differences in the regulative response; and (3) the onset of expression of Slug, a transcription factor present in premigratory and migrating Neural crest cells. Using DiI and HNK-1 antibody labeling techniques, we find that Neural crest regeneration occurs only after apposition of the remaining neuroepithelium with the epidermis, suggesting that the developmental mechanism underlying regeneration of the Neural crest may recapitulate initial generation of the Neural crest. The regulative response occurs maximally at the 3–5 somite stage, and slowly declines thereafter. Surprisingly, there are profound regional differences in the regenerative ability. Whereas a robust regulation occurs in the caudal midbrain/hindbrain, the caudal forebrain/rostral midbrain regenerates Neural crest to a much lesser extent. After Neural Fold removal in the hindbrain, regenerated Neural crest cells migrate in a segmental pattern analogous to that seen in unablated embryos; a decrease in regulative response appears to occur with increasing depth of the ablation. Up-regulation of Slug appears to be an early response after ablation, with Slug transcripts detectable proximal to the ablated region 5–8 hours after surgery and prior to emergence of Neural crest cells. Both bilateral and unilateral ablations yield substantial numbers of Neural crest cells, though the former recover less rapidly and have greater deficits in Neural crest-derived structures than the latter. These experiments demonstrate that the regulative ability of the cranial neuroepithelium to form Neural crest depends on the time, location and extent of Neural Fold ablation.