RESUMO
Maldevelopment of the pharyngeal endoderm, an embryonic tissue critical for patterning of the pharyngeal region and ensuing organogenesis, ultimately contributes to several classes of human developmental syndromes and disorders. Such syndromes are characterized by a spectrum of phenotypes that currently cannot be fully explained by known mutations or genetic variants due to gaps in characterization of critical drivers of normal and dysfunctional development. Despite the disease-relevance of pharyngeal endoderm, we still lack a comprehensive and integrative view of the molecular basis and gene regulatory networks driving pharyngeal endoderm development. To close this gap, we apply transcriptomic and chromatin accessibility single-cell sequencing technologies to generate a multi-omic developmental resource spanning pharyngeal endoderm patterning to the emergence of organ-specific epithelia in the developing mouse embryo. We identify cell-type specific gene regulation, distill GRN models that define developing organ domains, and characterize the role of an immunodeficiency-associated forkhead box transcription factor.
Assuntos
Cromatina/genética , Regulação da Expressão Gênica no Desenvolvimento , Faringe/embriologia , Transcriptoma , Animais , Cromatina/metabolismo , Endoderma/embriologia , Endoderma/metabolismo , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Organogênese , Faringe/metabolismo , Análise de Célula Única , Timócitos/citologia , Timócitos/metabolismoRESUMO
During animal ontogenesis, body axis patterning is finely regulated by complex interactions among several signaling pathways. Nitric oxide (NO) and retinoic acid (RA) are potent morphogens that play a pivotal role in vertebrate development. Their involvement in axial patterning of the head and pharynx shows conserved features in the chordate phylum. Indeed, in the cephalochordate amphioxus, NO and RA are crucial for the correct development of pharyngeal structures. Here, we demonstrate the functional cooperation between NO and RA that occurs during amphioxus embryogenesis. During neurulation, NO modulates RA production through the transcriptional regulation of Aldh1a.2 that irreversibly converts retinaldehyde into RA. On the other hand, RA directly or indirectly regulates the transcription of Nos genes. This reciprocal regulation of NO and RA pathways is essential for the normal pharyngeal development in amphioxus and it could be conserved in vertebrates.
Assuntos
Padronização Corporal , Anfioxos/embriologia , Óxido Nítrico/metabolismo , Transdução de Sinais , Tretinoína/metabolismo , Animais , Embrião não Mamífero/embriologia , Desenvolvimento Embrionário , Faringe/embriologiaRESUMO
The internalization of the central nervous system, termed neurulation in vertebrates, is a critical step in embryogenesis. Open questions remain regarding how force propels coordinated tissue movement during the process, and little is known as to how internalization happens in invertebrates. We show that in C. elegans morphogenesis, apical constriction in the retracting pharynx drives involution of the adjacent neuroectoderm. HMR-1/cadherin mediates this process via inter-tissue attachment, as well as cohesion within the neuroectoderm. Our results demonstrate that HMR-1 is capable of mediating embryo-wide reorganization driven by a centrally located force generator, and indicate a non-canonical use of cadherin on the basal side of an epithelium that may apply to vertebrate neurulation. Additionally, we highlight shared morphology and gene expression in tissues driving involution, which suggests that neuroectoderm involution in C. elegans is potentially homologous with vertebrate neurulation and thus may help elucidate the evolutionary origin of the brain.
Assuntos
Caderinas/genética , Proteínas de Caenorhabditis elegans/genética , Caenorhabditis elegans/embriologia , Neurulação , Animais , Caderinas/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Embrião não Mamífero/embriologia , Morfogênese , Placa Neural/embriologia , Faringe/embriologiaRESUMO
The vertebrate body plan is thought to be derived during the early Cambrian from a worm-like chordate ancestor. While all three germ layers were clearly involved in this innovation, the role of the endoderm remains elusive. According to the hourglass model, the optimal window for investigating the evolution of vertebrate endoderm-derived structures during cephalochordate development is from the Spemann's organizer stage to the opening of the mouth (Stages 1-7, described herein). Regulatory gene expression, examined during these stages, illustrate that the cephalochordate endoderm is patterned into 12 organ primordia. Early vertebrates inherited at least a portion of 6 of these primordia, while the remainder were lost. Of those that were preserved, we demonstrate that the vertebrate symmetric mouth was built on a vestige of the anterior pre-oral pit, that the pre-existing pharyngeal pouch in this chordate ancestor laid the foundation for the new neural crest cell (NCC)-derived vertebrate-type pharyngeal arches, that the thyroid evolved from the posterior endostyle primordim, that the pancreas was derived from the Pdx1-expressing diverticulum primordium, and the small and large intestines originated with the Cdx1-expressing hindgut rudiments. This investigation uncovers the evolutionary foundations of vertebrate endoderm-derived structures, and demonstrates that the number of organ primordia were reduced during evolution.
Assuntos
Padronização Corporal/genética , Endoderma/embriologia , Modelos Biológicos , Vertebrados/embriologia , Animais , Proteínas de Homeodomínio/genética , Pâncreas/embriologia , Faringe/embriologiaRESUMO
BACKGROUND: Vertebrate cranial neural crest cells (CNCCs) are multipotent, proximal to the source CNCC form the cranial ganglia. Distally, in the pharyngeal arches, they give rise to the craniofacial skeleton and connective tissues. Fate choices are made as CNCC pattern into distinct destination compartments. In spite of this importance, the mechanism patterning CNCC is poorly defined. RESULTS: Here, we report that a novel ß-catenin-dependent regulation of N-Cadherin levels may drive CNCC patterning. In mouse embryos, at the first pharyngeal arch axial level, membrane ß-catenin levels correlate with the extent of N-cadherin-mediated adhesion and thus suggest the presence of collective and dispersed states of CNCC. Using in vitro human neural crest model and chemical modulators of ß-catenin levels, we show a requirement for down-modulating ß-catenin for regulating N-cadherin levels and cell-cell adhesion. Similarly, in ß-catenin gain-of-function mutant mouse embryos, CNCC fail to lower N-cadherin levels. This indicates a failure to reduce cell-cell adhesion, which may underlie the failure of mutant CNCC to populate first pharyngeal arch. CONCLUSION: We suggest that ß-catenin-mediated regulation of CNCC adhesion, a previously underappreciated mechanism, underlies the patterning of CNCC into fate-specific compartments.
Assuntos
Padronização Corporal , Crista Neural/embriologia , Faringe/embriologia , Crânio/embriologia , beta Catenina/metabolismo , Animais , Camundongos , Camundongos Transgênicos , Crista Neural/citologia , Faringe/citologia , Crânio/citologia , beta Catenina/genéticaRESUMO
Cardiopharyngeal mesoderm (CPM) gives rise to muscles of the head and heart. Using genetic lineage analysis in mice, we show that CPM develops into a broad range of pharyngeal structures and cell types encompassing musculoskeletal and connective tissues. We demonstrate that CPM contributes to medial pharyngeal skeletal and connective tissues associated with both branchiomeric and somite-derived neck muscles. CPM and neural crest cells (NCC) make complementary mediolateral contributions to pharyngeal structures, in a distribution established in the early embryo. We further show that biallelic expression of the CPM regulatory gene Tbx1, haploinsufficient in 22q11.2 deletion syndrome patients, is required for the correct patterning of muscles with CPM-derived connective tissue. Our results suggest that CPM plays a patterning role during muscle development, similar to that of NCC during craniofacial myogenesis. The broad lineage contributions of CPM to pharyngeal structures provide new insights into congenital disorders and evolution of the mammalian pharynx.
Assuntos
Tecido Conjuntivo/embriologia , Desenvolvimento Muscular/genética , Faringe/embriologia , Somitos/fisiologia , Animais , Padronização Corporal/genética , Linhagem da Célula/genética , Regulação da Expressão Gênica no Desenvolvimento , Camundongos , Camundongos Transgênicos , Crista Neural/metabolismo , Faringe/citologia , Somitos/citologia , Proteínas com Domínio T/metabolismoRESUMO
Although cytokinesis has been intensely studied, the way it is executed during development is not well understood, despite a long-standing appreciation that various aspects of cytokinesis vary across cell and tissue types. To address this, we investigated cytokinesis during the invariant Caenorhabditis elegans embryonic divisions and found several parameters that are altered at different stages in a reproducible manner. During early divisions, furrow ingression asymmetry and midbody inheritance is consistent, suggesting specific regulation of these events. During morphogenesis, we found several unexpected alterations to cytokinesis, including apical midbody migration in polarizing epithelial cells of the gut, pharynx and sensory neurons. Aurora B kinase, which is essential for several aspects of cytokinesis, remains apically localized in each of these tissues after internalization of midbody ring components. Aurora B inactivation disrupts cytokinesis and causes defects in apical structures, even if inactivated post-mitotically. Therefore, we demonstrate that cytokinesis is implemented in a specialized way during epithelial polarization and that Aurora B has a role in the formation of the apical surface.
Assuntos
Aurora Quinase B/fisiologia , Proteínas de Caenorhabditis elegans/fisiologia , Caenorhabditis elegans/embriologia , Caenorhabditis elegans/enzimologia , Citocinese , Morfogênese , Animais , Caenorhabditis elegans/citologia , Polaridade Celular , Citocinese/fisiologia , Dendritos/fisiologia , Embrião não Mamífero/citologia , Células Epiteliais/fisiologia , Intestinos/embriologia , Neurônios/citologia , Faringe/embriologia , Propriedades de SuperfícieRESUMO
The pharyngeal arches are a prominent and significant feature of vertebrate embryos. These are visible as a series of bulges on the lateral surface of the embryonic head. In humans, and other amniotes, there are five pharyngeal arches numbered 1, 2, 3, 4 and 6; note the missing '5'. This is the standard scheme for the numbering of these structures, and it is a feature of modern anatomy textbooks. In this article, we discuss the rationale behind this odd numbering, and consider its origins. One reason given is that there is a transient 5th arch that is never fully realized, while another is that this numbering reflects considerations from comparative anatomy. We show here, however, that neither of these reasons has substance. There is no evidence from embryology for a '5th' arch, and the comparative argument does not hold as it does not apply across the vertebrates. We conclude that there is no justification for this strange numbering. We suggest that the pharyngeal arches should simply be numbered 1, 2, 3, 4 and 5 as this would be in keeping with the embryology and with the general numbering of the pharyngeal arches across the vertebrates.
Assuntos
Cabeça/embriologia , Animais , Evolução Biológica , Região Branquial/anatomia & histologia , Crista Neural/anatomia & histologia , Faringe/embriologia , Vertebrados/embriologiaRESUMO
Developmental defects affecting the heart and aortic arch arteries are a significant phenotype observed in individuals with 22q11 deletion syndrome and are caused by a microdeletion on chromosome 22q11. TBX1, one of the deleted genes, is expressed throughout the pharyngeal arches and is considered a key gene, when mutated, for the arch artery defects. Pax9 is expressed in the pharyngeal endoderm and is downregulated in Tbx1 mutant mice. We show here that Pax9-deficient mice are born with complex cardiovascular malformations that affect the outflow tract and aortic arch arteries with failure of the 3rd and 4th pharyngeal arch arteries to form correctly. Transcriptome analysis indicated that Pax9 and Tbx1 may function together, and mice double heterozygous for Tbx1/Pax9 presented with a significantly increased incidence of interrupted aortic arch when compared with Tbx1 heterozygous mice. Using a novel Pax9Cre allele, we demonstrated that the site of this Tbx1-Pax9 genetic interaction is the pharyngeal endoderm, therefore revealing that a Tbx1-Pax9-controlled signalling mechanism emanating from the pharyngeal endoderm is required for crucial tissue interactions during normal morphogenesis of the pharyngeal arch artery system.
Assuntos
Artérias/embriologia , Região Branquial/irrigação sanguínea , Sistema Cardiovascular/embriologia , Endoderma/embriologia , Morfogênese , Fator de Transcrição PAX9/metabolismo , Faringe/embriologia , Proteínas com Domínio T/metabolismo , Animais , Sistema Cardiovascular/metabolismo , Diferenciação Celular/genética , Embrião de Mamíferos/anormalidades , Deleção de Genes , Redes Reguladoras de Genes , Heterozigoto , Camundongos Endogâmicos C57BL , Modelos Biológicos , Mutação/genética , Crista Neural/patologia , Fator de Transcrição PAX9/deficiência , Ligação Proteica , Transdução de SinaisRESUMO
In recent years, there has been much discussion concerning the cervical fasciae. The aim of this study is to confirm and to describe the development of the alar fascia as well as its relationship with nearby structures. Histological preparations of 25 human embryos (6-8 weeks of development) and 25 human fetuses (9-12 weeks of development) were studied bilaterally using a conventional optical microscope. Our study confirms the existence of the alar fascia and permits three stages to be established during its development. The initial stage (1st), corresponding to the 6th week of development (Carnegie stages 18-19), is characterized by the beginning of the alar fascia primordium in the retroesophageal space at the level of C7-T1. In the formation stage (2nd), corresponding to the 7th and 8th weeks of development (Carnegie stages 20-23), the alar fascia primordium grows upwards and reaches the level of C2-C3. In the maturation stage (3rd), beginning in the 9th week of development, the visceral, alar and prevertebral fasciae can be identified. The alar fascia divides the retrovisceral space (retropharyngeal and retroesophageal) into two spaces: one anterior (between the alar fascia and the visceral fascia and extending from C1 to T1, named retropharyngeal or retroesophageal space according to the level) and the other posterior (between the alar fascia and the prevertebral fascia, named danger space). We suggest that this latter space be named the retroalar space. This study suggests that alar fascia development is related to mechanical factors and that the alar fascia permits the sliding of the pharynx and the oesophagus during swallowing.
Assuntos
Fáscia/embriologia , Pescoço/embriologia , Vértebras Cervicais/embriologia , Humanos , Disco Intervertebral/embriologia , Faringe/embriologiaRESUMO
The gnathostome pharyngeal cavity functions in food transport and respiration. In amniotes the mouth and nares are the only channels allowing direct contact between internal and external epithelia. In teleost fish, gill slits arise through opening of endodermal pouches and connect the pharynx to the exterior. Using transgenic zebrafish lines, cell tracing, live imaging and different markers, we investigated if pharyngeal openings enable epithelial invasion and how this modifies the pharyngeal epithelium. We conclude that in zebrafish the pharyngeal endoderm becomes overlain by cells with a peridermal phenotype. In a wave starting from pouch 2, peridermal cells from the outer skin layer invade the successive pouches until halfway their depth. Here the peridermal cells connect to a population of cells inside the pharyngeal cavity that express periderm markers, yet do not invade from outside. The latter population expands along the midline from anterior to posterior until the esophagus-gut boundary. Together, our results show a novel role for the periderm as an internal epithelium becomes adapted to function as an external surface.
Assuntos
Brânquias/anatomia & histologia , Faringe/anatomia & histologia , Peixe-Zebra/anatomia & histologia , Peixe-Zebra/embriologia , Animais , Animais Geneticamente Modificados , Endoderma/embriologia , Brânquias/embriologia , Faringe/embriologiaRESUMO
In the hindbrain and the adjacent cranial neural crest (NC) cells of jawed vertebrates (gnathostomes), nested and segmentally-restricted domains of Hox gene expression provide a combinatorial Hox-code for specifying regional properties during head development. Extant jawless vertebrates, such as the sea lamprey (Petromyzon marinus), can provide insights into the evolution and diversification of this Hox-code in vertebrates. There is evidence for gnathostome-like spatial patterns of Hox expression in lamprey; however, the expression domains of the majority of lamprey hox genes from paralogy groups (PG) 1-4 are yet to be characterized, so it is unknown whether they are coupled to hindbrain segments (rhombomeres) and NC. In this study, we systematically describe the spatiotemporal expression of all 14 sea lamprey hox genes from PG1-PG4 in the developing hindbrain and pharynx to investigate the extent to which their expression conforms to the archetypal gnathostome hindbrain and pharyngeal hox-codes. We find many similarities in Hox expression between lamprey and gnathostome species, particularly in rhombomeric domains during hindbrain segmentation and in the cranial neural crest, enabling inference of aspects of Hox expression in the ancestral vertebrate embryonic head. These data are consistent with the idea that a Hox regulatory network underlying hindbrain segmentation is a pan vertebrate trait. We also reveal differences in hindbrain domains at later stages, as well as expression in the endostyle and in pharyngeal arch (PA) 1 mesoderm. Our analysis suggests that many Hox expression domains that are observed in extant gnathostomes were present in ancestral vertebrates but have been partitioned differently across Hox clusters in gnathostome and cyclostome lineages after duplication.
Assuntos
Embrião não Mamífero/metabolismo , Evolução Molecular , Regulação da Expressão Gênica no Desenvolvimento , Genes Homeobox , Cabeça/embriologia , Petromyzon/embriologia , Petromyzon/genética , Animais , Faringe/embriologia , Rombencéfalo/embriologiaRESUMO
Pharyngeal pouches, a series of outpocketings that bud from the foregut endoderm, are essential to the formation of craniofacial skeleton as well as several important structures like parathyroid and thymus. However, whether pharyngeal pouch progenitors exist in the developing gut tube remains unknown. Here, taking advantage of cell lineage tracing and transgenic ablation technologies, we identified a population of nkx2.3+ pouch progenitors in zebrafish embryos and demonstrated an essential requirement of ectodermal BMP2b for their specification. At early somite stages, nkx2.3+ cells located at lateral region of pharyngeal endoderm give rise to the pouch epithelium except a subpopulation expressing pdgfαa rather than nkx2.3. A small-scale screen of chemical inhibitors reveals that BMP signaling is necessary to specify these progenitors. Loss-of-function analyses show that BMP2b, expressed in the pharyngeal ectoderm, actives Smad effectors in endodermal cells to induce nkx2.3+ progenitors. Collectively, our study provides in vivo evidence for the existence of pouch progenitors and highlights the importance of BMP2b signaling in progenitor specification.
Assuntos
Proteína Morfogenética Óssea 2/metabolismo , Proteínas de Homeodomínio/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/embriologia , Peixe-Zebra/metabolismo , Animais , Animais Geneticamente Modificados , Padronização Corporal/genética , Padronização Corporal/fisiologia , Proteína Morfogenética Óssea 2/genética , Linhagem da Célula/genética , Linhagem da Célula/fisiologia , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/metabolismo , Endoderma/embriologia , Endoderma/metabolismo , Proteínas de Homeodomínio/genética , Mutação , Faringe/embriologia , Faringe/metabolismo , Fator de Crescimento Derivado de Plaquetas/genética , Fator de Crescimento Derivado de Plaquetas/metabolismo , Transdução de Sinais , Proteínas Smad/metabolismo , Somitos/embriologia , Somitos/metabolismo , Peixe-Zebra/genética , Proteínas de Peixe-Zebra/genéticaRESUMO
In vertebrate embryos, the cardiopharyngeal mesoderm gives rise to both cardiac and branchiomeric head muscles. The canonical Wnt signaling pathway regulates many aspects of cardiomyocyte specification, and modulates a balance between skeletal and cardiac myogenesis during vertebrate head muscle development. However, the role of Wnt signaling during ascidian cardiopharyngeal development remains elusive. Here, we documented the expression of Wnt pathway components during cardiopharyngeal development in Ciona, and generated tools to investigate potential roles for Wnt signaling, and its transcriptional effector Tcf, on heart vs. pharyngeal muscle fate specification. Neither focused functional analyses nor lineage-specific transcriptome profiling uncovered a significant role for Tcf during early cardiac vs. pharyngeal muscle fate choice. By contrast, Wnt gene expression patterns of Frizzled4 and Lrp4/8 and CRISPR/Cas9-mediated Tcf knock-down suggested a later requirement for Wnt signaling during heart morphogenesis and/or cardiomyocyte differentiation. This study provides a provisional set of reagents to study Wnt signaling function in Ciona, and promising insights for future analyses of Wnt functions during heart organogenesis.
Assuntos
Ciona intestinalis/embriologia , Ciona intestinalis/genética , Coração/embriologia , Fatores de Transcrição TCF/metabolismo , Proteínas Wnt/metabolismo , Animais , Padronização Corporal/genética , Linhagem da Célula/genética , Desenvolvimento Embrionário/genética , Regulação da Expressão Gênica no Desenvolvimento , Organogênese/genética , Faringe/embriologia , Fatores de Transcrição TCF/genética , Transcriptoma/genética , Regulação para Cima/genética , Proteínas Wnt/genética , Via de Sinalização Wnt/genéticaRESUMO
Defects in the middle ear ossicles - malleus, incus and stapes - can lead to conductive hearing loss. During development, neural crest cells (NCCs) migrate from the dorsal hindbrain to specific locations in pharyngeal arch (PA) 1 and 2, to form the malleus-incus and stapes, respectively. It is unclear how migratory NCCs reach their proper destination in the PA and initiate mesenchymal condensation to form specific ossicles. We show that secreted molecules sonic hedgehog (SHH) and bone morphogenetic protein 4 (BMP4) emanating from the pharyngeal endoderm are important in instructing region-specific NCC condensation to form malleus-incus and stapes, respectively, in mouse. Tissue-specific knockout of Shh in the pharyngeal endoderm or Smo (a transducer of SHH signaling) in NCCs causes the loss of malleus-incus condensation in PA1 but only affects the maintenance of stapes condensation in PA2. By contrast, knockout of Bmp4 in the pharyngeal endoderm or Smad4 (a transducer of TGFß/BMP signaling) in the NCCs disrupts NCC migration into the stapes region in PA2, affecting stapes formation. These results indicate that region-specific endodermal signals direct formation of specific middle ear ossicles.
Assuntos
Ossículos da Orelha/embriologia , Endoderma/embriologia , Endoderma/metabolismo , Crista Neural/citologia , Transdução de Sinais , Animais , Proteínas Morfogenéticas Ósseas/metabolismo , Movimento Celular , Sobrevivência Celular , Deleção de Genes , Proteínas Hedgehog , Bigorna/embriologia , Bigorna/metabolismo , Martelo/embriologia , Martelo/metabolismo , Camundongos , Modelos Biológicos , Crista Neural/embriologia , Crista Neural/metabolismo , Especificidade de Órgãos , Faringe/embriologia , Fenótipo , Estribo/embriologia , Estribo/metabolismo , Fatores de Tempo , Fator de Crescimento Transformador beta/metabolismoRESUMO
Craniofacial malformations are common congenital birth defects and usually caused by abnormal development of the cranial neural crest cells. Some nucleolar ribosome biogenesis factors are implicated in neural crest disorders also known as neurocristopathies. However, the underlying mechanisms linking ribosome biogenesis and neural crest cell (NCC) development remain to be elucidated. Here we report a novel zebrafish model with a CRISPR/Cas9-generated esf1 mutation, which exhibits severe NCC-derived pharyngeal cartilage loss and defects in the eyes, brain, and heart. The expression of several typical NCC markers, including sox10, dlx2a, nrp2b, crestin, vgll2a, and sox9a, was reduced in the head of the esf1 mutants, which indicates that esf1 plays a role in the development of zebrafish NCCs. We demonstrate that, similar to the yeast, loss of esf1 in zebrafish leads to defects in 18S rRNA biogenesis and ribosome biogenesis. We also show strong upregulation of p53 signaling as well as apoptosis, and poor proliferation in mutants. Inactivation of p53 rescues the early tissue defects and pharyngeal cartilage loss observed in esf1 mutants, indicating that increased cell death and pharyngeal cartilage defects observed in esf1 mutants are mediated via upregulated p53 signaling pathways. Based on transplantation analysis, we found esf1 functions in NCC in a cell autonomous fashion. Together, our results suggest that esf1 is required for NCC development and pharyngeal cartilage formation. These studies provide a potential model for investigating the relationship between ribosome biogenesis defects and craniofacial neurocristopathies.
Assuntos
Cartilagem/embriologia , Embrião não Mamífero/citologia , Proteínas Nucleares/metabolismo , Faringe/embriologia , Proteína Supressora de Tumor p53/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/embriologia , Animais , Animais Geneticamente Modificados/embriologia , Animais Geneticamente Modificados/genética , Animais Geneticamente Modificados/metabolismo , Padronização Corporal , Cartilagem/metabolismo , Embrião não Mamífero/metabolismo , Mutação , Proteínas Nucleares/genética , Faringe/metabolismo , Ribossomos/metabolismo , Proteína Supressora de Tumor p53/genética , Peixe-Zebra/genética , Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/genéticaRESUMO
Microdeletions involving TBX1 result in variable congenital malformations known collectively as 22q11.2 deletion syndrome (22q11.2DS). Tbx1-deficient mice and zebrafish recapitulate several disease phenotypes, including pharyngeal arch artery (PAA), head muscle (HM), and cardiac outflow tract (OFT) deficiencies. In zebrafish, these structures arise from nkx2.5+ progenitors in pharyngeal arches 2-6. Because pharyngeal arch morphogenesis is compromised in Tbx1-deficient animals, the malformations were considered secondary. Here, we report that the PAA, HM, and OFT phenotypes in tbx1 mutant zebrafish are primary and arise prior to pharyngeal arch morphogenesis from failed specification of the nkx2.5+ pharyngeal lineage. Through in situ analysis and lineage tracing, we reveal that nkx2.5 and tbx1 are co-expressed in this progenitor population. Furthermore, we present evidence suggesting that gdf3-ALK4 signaling is a downstream mediator of nkx2.5+ pharyngeal lineage specification. Collectively, these studies support a cellular mechanism potentially underlying the cardiovascular and craniofacial defects observed in the 22q11.2DS population.
Assuntos
Síndrome da Deleção 22q11/patologia , Diferenciação Celular , Células-Tronco Embrionárias/citologia , Faringe/embriologia , Síndrome da Deleção 22q11/genética , Animais , Linhagem da Célula , Células-Tronco Embrionárias/metabolismo , Proteína Homeobox Nkx-2.5/genética , Proteína Homeobox Nkx-2.5/metabolismo , Faringe/citologia , Fenótipo , Proteínas com Domínio T/genética , Peixe-Zebra , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismoRESUMO
The larval pharynx of the cephalochordate Branchiostoma (amphioxus) is asymmetrical. The mouth is on the left, and endostyle and gill slits are on the right. At the neurula, Nodal and Hedgehog (Hh) expression becomes restricted to the left. To dissect their respective roles in gill slit formation, we inhibited each pathway separately for 20â min at intervals during the neurula stage, before gill slits penetrate, and monitored the effects on morphology and expression of pharyngeal markers. The results pinpoint the short interval spanning the gastrula/neurula transition as the critical period for specification and positioning of future gill slits. Thus, reduced Nodal signaling shifts the gill slits ventrally, skews the pharyngeal domains of Hh, Pax1/9, Pax2/5/8, Six1/2 and IrxC towards the left, and reduces Hh and Tbx1/10 expression in endoderm and mesoderm, respectively. Nodal auto-regulates. Decreased Hh signaling does not affect gill slit positions or Hh or Nodal expression, but it does reduce the domain of Gli, the Hh target, in the pharyngeal endoderm. Thus, during the neurula stage, Nodal and Hh cooperate in gill slit development - Hh mediates gill slit formation and Nodal establishes their left-right position.
Assuntos
Padronização Corporal , Brânquias/metabolismo , Proteínas Hedgehog/metabolismo , Anfioxos/embriologia , Anfioxos/metabolismo , Proteína Nodal/metabolismo , Animais , Benzodioxóis/farmacologia , Padronização Corporal/efeitos dos fármacos , Padronização Corporal/genética , Embrião não Mamífero/efeitos dos fármacos , Embrião não Mamífero/metabolismo , Epistasia Genética/efeitos dos fármacos , Gástrula/efeitos dos fármacos , Gástrula/embriologia , Gástrula/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Brânquias/efeitos dos fármacos , Proteínas Hedgehog/genética , Imidazóis/farmacologia , Anfioxos/efeitos dos fármacos , Anfioxos/genética , Larva/efeitos dos fármacos , Larva/metabolismo , Mesoderma/efeitos dos fármacos , Mesoderma/embriologia , Mesoderma/metabolismo , Proteína Nodal/genética , Faringe/efeitos dos fármacos , Faringe/embriologia , Faringe/metabolismo , Piridinas/farmacologia , Alcaloides de Veratrum/farmacologiaRESUMO
The pharyngeal pouches are a series of epithelial outgrowths of the foregut endoderm. Pharyngeal pouches segment precursors of the vertebrate face into pharyngeal arches and pattern the facial skeleton. These pouches fail to develop normally in zebrafish foxi1 mutants, yet the role Foxi1 plays in pouch development remains to be determined. Here we show that ectodermal Foxi1 acts downstream of Fgf8a during the late stage of pouch development to promote rearrangement of pouch-forming cells into bilayers. During this phase, foxi1 and wnt4a are coexpressed in the facial ectoderm and their expression is expanded in fgf8a mutants. foxi1 expression is unaffected in wnt4a mutants; conversely, ectodermal wnt4a expression is abolished in foxi1 mutants. Consistent with this, foxi1 mutant pouch and facial skeletal defects resemble those of wnt4a mutants. These findings suggest that ectodermal Foxi1 mediates late-stage pouch morphogenesis through wnt4a expression. We therefore propose that Fox1 activation of Wnt4a in the ectoderm signals the epithelial stabilization of pouch-forming cells during late-stage of pouch morphogenesis.
Assuntos
Ectoderma/embriologia , Fatores de Transcrição Forkhead/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Organogênese/fisiologia , Faringe/embriologia , Proteína Wnt4/biossíntese , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/embriologia , Animais , Fatores de Crescimento de Fibroblastos/genética , Fatores de Crescimento de Fibroblastos/metabolismo , Fatores de Transcrição Forkhead/genética , Mutação , Via de Sinalização Wnt/fisiologia , Proteína Wnt4/genética , Peixe-Zebra/genética , Proteínas de Peixe-Zebra/genéticaRESUMO
The migration and distribution patterns of neural crest (NC) cells reflect the distinct embryonic environments of the head and trunk: cephalic NC cells migrate predominantly along the dorsolateral pathway to populate the craniofacial and pharyngeal regions, whereas trunk crest cells migrate along the ventrolateral pathways to form the dorsal root ganglia. These two patterns thus reflect the branchiomeric and somitomeric architecture, respectively, of the vertebrate body plan. The so-called vagal NC occupies a postotic, intermediate level between the head and trunk NC. This level of NC gives rise to both trunk- and cephalic-type (circumpharyngeal) NC cells. The anatomical pattern of the amphioxus, a basal chordate, suggests that somites and pharyngeal gills coexist along an extensive length of the body axis, indicating that the embryonic environment is similar to that of vertebrate vagal NC cells and may have been ancestral for vertebrates. The amniote-like condition in which the cephalic and trunk domains are distinctly separated would have been brought about, in part, by anteroposterior reduction of the pharyngeal domain.