Search

Background: Skeletal muscle in the trunk derives from the somites, paired segments of paraxial mesoderm. Whereas axial musculature develops within the somite, appendicular muscle develops following migration of muscle precursors into lateral plate mesoderm. The development of muscles bridging axial and appendicular systems appears mixed., Results: We examine development of three migratory muscle precursor-derived muscles in zebrafish: the sternohyoideus (SH), pectoral fin (PF), and posterior hypaxial (PHM) muscles. We show there is an anterior to posterior gradient to the developmental gene expression and maturation of these three muscles. SH muscle precursors exhibit a long delay between migration and differentiation, PF muscle precursors exhibit a moderate delay in differentiation, and PHM muscle precursors show virtually no delay between migration and differentiation. Using lineage tracing, we show that lateral plate contribution to the PHM muscle is minor, unlike its known extensive contribution to the PF muscle and absence in the ventral extension of axial musculature., Conclusions: We propose that PHM development is intermediate between a migratory muscle mode and an axial muscle mode of development, wherein the PHM differentiates after a very short migration of its precursors and becomes more anterior primarily by elongation of differentiated muscle fibers., (© 2019 Wiley Periodicals, Inc.)

The embryonic development of the bullseye puffer, Sphoeroides annulatus, was characterized on the basis of the theory of saltatory ontogeny. This theory predicts a correlative relationship between the ontogeny-type in an altricial-precocial spectrum and the habitat that a species occupies within an unstable-stable environmental spectrum. Because S. annulatus inhabits a variety of unstable environments along a wide latitudinal range, the hypothesis that this species presents one of the most altricial embryonic developments among tetraodontids was tested. Based on major developmental events that marked the ontogenetic thresholds nine embryonic steps were identified. Developmental features such as small adhesives eggs, lack of vitelline circulation, small free embryos swimming up at hatching guided by positive phototaxis, and small first-feeding larvae actively swam in the water column, suggest that S. annulatus belongs to the reproductive guild of the nonguarders-lithopelagophils. Moreover, a comparative analysis of the developmental sequences, egg size, and first-feeding larvae size between tetraodontids confirms the hypothesis of this study and supports the evolutionary principle of the altricial-precocial spectrum postulated in the theory of saltatory ontogeny., (© 2019 Wiley Periodicals, Inc.)

Background: Myostatin (MSTN), a member of the transforming growth factor-β (TGF-β) superfamily, has been implicated in the negative regulation of skeletal myogenesis. However, the molecular mechanism through which MSTN regulates early embryonic myogenesis is not well understood., Results: We demonstrate that MSTN regulates early embryonic myogenesis by promoting the epithelial-to-mesenchymal transition (EMT) of the dermomyotome during somitogenesis in chicks. We show that the MSTN gene is first expressed at the center of the dermomyotome. As somitogenesis progresses, its expression extends dorsally and ventrally along the plane of the dermomyotome. By combining in situ hybridization and immunofluorescence assays, we demonstrate that the expression pattern of MSTN is spatiotemporally well correlated with EMT of the dermomyotome. Our gain- and loss-of-function experiments further reveal that MSTN can induce EMT of the chick dermomyotome. We also show that MSTN induces EMT of a nonsmall cell lung carcinoma cell line (A549) and Madin-Darby canine kidney cells in vitro., Conclusions: Our experimental data suggest that MSTN regulates myogenesis by promoting EMT during somitogenesis. These findings provide novel insights into the functions of MSTN during early embryonic myogenesis. Developmental Dynamics 247:1241-1252, 2018. © 2018 Wiley Periodicals, Inc., (© 2018 Wiley Periodicals, Inc.)

Using multiple taxa to research development is necessary for making general conclusions about developmental patterns and mechanisms. We present a staging series for Cyprinodon variegatus as a basis for further study of the developmental biology of fishes in the genus Cyprinodon and for comparative work on teleost fishes beyond the standard models. Cyprinodon are small, euryhaline fishes, widely distributed in fresh, brackish, and hypersaline waters of southern and eastern North America. Cyprinodontids are closely related to fundulids, providing a comparative reference point to the embryological model, Fundulus heteroclitus. Ecologists and evolutionary biologists commonly study Cyprinodon, and we have been using Cyprinodon to study skull variation and its genetic basis among closely related species. We divided embryonic development of C. variegatus into 34 morphologically identifiable stages. We reference our staging series to that already defined for a related model species, Oryzias latipes (medaka) that is studied by a large community of researchers. We provide a description of the early chondrogenesis and ossification of skull and caudal fin bones during the latter stages of embryonic development. We show that Cyprinodon are tractable for studying development. Eggs can be obtained easily from breeding pairs and our study provides a staging system to facilitate future developmental studies., (© 2018 Wiley Periodicals, Inc.)

Background: Hematopoietic stem and progenitor cells (HSPCs) are generated de novo in the embryo in a process termed the endothelial to hematopoietic transition (EHT). EHT is most extensively studied in the yolk sac and dorsal aorta. Recently new sites of hematopoiesis have been described, including the heart, somites, head, and venous plexus of the yolk sac., Results: We examined sites of HSPC formation in well-studied and in less well-known sites by mapping the expression of the key EHT factor Runx1 along with several other markers by means of confocal microscopy. We identified sites of HSPC formation in the head, heart and somites. We also identified sites of HSPC formation in both the arterial and venous plexuses of the yolk sac, and show that progenitors with lymphoid potential are enriched in hematopoietic clusters in close proximity to arteries. Furthermore, we demonstrate that many of the cells in hematopoietic clusters resemble monocytes or granulocytes based on nuclear shape., Conclusions: We identified sites of HSPC formation in the head, heart, and somites, confirming that embryonic hematopoiesis is less spatially restricted than previously thought. Furthermore, we show that HSPCs in the yolk sac with lymphoid potential are located in closer proximity to arteries than to veins. Developmental Dynamics 245:1011-1028, 2016. © 2016 Wiley Periodicals, Inc., (© 2016 Wiley Periodicals, Inc.)

Background: In most vertebrates, the segmentation of the paraxial mesoderm involves the formation of metameric units called somites through a mesenchymal-epithelial transition. However, this process is different in Xenopus laevis because it does not form an epithelial somite. Xenopus somitogenesis is characterized by a complex cells rearrangement that requires the coordinated regulation of cell shape, adhesion, and motility. The molecular mechanisms that control these cell behaviors underlying somite formation are little known. Although the Paraxis has been implicated in the epithelialization of somite in chick and mouse, its role in Xenopus somite morphogenesis has not been determined., Results: Using a morpholino and hormone-inducible construction approaches, we showed that both gain and loss of function of paraxis affect somite elongation, rotation and alignment, causing a severe disorganization of somitic tissue. We further found that depletion or overexpression of paraxis in the somite led to the downregulation or upregulation, respectively, of cell adhesion expression markers. Finally, we demonstrated that paraxis is necessary for the proper expression of myotomal and sclerotomal differentiation markers., Conclusions: Our results demonstrate that paraxis regulates the cell rearrangements that take place during the somitogenesis of Xenopus by regulating cell adhesion. Furthermore, paraxis is also required for somite differentiation., (© 2015 Wiley Periodicals, Inc.)

Background: Six1 plays an important role in the development of several vertebrate organs, including cranial sensory placodes, somites, and kidney. Although Six1 mutations cause one form of branchio-otic syndrome (BOS), the responsible gene in many patients has not been identified; genes that act downstream of Six1 are potential BOS candidates., Results: We sought to identify novel genes expressed during placode, somite and kidney development by comparing gene expression between control and Six1-expressing ectodermal explants. The expression patterns of 19 of the significantly up-regulated and 11 of the significantly down-regulated genes were assayed from cleavage to larval stages. A total of 28/30 genes are expressed in the otocyst, a structure that is functionally disrupted in BOS, and 26/30 genes are expressed in the nephric mesoderm, a structure that is functionally disrupted in the related branchio-otic-renal (BOR) syndrome. We also identified the chick homologues of five genes and show that they have conserved expression patterns., Conclusions: Of the 30 genes selected for expression analyses, all are expressed at many of the developmental times and appropriate tissues to be regulated by Six1. Many have the potential to play a role in the disruption of hearing and kidney function seen in BOS/BOR patients., (© 2014 Wiley Periodicals, Inc.)

Background: Stromal derived factor-1α (sdf-1α), a chemoattractant chemokine, plays a major role in tumor growth, angiogenesis, metastasis, and in embryogenesis. The sdf-1α signaling pathway has also been shown to be important for somite rotation in zebrafish (Hollway et al., 2007). Given the known similarities and differences between zebrafish and Xenopus laevis somitogenesis, we sought to determine whether the role of sdf-1α is conserved in Xenopus laevis., Results: Using a morpholino approach, we demonstrate that knockdown of sdf-1α or its receptor, cxcr4, leads to a significant disruption in somite rotation and myotome alignment. We further show that depletion of sdf-1α or cxcr4 leads to the near absence of β-dystroglycan and laminin expression at the intersomitic boundaries. Finally, knockdown of sdf-1α decreases the level of activated RhoA, a small GTPase known to regulate cell shape and movement., Conclusion: Our results show that sdf-1α signaling regulates somite cell migration, rotation, and myotome alignment by directly or indirectly regulating dystroglycan expression and RhoA activation. These findings support the conservation of sdf-1α signaling in vertebrate somite morphogenesis; however, the precise mechanism by which this signaling pathway influences somite morphogenesis is different between the fish and the frog., (Copyright © 2013 Wiley Periodicals, Inc.)

The segmental series of somites in the vertebrate embryo gives rise to the axial skeleton. In amniote models, single vertebrae are derived from the sclerotome of two adjacent somites. This process, known as resegmentation, is well-studied using the quail-chick chimeric system, but the presumed generality of resegmentation across vertebrates remains poorly evaluated. Resegmentation has been questioned in anamniotes, given that the sclerotome is much smaller and lacks obvious differentiation between cranial and caudal portions. Here, we provide the first experimental evidence that resegmentation does occur in a species of amphibian. Fate mapping of individual somites in the Mexican axolotl (Ambystoma mexicanum) revealed that individual vertebrae receive cells from two adjacent somites as in the chicken. These findings suggest that large size and segmentation of the sclerotome into distinct cranial and caudal portions are not requirements for resegmentation. Our results, in addition to those for zebrafish, indicate that resegmentation is a general process in building the vertebral column in vertebrates, although it may be achieved in different ways in different groups., (Copyright © 2013 Wiley Periodicals, Inc.)

Background: In the vertebrate embryo, skeletal muscle and the axial skeleton arise from the somites. Patterning of the somites into the respective somite compartments, namely dermomyotome, myotome, and sclerotome, depends on molecular signals from neighboring structures, including surface ectoderm, neural tube, notochord, and lateral plate mesoderm. A potential role of the intermediate mesoderm, notably the Wolffian or nephric duct, in somite development is poorly understood., Results: We studied somite compartmentalization as well as muscular and skeletal development after surgical ablation of the early Wolffian duct anlage, which lead to loss of the Wolffian duct and absence of the mesonephros, whereas Pax2 expression in the nephrogenic mesenchyme was temporarily maintained. We show that somite compartments, as well as the somite derivatives, skeletal muscle and the cartilaginous skeleton, develop normally in the absence of the Wolffian duct., Conclusions: Our results indicate that development of the musculoskeletal system is independent of the Wolffian duct as a signaling center., (Copyright © 2013 Wiley Periodicals, Inc.)

Background: During segmentation of the zebrafish embryo, inside-out signaling activates Integrin α5, which is necessary for somite border morphogenesis. The direct activator of Integrin α5 during this process is unknown. One candidate is Rap1b, a small monomeric GTPase implicated in Integrin activation in the immune system., Results: Knockdown of rap1b, or overexpression of a dominant negative rap1b, causes a mild axis elongation defect in zebrafish. However, disruption of rap1b function in integrin α5(-/-) mutants results in a strong reduction in Fibronectin (FN) matrix assembly in the paraxial mesoderm and a failure in somite border morphogenesis along the entire anterior-posterior axis. Somite patterning appears unaffected, as her1 oscillations are maintained in single and double morphants/mutants, but somite polarity is gradually lost in itgα5(-/-) ; rap1b MO embryos., Conclusions: In itgα5(-) (/) (-) mutants, rap1b is required for proper somite border morphogenesis in zebrafish. The loss of somite borders is not a result of aberrant segmental patterning. Rather, somite boundary formation initiates but is not completed, due to the failure to assemble FN matrix along the nascent boundary. We propose a model in which Rap1b activates Integrin/Fibronectin receptors as part of an "inside-out" signaling pathway that promotes Integrin binding to FN, FN matrix assembly, and subsequent stabilization of morphological somite boundaries., (Copyright © 2012 Wiley Periodicals, Inc.)

Background: Repulsive guidance molecules (RGM) are high-affinity ligands for the Netrin receptor Neogenin, and they are crucial for nervous system development including neural tube closure; neuronal and neural crest cell differentiation and axon guidance. Recent studies implicated RGM molecules in bone morphogenetic protein signaling, which regulates a variety of developmental processes. Moreover, a role for RGMc in iron metabolism has been established. This suggests that RGM molecules may play important roles in non-neural tissues., Results: To explore which tissues and processed may be regulated by RGM molecules, we systematically investigated the expression of RGMa and RGMb, the only RGM molecules currently known for avians, in the chicken embryo., Conclusions: Our study suggests so far unknown roles of RGM molecules in notochord, somite and skeletal muscle development., (Copyright © 2012 Wiley Periodicals, Inc.)

Somitogenesis is a complex process during early vertebrate development involving interactions between many factors to form a bilateral somite series. A role for chromatin remodelers in somitogenesis has not yet been demonstrated. Here, we investigate the function of chromodomain helicase DNA binding protein 7 (chd7) during zebrafish somitogenesis. We show that Chd7 deficiency leads to asymmetric segmentation of the presomitic mesoderm (PSM), as revealed by expression of the somitogenesis genes, cdx1a, dlc, her7, mespa, and ripply1. Moreover, we show that abrogation of Chd7 results in the loss of asymmetric expression of spaw in the lateral plate mesoderm, which is consistent with more general laterality defects. Based on the observation that insufficient Chd7 leads to left-right asymmetry defects during PSM segmentation, and because CHD7 has been linked to human spinal deformities, we suggest that zebrafish chd7 morphants may be a good in vivo model to examine the pathophysiology of these diseases., (Copyright © 2011 Wiley-Liss, Inc.)

The highly conserved POU genes encode homeodomain transcription factors involved in various developmental events, with some, the Brn genes, playing key roles in neurogenesis. We investigated the evolutionary relationships between these genes, by studying the POU gene complement of a model teleost, the medaka (Oryzias latipes). We identified 17 POU genes and carried out a comprehensive in situ hybridization analysis focusing on the optic tectum, a cortical structure of the mesencephalon, in which cell positions and their differentiation states are spatially and temporally correlated. Six POU genes displayed patterned expression in the optic tectum: two genes were expressed in the center of the organ (a zone with differentiated neurons), two in an intermediate zone in which cells exit the cell cycle and two in the peripheral proliferation zone. These results suggest that POU genes may play key roles in both late neurogenesis and in multipotent neural progenitors., (Copyright © 2011 Wiley-Liss, Inc.)

Ephrins and their tyrosine kinase receptors (Ephs) are a highly conserved family of signaling proteins with various functions during embryonic development. Among others, Eph/ephrin signaling is involved in regulating axon guidance, cell migration, and tissue border formation through inducing modifications of the actin cytoskeleton and cell adhesion. During development ephrins and Ephs are expressed in spatially and temporarily regulated patterns in a wide range of tissues. Here, we analyzed the expression of seven members of the Eph and four member of the ephrin family during early stages of mouse inner ear development by whole-mount in situ hybridization. We detected expressions of EphA2, EphA4, EphA7, EphB1, ephrinA4, and ephrinA5 in and around the forming otic placode between embryonic day (E) 8.5 and E10, and report their detailed expression patterns. Our results reveal dynamic expression of several members of the ephrin/Eph family consistent with functions in otic placode development, invagination and neuroblast delamination., (Copyright © 2011 Wiley-Liss, Inc.)

The chick, Gallus gallus, is the traditional model in avian developmental studies. Data on other bird species are scarce. Here, we present a comparative study of the embryonic development of the chick and the emu Dromaius novaehollandiae, a member of Paleognathae, which also includes the ostrich, rhea, tinamou, kiwi, and cassowary. Emu embryos ranging from Hamburger and Hamilton (HH) equivalent stages 1 to 43 were collected and their gross morphology analyzed. Its early development was studied in detail with time-lapse imaging and molecular techniques. Emu embryos in general take 2-3 times longer incubation time to reach equivalent chicken stages, requiring 1 day for HH2, 2.5 days for HH4, 7 days for limb bud initiation, 23 days for feather germ appearance, and approximately 50-56 days for hatching. Chordin gene expression is similar in emu and chick embryos, and emu Brachyury is not expressed until HH3. Circulation is established at approximately the 27- to 30-somite stage. Forelimb buds are formed and patterned initially, but their growth is severely retarded. The size difference between an emu and a chick embryo only becomes apparent after limb bud formation. Overall, emu and chick embryogenesis proceeds through similar stages, but developmental heterochrony between these two species is widely observed., (© 2010 Wiley-Liss, Inc.)

Filamin proteins cross-link F-actin and form a scaffold for numerous signal transduction systems. In this study, we show that filaminB is apically enriched in avian embryonic epithelium, and colocalizes with cell adhesion molecules and circumferential F-actin. FilaminB knockdown in the neural tube and somites decreases the accumulation of N-cadherin and ZO-1 protein at cell junctions, and promotes disruption of these tissues and the presence of neuronal aggregates within the lumen of the neural tube. This phenotype resembles that of human congenital condition, periventricular heterotopia (PH). FilaminB knockdown in MDCK cells suggests that filaminB is required for the apical accumulation of adhesion molecules in the junctional complex and subsequent epithelium formation. We further suggest that the reduction of structural integrity of the neural epithelium caused by the loss of Filamin function may also result in formation of the neuronal nodules found in PH patients., (© 2010 Wiley-Liss, Inc.)

The Notch signalling pathway mutants, after-eight (aei), beamter (bea), and deadly-seven (des) have previously been used to study somitogenesis and neurogenesis. Notch signalling has also been shown to have roles in vascular development. However, vascular development in each of these three Notch mutants has not been described, and so their potential usefulness for further understanding the role of Notch signalling in angiogenesis is unknown. Here we demonstrate each of the mutants also exhibit vascular defects in inter-somitic vessel (ISV) positioning and patterning. Ectopic filopodia were also observed on the ISVs of the mutants. Ectopic filopodia are not due to loss of dll4. Somite expression of known vascular guidance cues, efnb2, sema3a2, and plexinD1 are disrupted, suggesting that the ISV vascular phenotype is due to disruption of these cues.

Somites give rise to the vertebral column and segmented musculature of adult vertebrates. The cell movements that position cells within somites along the anteroposterior and dorsoventral axes are not well understood. Using a fate mapping approach, we show that at the onset of Xenopus laevis gastrulation, mesoderm cells undergo distinct cell movements to form myotome fibers positioned in discrete locations within somites and along the anteroposterior axis. We show that the distribution of presomitic cells along the anteroposterior axis is influenced by convergent and extension movements of the notochord. Heterochronic and heterotopic transplantations between presomitic gastrula and early tail bud stages show that these cells are interchangeable and can form myotome fibers in locations determined by the host embryo. However, additional transplantation experiments revealed differences in the competency of presomitic cells to form myotome fibers, suggesting that maturation within the tail bud presomitic mesoderm is required for myotome fiber differentiation.

Somitic beta-catenin is involved in both maintaining a stem cell population and controlling myogenic differentiation. It is unclear how beta-catenin-dependent Wnt signaling accomplishes these disparate roles. The present study shows that only dorsal cells in the early somite respond to beta-catenin-dependent Wnt signaling and as the somites compartmentalize to form the dermomyotome and myotome, responding cells are detected primarily in the dorsomedial lip (DML). Forced activation of Wnt target genes in DML cells prevents their progeny from entering the myotome, while blocking activation allows myotomal entry. This suggests a role for beta-catenin-dependent/Wnt signaling in maintaining progenitor cells in the DML and that if beta-catenin-dependent/Wnt signaling is required to induce myogenesis, the response is transitory and rapidly down-regulated., ((c) 2009 Wiley-Liss, Inc.)

The process of segmentation in vertebrates is described by a clock and wavefront model consisting of a Notch signal and an fibroblast growth factor-8 (FGF8) gradient, respectively. To further investigate the segmentation process, we screened gene expression profiles for downstream targets of the segmentation clock. The Rnd1 and Rnd3 GTP-binding proteins comprise a subgroup of the Rho GTPase family that show a specific expression pattern similar to the Notch signal component ESR5, suggesting an association between Rnd1/3 and the segmentation clock. Rnd1/3 expression patterns are disrupted by overexpression of dominant-negative or active forms of Notch signaling genes, and responds to the FGF inhibitor SU5402 by a posterior shift analogous to other segmentation-related genes, suggesting that Rnd1/3 expressions are regulated by the segmentation clock machinery. We also show that antisense morpholino oligonucleotides to Rnd1/3 inhibit somite segmentation and differentiation in Xenopus embryos. These results suggest that Rnd1/3 are required for Xenopus somitogenesis., (Copyright 2009 Wiley-Liss, Inc.)

Somites are segmental units of the mesoderm in vertebrate embryos that give rise to the axial skeleton, muscle, and dermis. Somitogenesis occurs in a periodic manner and is governed by a segmentation clock that causes cells to undergo repeated oscillations of gene expression. Here, we present a detailed analysis of cis-regulatory elements that control oscillating expression of the zebrafish her1 gene in the anterior presomitic mesoderm. We identify binding sites for Her proteins and demonstrate that they are necessary for transcriptional repression. This result confirms that direct negative autoregulation of her gene expression constitutes part of the oscillator mechanism. We also characterize binding sites for fused somites/Tbx24 and Suppressor of Hairless proteins and show that they are required for activation of her1 expression. These data provide the foundation for a precise description of the regulatory grammar that defines oscillating gene expression in the zebrafish segmentation clock., (Copyright 2009 Wiley-Liss, Inc.)

Three consecutive pairs of head cavities (premandibular, mandibular, and hyoid) found in elasmobranchs have been considered as remnants of preotic 'head' somites-serial homologues of the myotomic compartments of trunk somites that give rise to the extraoccular musculature. Here, we study a more derived vertebrate, and show that cavitation is more complex in the head of Crocodylus niloticus, than just the occurrence of three pairs of cavities. Apart from the premandibular cavities, paired satellite microcavities, and unpaired extrapremandibular microcavities are recognized in the prechordal region as well. We observed that several developmental phenomena occur at the same time as the formation of the head cavities (premandibular, satellite, extrapremandibular, mandibular, and hyoid) appear temporarily in the crocodile embryo. These are 1) rapid growth of the optic stalk and inflation of the optic vesicle; 2) release of the intimate topographical relationships between the neural tube, notochord and oral gut; 3) tendency of the prechordal mesenchyme to follow the curvature of the forebrain; and 4) proliferation of the prechordal mesenchyme. On the basis of volumetric characters, only the hyoid cavity and hyoid condensation is comparable to the trunk somitocoel and somite, respectively., (Copyright 2009 Wiley-Liss, Inc.)

Dishevelled (Dvl) proteins are key transducers of Wnt signaling encoded by members of a multi-gene family in vertebrates. We report here the divergent, tissue-specific expression patterns for all three Dvl genes in Xenopus embryos, which contrast dramatically with their expression patterns in mice. Moreover, we find that the expression patterns of Dvl genes in the chick diverge significantly from those of Xenopus. In addition, in hemichordates, an outgroup to chordates, we find that the one Dvl gene is dynamically expressed in a tissue-specific manner. Using knockdowns, we find that Dvl1 and Dvl2 are required for early neural crest specification and for somite segmentation in Xenopus. Most strikingly, we report a novel role for Dvl3 in the maintenance of gene expression in muscle and in the development of the Xenopus sclerotome. These data demonstrate that the expression patterns and developmental functions of specific Dvl genes have diverged significantly during chordate evolution., (Copyright (c) 2009 Wiley-Liss, Inc.)

Dystroglycan (Dg) is a cell adhesion receptor for laminin that has been reported to play a role in skeletal muscle cell stability, cytoskeletal organization, cell polarity, and signaling. Here we show that Dg is expressed at both the notochord/somite and the intersomitic boundaries, where laminin and fibronectin are accumulated during somitogenesis. Inhibition of Dg function with morpholino antisense oligonucleotides or a dominant negative mutant results in the normal segmentation of the presomitic mesoderm but affects the number, the size, and the integrity of somites. Depletion of Dg disrupts proliferation and alignment of myoblasts without affecting XMyoD and XMRF4 expression. It also leads to defects in laminin deposition at the intersomitic junctions, whereas expression of integrin beta1 subunits and fibronectin assembly occur normally. Our results show that Dg is critical for both proliferation and elongation of somitic cells and that the Dg-cytoplasmic domain is required for the laminin assembly at the intersomitic boundaries. Developmental Dynamics 238:1332-1345, 2009. (c) 2008 Wiley-Liss, Inc.

The vertebrate mid-hindbrain boundary (MHB) is a crucial morphological structure required for patterning and neural differentiation of the midbrain and anterior hindbrain. We isolated a novel zebrafish mutant, MHB gone (mgo), that exhibited a defective MHB. Expression of engrailed3 in the prospective MHB was absent at the 1-somite stage, suggesting that initiation of the isthmic organizer was disrupted in mgo mutants. Complementation test with mgo and noi, in which the pax2a gene is mutated, infer that the mgo mutant may represent a novel noi allele. However, pronephric, otic vesicle, and commissural axonal defects described in noi mutants were not associated with mgo mutants. Genetic mapping revealed that the mgo mutation is linked to the Pax2a locus, but no mutation was detected in pax2a exons or within intron-exon boundaries. Based on these findings, we propose that the mgo mutation genetically interacts with pax2a required for the initiation of MHB formation., (Copyright 2009 Wiley-Liss, Inc.)

The vertebrate retina is very well conserved in evolution. Its structure and functional features are very similar in phyla as different as primates and teleost fish. Here, we describe the spatiotemporal characteristics of neurogenesis in the retina of a teleost, medaka, and compare them with other species, primarily the zebrafish. Several intriguing differences are observed between medaka and zebrafish. For example, photoreceptor differentiation in the medaka retina starts independently in two different areas, and at more advanced stages of differentiation, medaka and zebrafish retinae display obviously different patterns of the photoreceptor cell mosaic. Medaka and zebrafish evolutionary lineages are thought to have separated from each other 110 million years ago, and so the differences between these species are not unexpected, and may be exploited to gain insight into the architecture of developmental pathways. Importantly, this work highlights the benefits of using multiple teleost models in parallel to understand a developmental process., ((c) 2008 Wiley-Liss, Inc.)

T-box transcription factor tbx6 and basic-helix-loop-helix transcription factor pMesogenin1 are reported to be involved in paraxial mesodermal differentiation. To clarify the relationship between these genes in Xenopus laevis, we isolated pMesogenin2, which showed high homology with pMesogenin1. Both pMesogenin1 and 2 appeared to be transcriptional activators and were induced by a hormone-inducible version of Xtbx6 without secondary protein synthesis in animal cap assays. The pMesogenin2 promoter contained three potential T-box binding sites with which Xtbx6 protein was shown to interact, and a reporter gene construct containing these sites was activated by Xtbx6. Xtbx6 knockdown reduced pMesogenin1 and 2 expressions, but not vice versa. Xtbx6 and pMesogenin1 and 2 knockdowns caused similar phenotypic abnormalities including somite malformation and ventral body wall muscle hypoplasia, suggesting that Xtbx6 is a direct regulator of pMesogenin1 and 2, which are both involved in somitogenesis and myogenesis including that of body wall muscle in Xenopus laevis., ((c) 2008 Wiley-Liss, Inc.)

Eya1 is a homolog of eyes absent in Drosophila, and essential for various organ formations in vertebrates. Mouse and chick Eya1 shows dynamic expression pattern in early development. We identified ten independent Eya1 enhancers by screening evolutionarily conserved sequences. They exhibited enhancer activities in Hensen's node, neural tube, migrating neural crest cells, otic vesicle, olfactory placode, cranial ganglia, and somites at HH6-17 of chick embryo. The sum of the enhancer activities of the enhancers covers the endogenous expression domains of Eya1 common to chick and mouse. Enhancer activities were also observed in species-specific expression domains such as trigeminal ganglia and brain. Mutational study of one of the enhancers revealed that the enhancer is composed of positive and negative cis-regulatory elements. Thus, we successfully identified a comprehensive group of enhancers around Eya1 locus, which are probably involved in the control of the complex expression pattern of Eya1 in vivo., (Copyright (c) 2008 Wiley-Liss, Inc.)

The predominant source of myogenic cells in vertebrates is the dermomyotome (DM). In teleost fish, recent research has provided a useful but limited picture of how myogenic precursors originate from the DM and how they develop into muscle fibers. Here, we combine detailed morphological analysis with examination of molecular markers in trout to describe the cellular mechanisms by which the lateral fast muscle growth zone is created during second phase myogenesis. Results suggest that this occurs by lateral-to-medial immigration of myogenic cells de-epithelializing from the posterior DM lip. These cells then appear to stop proliferation and migrate anteriorly to finally differentiate into muscle fibres. This seems to be a continuation of the rotational cell movement that creates the teleost DM during early somite development. These findings suggest an evolutionary conserved role of the posterior DM lip in amniotes and fish., (Copyright (c) 2008 Wiley-Liss, Inc.)

Vertebrate craniofacial and trunk myogenesis are regulated by distinct genetic programs. Tbx1, homologue of the del22q11.2 syndrome candidate gene TBX1, controls branchiomeric craniofacial muscle development. Here, we demonstrate using immunohistochemistry that myogenic regulatory factors are activated in Tbx1-positive cells within pharyngeal mesoderm. These cells are also Islet1 and Capsulin-positive and in the absence of Tbx1 persist in the core of the first arch. Sporadic hypoplastic mandibular muscles in Tbx1-/- embryos contain Pax7-positive myocytes with indistinguishable differentiation properties from wild-type muscles and have normal tendon attachments and fiber-type patterning. In contrast to TBX1 haploinsufficient del22q11.2 syndrome patients, no alteration in fiber-type distribution was detected in Tbx1+/- adult masseter and pharyngeal constrictor muscles. Furthermore, Tbx1-expressing limb muscles display normal patterning, differentiation, fiber-type growth, fiber-type distribution and fetal maturation in the absence of Tbx1. The critical requirement for Tbx1 during muscle development is thus in the robust onset of myogenic specification in pharyngeal mesoderm., (Copyright (c) 2008 Wiley-Liss, Inc.)

After somitogenesis, skeletal muscle precursors elongate into muscle fibers that anchor to the somite boundary, which becomes the myotome boundary. Fibronectin (Fn) is a major component of the extracellular matrix in both boundaries. Although Fn is required for somitogenesis, effects of Fn disruption on subsequent muscle development are unknown. We show that fn knockdown disrupts myogenesis. Muscle morphogenesis is more disrupted in fn morphants than in a mutant where initial somite boundaries did not form, aei/deltaD. We quantified this disruption using the two-dimensional Wavelet-Transform Modulus Maxima method, which uses the variation of intensity in an image with respect to the direction considered to characterize the structure in a cell lattice. We show that fibers in fn morphants are less organized than in aei/deltaD mutant embryos. Fast- and slow-twitch muscle lengths are also more frequently uncoupled. These data suggest that fn may function to regulate fiber organization and limit fast-twitch muscle fiber length.

Pax3 and Pax7 are closely related paired-boxed family transcription factors that are known to play important roles in embryonic and adult myogenesis. Previous reports describing the expression of Pax3 and Pax7 transcripts reveal expression in many overlapping domains. In this manuscript, we extend these studies by examining the protein expression profiles for Pax3 and Pax7 in developing chick somites and limbs with cellular resolution. Our studies show the existence of distinct subpopulations of cells in the somite and developing limb that are defined by the relative expression levels of Pax3 and Pax7. We also show that Pax3 and Pax7 negatively regulate each other's expression in the dermomyotome, thus providing a possible mechanism for the maintenance of observed expression patterns in the dermomyotome. Further characterization of Pax3- and/or Pax7-positive cells in the dermomyotome and myotome with respect to proliferation and differentiation reveals subpopulations of cells with distinct properties.

Vesicle-associated membrane protein 2 (VAMP2) is a member of the SNARE family of proteins that regulate the intracellular vesicle fusion process. This study investigated the developmental expression of VAMP2 in the rat embryo. In the trunk, VAMP2 was primarily found in the heart on embryonic day (E) 10. On E12.5, VAMP2 expression was found in nerve fibers, somites, and heart. In somites, epithelial cells in the dorsomedial lip, and elongated myoblasts in myotome were positive for VAMP2. On E16.5, VAMP2 was expressed in the heart, nerve fibers, and skeletal muscles. In skeletal muscles, multinuclear myotubes were positive for VAMP2. In the head, where muscles are derived both from somitic and non-somitic origin, VAMP2 was found in myotubes of the extrinsic ocular muscles and masseter muscle on E16.5. These findings suggest the involvement of VAMP2 in the development of skeletal muscles of somitic and non-somitic origins.

Vertebrate somitogenesis is a rhythmically repeated morphogenetic process. The dependence of somitogenesis dynamics on axial position and temperature has not been investigated systematically in any species. Here we use multiple embryo time-lapse imaging to precisely estimate somitogenesis period and somite length under various conditions in the zebrafish embryo. Somites form at a constant period along the trunk, but the period gradually increases in the tail. Somite length varies along the axis in a stereotypical manner, with tail somites decreasing in size. Therefore, our measurements prompt important modifications to the steady-state Clock and Wavefront model: somitogenesis period, somite length, and wavefront velocity all change with axial position. Finally, we show that somitogenesis period changes more than threefold across the standard developmental temperature range, whereas the axial somite length distribution is temperature invariant. This finding indicates that the temperature-induced change in somitogenesis period exactly compensates for altered axial growth., ((c) 2008 Wiley-Liss, Inc.)

Vertebrate developmental biologists typically rely on a limited number of model organisms to understand the evolutionary bases of morphological change. Unfortunately, a typical model system for squamates (lizards and snakes) has not yet been developed leaving many fundamental questions about morphological evolution unaddressed. New model systems would ideally include clades, rather than single species, that are amenable to both laboratory studies of development and field-based analyses of ecology and evolution. Combining an understanding of development with an understanding of ecology and evolution within and between closely related species has the potential to create a seamless understanding of how genetic variation underlies ecologically and evolutionarily relevant variation within populations and between species. Here we briefly introduce a new model system for the integration of development, evolution, and ecology, the lizard genus Anolis, a diverse group of lizards whose ecology and evolution is well understood, and whose genome has recently been sequenced. We present a developmental staging series for Anolis lizards that can act as a baseline for later comparative and experimental studies within this genus., (Copyright (c) 2007 Wiley-Liss, Inc.)

Somites are blocks of mesoderm that form when segment boundaries are periodically generated in the anterior presomitic mesoderm (PSM). Periodicity is thought to be driven by an oscillating Notch-centered segmentation clock, whereas boundaries are spatially positioned by the secreted signaling molecules Wnt3a and Fgf8. We identified the putative transcriptional corepressor Ripply2 as a differentially expressed gene in wild-type and Wnt3a(-/-) embryos. Here, we show that Ripply2 is expressed in the anterior PSM and that it indeed lies downstream of Wnt3a. Dynamic Ripply2 expression in prospective somites S0 and S-I overlaps with the rostral expression of cycling genes in the Notch pathway, suggesting that Ripply2 may be controlled by the segmentation clock. Continued expression of Ripply2 in embryos lacking Hes7, a molecular oscillator in the Notch clock, indicates that Hes7 is not a major regulator of Ripply2. Our data are consistent with Ripply2 functioning as a segment boundary determination gene during mammalian embryogenesis. Developmental, (Published 2007 Wiley-Liss, Inc.)

The three Enigma subfamily proteins, Enigma, Enigma homologue, and Cypher/ZASP belong to the PDZ and LIM encoding protein family, which is characterized by the presence of a PDZ- and one or more LIM domains. PDZ/LIM proteins play important biological roles, and all members have been shown to associate with the actin cytoskeleton. We describe here the splice form specific expression patterns for the three Enigma subfamily members during zebrafish embryogenesis. Whole-mount in situ hybridization revealed common and distinct expression patterns for the different PDZ or LIM domain encoding splice variants. We further studied the role of enigma in zebrafish development. Enigma knockdown appeared to be embryonic lethal shortly after the end of gastrulation and in few surviving embryos led to elongation defects and disorganized somites. In summary, we show here the temporal and spatial expression patterns of the three Enigma family members and their PDZ and LIM domain encoding splice forms during zebrafish embryogenesis. Our results suggest that enigma is important for the formation and organization of somites and might play an important role for actin cytoskeleton organization during development., (Copyright 2007 Wiley-Liss, Inc.)