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Sample records for mesoderm

  1. Efficient ectopic gene expression targeting chick mesoderm.

    PubMed

    Oberg, Kerby C; Pira, Charmaine U; Revelli, Jean-Pierre; Ratz, Beate; Aguilar-Cordova, Estuardo; Eichele, Gregor

    2002-07-01

    The chick model has been instrumental in illuminating genes that regulate early vertebrate development and pattern formation. Targeted ectopic gene expression is critical to dissect further the complicated gene interactions that are involved. In an effort to develop a consistent method to ectopically introduce and focally express genes in chick mesoderm, we evaluated and optimized several gene delivery methods, including implantation of 293 cells laden with viral vectors, direct adenoviral injection, and electroporation (EP). We targeted the mesoderm of chick wing buds between stages 19 and 21 (Hamburger and Hamilton stages) and used beta-galactosidase and green fluorescent protein (GFP) to document gene transfer. Expression constructs using the cytomegalovirus (CMV) promoter, the beta-actin promoter, and vectors with an internal ribosomal entry sequence linked to GFP (IRES-GFP) were also compared. After gene transfer, we monitored expression for up to 3 days. The functionality of ectopic expression was demonstrated with constructs containing the coding sequences for Shh, a secreted signaling protein, or Hoxb-8, a transcription factor, both of which can induce digit duplication when ectopically expressed in anterior limb mesoderm. We identified several factors that enhance mesodermal gene transfer. First, the use of a vector with the beta-actin promoter coupled to the 69% fragment of the bovine papilloma virus yielded superior mesodermal expression both by markers and functional results when compared with several CMV-driven vectors. Second, we found the use of mineral oil to be an important adjuvant for EP and direct viral injection to localize and contain vector within the mesoderm at the injection site. Lastly, although ectopic expression could be achieved with all three methods, we favored EP confined to the mesoderm with insulated microelectrodes (confined microelectroporation- CMEP), because vector construction is rapid, the method is efficient, and results

  2. Evolutionary conservation of early mesoderm specification by mechanotransduction in Bilateria

    PubMed Central

    Brunet, Thibaut; Bouclet, Adrien; Ahmadi, Padra; Mitrossilis, Démosthène; Driquez, Benjamin; Brunet, Anne-Christine; Henry, Laurent; Serman, Fanny; Béalle, Gaëlle; Ménager, Christine; Dumas-Bouchiat, Frédéric; Givord, Dominique; Yanicostas, Constantin; Le-Roy, Damien; Dempsey, Nora M.; Plessis, Anne; Farge, Emmanuel

    2013-01-01

    The modulation of developmental biochemical pathways by mechanical cues is an emerging feature of animal development, but its evolutionary origins have not been explored. Here we show that a common mechanosensitive pathway involving β-catenin specifies early mesodermal identity at gastrulation in zebrafish and Drosophila. Mechanical strains developed by zebrafish epiboly and Drosophila mesoderm invagination trigger the phosphorylation of β-catenin–tyrosine-667. This leads to the release of β-catenin into the cytoplasm and nucleus, where it triggers and maintains, respectively, the expression of zebrafish brachyury orthologue notail and of Drosophila Twist, both crucial transcription factors for early mesoderm identity. The role of the β-catenin mechanosensitive pathway in mesoderm identity has been conserved over the large evolutionary distance separating zebrafish and Drosophila. This suggests mesoderm mechanical induction dating back to at least the last bilaterian common ancestor more than 570 million years ago, the period during which mesoderm is thought to have emerged. PMID:24281726

  3. The origin of mesoderm in phoronids

    NASA Technical Reports Server (NTRS)

    Freeman, Gary; Martindale, Mark Q.

    2002-01-01

    Descriptive studies of phoronid development have concluded that the mesoderm of these animals originates from the endoderm during gastrulation. This interpretation has been tested by labeling one blastomere of 4- through 16-cell embryos and examining the position and germ layers occupied by the labeled clones of cells in the larva. No 2 injections gave rise to identical clones of cells, suggesting that the cleavage program does not generate cells of unique identity and that cell fates are established at later developmental time points. In many cases, a relatively large sector composed of ectodermal cells was labeled. When these labeled cells were adjacent to the mouth or anus of the larva, muscle and mesenchyme cells originated from the labeled clones. Under these circumstances, nerve cells also originated from these labeled sectors. These labeling studies also showed that endodermal cells can give rise to mesodermal and neural cells. These results suggest that nerve and muscle cells are induced to form at ectodermal-endodermal boundaries from both germ layers. These marking experiments also confirmed the observation that nerve cells originate both from the apical organ and the trunk region and show for the first time that the intestine originates by ingression of posterior ectoderm.

  4. Mesoderm layer formation in Xenopus and Drosophila gastrulation

    NASA Astrophysics Data System (ADS)

    Winklbauer, Rudolf; Müller, H.-Arno J.

    2011-08-01

    During gastrulation, the mesoderm spreads out between ectoderm and endoderm to form a mesenchymal cell layer. Surprisingly the underlying principles of mesoderm layer formation are very similar in evolutionarily distant species like the fruit fly, Drosophila melanogaster, and the frog, Xenopus laevis, in which the molecular and the cellular basis of mesoderm layer formation have been extensively studied. Complementary expression of growth factors in the ectoderm and their receptors in the mesoderm act to orient cellular protrusive activities and direct cell movement, leading to radial cell intercalation and the spreading of the mesoderm layer. This mechanism is contrasted with generic physical mechanisms of tissue spreading that consider the adhesive and physical properties of the cells and tissues. Both mechanisms need to be integrated to orchestrate mesenchymal morphogenesis.

  5. Hedgehog signaling patterns mesoderm in the sea urchin

    PubMed Central

    Walton, Katherine D.; Warner, Jacob; Hertzler, Philip H.; McClay, David R.

    2009-01-01

    The Hedgehog (Hh) signaling pathway is essential for patterning many structures in vertebrates including the nervous system, chordamesoderm, limb and endodermal organs. In the sea urchin, a basal deuterostome, Hh signaling is shown to participate in organizing the mesoderm. At gastrulation the Hh ligand is expressed by the endoderm downstream of the Brachyury and FoxA transcription factors in the endomesoderm gene regulatory network. The co-receptors Patched (Ptc) and Smoothened (Smo) are expressed by the neighboring skeletogenic and non-skeletogenic mesoderm. Perturbations of Hh, Ptc and Smo cause embryos to develop with skeletal defects and inappropriate non-skeletogenic mesoderm patterning, although initial specification of mesoderm occurs without detectable abnormalities. Perturbations of the pathway caused late defects in skeletogenesis and in the non-skeletogenic mesoderm, including altered numbers of pigment and blastocoelar cells, randomized left-right asymmetry of coelomic pouches, and disorganized circumesophageal muscle causing an inability to swallow. Together the data support the requirement of Hh signaling in patterning each of the mesoderm subtypes in the sea urchin embryo. PMID:19393640

  6. Genetic Tagging During Human Mesoderm Differentiation Reveals Tripotent Lateral Plate Mesodermal Progenitors.

    PubMed

    Chin, Chee Jia; Cooper, Aaron R; Lill, Georgia R; Evseenko, Denis; Zhu, Yuhua; He, Chong Bin; Casero, David; Pellegrini, Matteo; Kohn, Donald B; Crooks, Gay M

    2016-05-01

    Although clonal studies of lineage potential have been extensively applied to organ specific stem and progenitor cells, much less is known about the clonal origins of lineages formed from the germ layers in early embryogenesis. We applied lentiviral tagging followed by vector integration site analysis (VISA) with high-throughput sequencing to investigate the ontogeny of the hematopoietic, endothelial and mesenchymal lineages as they emerge from human embryonic mesoderm. In contrast to studies that have used VISA to track differentiation of self-renewing stem cell clones that amplify significantly over time, we focused on a population of progenitor clones with limited self-renewal capability. Our analyses uncovered the critical influence of sampling on the interpretation of lentiviral tag sharing, particularly among complex populations with minimal clonal duplication. By applying a quantitative framework to estimate the degree of undersampling we revealed the existence of tripotent mesodermal progenitors derived from pluripotent stem cells, and the subsequent bifurcation of their differentiation into bipotent endothelial/hematopoietic or endothelial/mesenchymal progenitors. Stem Cells 2016;34:1239-1250. PMID:26934332

  7. Brg1 modulates enhancer activation in mesoderm lineage commitment

    DOE PAGESBeta

    Alexander, Jeffrey M.; Hota, Swetansu K.; He, Daniel; Thomas, Sean; Ho, Lena; Pennacchio, Len A.; Bruneau, B. G.

    2015-03-26

    The interplay between different levels of gene regulation in modulating developmental transcriptional programs, such as histone modifications and chromatin remodeling, is not well understood. Here, we show that the chromatin remodeling factor Brg1 is required for enhancer activation in mesoderm induction. In an embryonic stem cell-based directed differentiation assay, the absence of Brg1 results in a failure of cardiomyocyte differentiation and broad deregulation of lineage-specific gene expression during mesoderm induction. We find that Brg1 co-localizes with H3K27ac at distal enhancers and is required for robust H3K27 acetylation at distal enhancers that are activated during mesoderm induction. Brg1 is also requiredmore » to maintain Polycomb-mediated repression of non-mesodermal developmental regulators, suggesting cooperativity between Brg1 and Polycomb complexes. Thus, Brg1 is essential for modulating active and repressive chromatin states during mesoderm lineage commitment, in particular the activation of developmentally important enhancers. In conclusion, these findings demonstrate interplay between chromatin remodeling complexes and histone modifications that, together, ensure robust and broad gene regulation during crucial lineage commitment decisions.« less

  8. Brg1 modulates enhancer activation in mesoderm lineage commitment

    SciTech Connect

    Alexander, Jeffrey M.; Hota, Swetansu K.; He, Daniel; Thomas, Sean; Ho, Lena; Pennacchio, Len A.; Bruneau, B. G.

    2015-03-26

    The interplay between different levels of gene regulation in modulating developmental transcriptional programs, such as histone modifications and chromatin remodeling, is not well understood. Here, we show that the chromatin remodeling factor Brg1 is required for enhancer activation in mesoderm induction. In an embryonic stem cell-based directed differentiation assay, the absence of Brg1 results in a failure of cardiomyocyte differentiation and broad deregulation of lineage-specific gene expression during mesoderm induction. We find that Brg1 co-localizes with H3K27ac at distal enhancers and is required for robust H3K27 acetylation at distal enhancers that are activated during mesoderm induction. Brg1 is also required to maintain Polycomb-mediated repression of non-mesodermal developmental regulators, suggesting cooperativity between Brg1 and Polycomb complexes. Thus, Brg1 is essential for modulating active and repressive chromatin states during mesoderm lineage commitment, in particular the activation of developmentally important enhancers. In conclusion, these findings demonstrate interplay between chromatin remodeling complexes and histone modifications that, together, ensure robust and broad gene regulation during crucial lineage commitment decisions.

  9. Mesodermal expression of Moz is necessary for cardiac septum development.

    PubMed

    Vanyai, Hannah K; Thomas, Tim; Voss, Anne K

    2015-07-01

    Ventricular septal defects (VSDs) are the most commonly occurring congenital heart defect. They are regularly associated with complex syndromes, including DiGeorge syndrome and Holt-Oram syndrome, which are characterised by haploinsufficiency for the T-box transcription factors TBX1 and TBX5, respectively. The histone acetyltransferase monocytic leukaemia zinc finger protein, MOZ (MYST3/KAT6A), is required for the expression of the Tbx1 and Tbx5 genes. Homozygous loss of MOZ results in DiGeorge syndrome-like defects including VSD. The Moz gene is expressed in the ectodermal, mesodermal and endodermal aspects of the developing pharyngeal apparatus and heart; however it is unclear in which of these tissues MOZ is required for heart development. The role of MOZ in the activation of Tbx1 would suggest a requirement for MOZ in the mesoderm, because deletion of Tbx1 in the mesoderm causes VSDs. Here, we investigated the tissue-specific requirements for MOZ in the mesoderm. We demonstrate that Mesp1-cre-mediated deletion of Moz results in high penetrance of VSDs and overriding aorta and a significant decrease in MOZ-dependant Tbx1 and Tbx5 expression. Together, our data suggest that the molecular pathogenesis of VSDs in Moz germline mutant mice is due to loss of MOZ-dependant activation of mesodermal Tbx1 and Tbx5 expression. PMID:25912687

  10. Mesoderm, Cooked Up Fast and Served to Order.

    PubMed

    Kyba, Michael

    2016-08-01

    New sequencing technologies have made it possible to interrogate with unprecedented depth the intrinsic changes experienced by cells as they transit the arena of development. Recently in Cell, Loh, Chen, and colleagues investigated early lineage-restricted human mesoderm cell types and their precursors going back to pluripotency (Loh et al., 2016). PMID:27494669

  11. Gut endoderm takes flight from the wings of mesoderm.

    PubMed

    McDonald, Angela C H; Rossant, Janet

    2014-12-01

    The endoderm layer destined to be primitive gut is a mosaic of earlier visceral endoderm and definitive endoderm that arises later, during gastrulation. Live imaging now reveals that in mouse embryos, definitive endoderm cells egress from underlying mesoderm and intercalate into the overlying cell layer. This process requires SOX17-mediated control of basement membrane organization. PMID:25434463

  12. Trunk exoskeleton in teleosts is mesodermal in origin

    PubMed Central

    Shimada, Atsuko; Kawanishi, Toru; Kaneko, Takuya; Yoshihara, Hiroki; Yano, Tohru; Inohaya, Keiji; Kinoshita, Masato; Kamei, Yasuhiro; Tamura, Koji; Takeda, Hiroyuki

    2013-01-01

    The vertebrate mineralized skeleton is known to have first emerged as an exoskeleton that extensively covered the fossil jawless fish. The evolutionary origin of this exoskeleton has long been attributed to the emergence of the neural crest, but experimental evaluation for this is still poor. Here we determine the embryonic origin of scales and fin rays of medaka (teleost trunk exoskeletons) by applying long-term cell labelling methods, and demonstrate that both tissues are mesodermal in origin. Neural crest cells, however, fail to contribute to these tissues. This result suggests that the trunk neural crest has no skeletogenic capability in fish, instead highlighting the dominant role of the mesoderm in the evolution of the trunk skeleton. This further implies that the role of the neural crest in skeletogenesis has been predominant in the cephalic region from the early stage of vertebrate evolution. PMID:23535660

  13. Tbx16 regulates hox gene activation in mesodermal progenitor cells.

    PubMed

    Payumo, Alexander Y; McQuade, Lindsey E; Walker, Whitney J; Yamazoe, Sayumi; Chen, James K

    2016-09-01

    The transcription factor T-box 16 (Tbx16, or Spadetail) is an essential regulator of paraxial mesoderm development in zebrafish (Danio rerio). Mesodermal progenitor cells (MPCs) fail to differentiate into trunk somites in tbx16 mutants and instead accumulate within the tailbud in an immature state. However, the mechanisms by which Tbx16 controls mesoderm patterning have remained enigmatic. We describe here the use of photoactivatable morpholino oligonucleotides to determine the Tbx16 transcriptome in MPCs. We identified 124 Tbx16-regulated genes that were expressed in zebrafish gastrulae, including several developmental signaling proteins and regulators of gastrulation, myogenesis and somitogenesis. Unexpectedly, we observed that a loss of Tbx16 function precociously activated posterior hox genes in MPCs, and overexpression of a single posterior hox gene was sufficient to disrupt MPC migration. Our studies support a model in which Tbx16 regulates the timing of collinear hox gene activation to coordinate the anterior-posterior fates and positions of paraxial MPCs. PMID:27376691

  14. Mesoderm patterning and morphogenesis in the polychaete Alitta virens (Spiralia, Annelida): Expression of mesodermal markers Twist, Mox, Evx and functional role for MAP kinase signaling.

    PubMed

    Kozin, Vitaly V; Filimonova, Daria A; Kupriashova, Ekaterina E; Kostyuchenko, Roman P

    2016-05-01

    Mesoderm represents the evolutionary youngest germ layer and forms numerous novel tissues in bilaterian animals. Despite the established conservation of the gene regulatory networks that drive mesoderm differentiation (e.g. myogenesis), mechanisms of mesoderm specification are highly variable in distant model species. Thus, broader phylogenetic sampling is required to reveal common features of mesoderm formation across bilaterians. Here we focus on a representative of Spiralia, the marine annelid Alitta virens, whose mesoderm development is still poorly investigated on the molecular level. We characterize three novel early mesodermal markers for A. virens - Twist, Mox, and Evx - which are differentially expressed within the mesodermal lineages. The Twist mRNA is ubiquitously distributed in the fertilized egg and exhibits specific expression in endomesodermal- and ectomesodermal-founder cells at gastrulation. Twist is expressed around the blastopore and later in a segmental metameric pattern. We consider this expression to be ancestral, and in support of the enterocoelic hypothesis of mesoderm evolution. We also revealed an early pattern of the MAPK activation in A. virens that is different from the previously reported pattern in spiralians. Inhibition of the MAPK pathway by U0126 disrupts the metameric Twist and Mox expression, indicating an early requirement of the MAPK cascade for proper morphogenesis of endomesodermal tissues. PMID:27000638

  15. Heterogeneity of Mesp1+ mesoderm revealed by single-cell RNA-seq.

    PubMed

    Chan, Sunny Sun-Kin; Chan, Howe H W; Kyba, Michael

    2016-06-01

    Mesp1 is a transcription factor that promotes differentiation of pluripotent cells into different mesoderm lineages including hematopoietic, cardiac and skeletal myogenic. This occurs via at least two transient cell populations: a common hematopoietic/cardiac progenitor population and a common cardiac/skeletal myogenic progenitor population. It is not established whether Mesp1-induced mesoderm cells are intrinsically heterogeneous, or are simply capable of multiple lineage decisions. In the current study, we applied single-cell RNA-seq to analyze Mesp1+ mesoderm. Initial whole transcriptome analysis showed a surprising homogeneity among Mesp1-induced mesoderm cells. However, this apparent global homogeneity masked an intrinsic heterogeneity revealed by interrogating a panel of early mesoderm patterning factors. This approach enabled discovery of subpopulations primed for hematopoietic or cardiac development. These studies demonstrate the heterogeneic nature of Mesp1+ mesoderm. PMID:27131741

  16. The role of FGF signaling in establishment and maintenance of mesodermal gene expression in Xenopus

    PubMed Central

    Fletcher, Russell B.; Harland, Richard M.

    2010-01-01

    FGF signaling is important for the formation of mesoderm in vertebrates, and when it is perturbed in Xenopus, most trunk and tail mesoderm fails to form. Here we have further dissected the activities of FGF in patterning the embryo by addressing its inductive and maintenance roles. We show that FGF signaling is necessary for the establishment of xbra expression in addition to its well-characterized role in maintaining xbra expression. The role of FGF signaling in organizer formation is not clear in Xenopus. We find that FGF signaling is essential for the initial specification of paraxial mesoderm but not for activation of several pan-mesodermal and most organizer genes; however, early FGF signaling is necessary for the maintenance of organizer gene expression into the neurula stage. Inhibition of FGF signaling prevents VegT activation of specific mesodermal transcripts. These findings illuminate how FGF signaling contributes to the establishment of distinct types of mesoderm. PMID:18386826

  17. Factors that coordinate mesoderm specification from neuromesodermal progenitors with segmentation during vertebrate axial extension.

    PubMed

    Martin, Benjamin L

    2016-01-01

    The formation of the vertebrate body depends on the precise timing and coordination of molecular and morphological events. During vertebrate embryogenesis, the paraxial mesoderm is segmented into structures called somites in a progressive fashion from the anterior to the posterior at the same time as the entire body axis elongates in the posterior direction. Evidence from several vertebrate species indicates that new paraxial mesoderm is continuously induced from neuromesodermal progenitors at the posterior-most end of the embryo. The newly forming mesoderm exists in a specialized environment called the mesodermal progenitor niche. This review will discuss how the progenitor niche coordinates the continuous addition of new mesoderm to the body axis with proper segmentation of this mesoderm upon exit from the niche. I will focus on evidence that the t-box transcription factor Brachyury and its downstream transcriptional targets serve as the primary factors coordinating mesoderm specification with somitogenesis. I will end with a discussion of recent exciting work regarding the cell-cycle and migratory behavior of mesodermal cells as they exit the progenitor niche, which may serve to further integrate new mesoderm production with proper segmentation. PMID:26658097

  18. Resolving early mesoderm diversification through single-cell expression profiling.

    PubMed

    Scialdone, Antonio; Tanaka, Yosuke; Jawaid, Wajid; Moignard, Victoria; Wilson, Nicola K; Macaulay, Iain C; Marioni, John C; Göttgens, Berthold

    2016-07-14

    In mammals, specification of the three major germ layers occurs during gastrulation, when cells ingressing through the primitive streak differentiate into the precursor cells of major organ systems. However, the molecular mechanisms underlying this process remain unclear, as numbers of gastrulating cells are very limited. In the mouse embryo at embryonic day 6.5, cells located at the junction between the extra-embryonic region and the epiblast on the posterior side of the embryo undergo an epithelial-to-mesenchymal transition and ingress through the primitive streak. Subsequently, cells migrate, either surrounding the prospective ectoderm contributing to the embryo proper, or into the extra-embryonic region to form the yolk sac, umbilical cord and placenta. Fate mapping has shown that mature tissues such as blood and heart originate from specific regions of the pre-gastrula epiblast, but the plasticity of cells within the embryo and the function of key cell-type-specific transcription factors remain unclear. Here we analyse 1,205 cells from the epiblast and nascent Flk1(+) mesoderm of gastrulating mouse embryos using single-cell RNA sequencing, representing the first transcriptome-wide in vivo view of early mesoderm formation during mammalian gastrulation. Additionally, using knockout mice, we study the function of Tal1, a key haematopoietic transcription factor, and demonstrate, contrary to previous studies performed using retrospective assays, that Tal1 knockout does not immediately bias precursor cells towards a cardiac fate. PMID:27383781

  19. Case report of a 22-week fetus with 47,XXX karyotype and multiple lower mesodermal defects.

    PubMed

    Hoang, M P; Wilson, K S; Schneider, N R; Timmons, C F

    1999-01-01

    A 22-week stillborn fetus with 47,XXX karyotype had lower mesodermal defects consisting of irregular fusion of the sacral vertebrae, anal agenesis, multicystic dysplasia of a horseshoe kidney, a single umbilical artery, dysplastic ovaries, and uterine hypoplasia. This case provides additional evidence for an association between trisomy X and genitourinary defects including lower mesodermal defects sequence. PMID:9841707

  20. Expression of the prospective mesoderm genes twist, snail, and mef2 in penaeid shrimp.

    PubMed

    Wei, Jiankai; Glaves, Richard Samuel Elliot; Sellars, Melony J; Xiang, Jianhai; Hertzler, Philip L

    2016-07-01

    In penaeid shrimp, mesoderm forms from two sources: naupliar mesoderm founder cells, which invaginate during gastrulation, and posterior mesodermal stem cells called mesoteloblasts, which undergo characteristic teloblastic divisions. The primordial mesoteloblast descends from the ventral mesendoblast, which arrests in cell division at the 32-cell stage and ingresses with its sister dorsal mesendoblast prior to naupliar mesoderm invagination. The naupliar mesoderm forms the muscles of the naupliar appendages (first and second antennae and mandibles), while the mesoteloblasts form the mesoderm, including the muscles, of subsequently formed posterior segments. To better understand the mechanism of mesoderm and muscle formation in penaeid shrimp, twist, snail, and mef2 cDNAs were identified from transcriptomes of Penaeus vannamei, P. japonicus, P. chinensis, and P. monodon. A single Twist ortholog was found, with strong inferred amino acid conservation across all three species. Multiple Snail protein variants were detected, which clustered in a phylogenetic tree with other decapod crustacean Snail sequences. Two closely-related mef2 variants were found in P. vannamei. The developmental mRNA expression of these genes was studied by qPCR in P. vannamei embryos, larvae, and postlarvae. Expression of Pv-twist and Pv-snail began during the limb bud stage and continued through larval stages to the postlarva. Surprisingly, Pv-mef2 expression was found in all stages from the zygote to the postlarva, with the highest expression in the limb bud and protozoeal stages. The results add comparative data on the development of anterior and posterior mesoderm in malacostracan crustaceans, and should stimulate further studies on mesoderm and muscle development in penaeid shrimp. PMID:27129985

  1. Wnt signaling and tbx16 form a bistable switch to commit bipotential progenitors to mesoderm

    PubMed Central

    Bouldin, Cortney M.; Manning, Alyssa J.; Peng, Yu-Hsuan; Farr, Gist H.; Hung, King L.; Dong, Alice; Kimelman, David

    2015-01-01

    Anterior to posterior growth of the vertebrate body is fueled by a posteriorly located population of bipotential neuro-mesodermal progenitor cells. These progenitors have a limited rate of proliferation and their maintenance is crucial for completion of the anterior-posterior axis. How they leave the progenitor state and commit to differentiation is largely unknown, in part because widespread modulation of factors essential for this process causes organism-wide effects. Using a novel assay, we show that zebrafish Tbx16 (Spadetail) is capable of advancing mesodermal differentiation cell-autonomously. Tbx16 locks cells into the mesodermal state by not only activating downstream mesodermal genes, but also by repressing bipotential progenitor genes, in part through a direct repression of sox2. We demonstrate that tbx16 is activated as cells move from an intermediate Wnt environment to a high Wnt environment, and show that Wnt signaling activates the tbx16 promoter. Importantly, high-level Wnt signaling is able to accelerate mesodermal differentiation cell-autonomously, just as we observe with Tbx16. Finally, because our assay for mesodermal commitment is quantitative we are able to show that the acceleration of mesodermal differentiation is surprisingly incomplete, implicating a potential separation of cell movement and differentiation during this process. Together, our data suggest a model in which high levels of Wnt signaling induce a transition to mesoderm by directly activating tbx16, which in turn acts to irreversibly flip a bistable switch, leading to maintenance of the mesodermal fate and repression of the bipotential progenitor state, even as cells leave the initial high-Wnt environment. PMID:26062939

  2. The distribution of Dishevelled in convergently extending mesoderm.

    PubMed

    Panousopoulou, Eleni; Tyson, Richard A; Bretschneider, Till; Green, Jeremy B A

    2013-10-15

    Convergent extension (CE) is a conserved morphogenetic movement that drives axial lengthening of the primary body axis and depends on the planar cell polarity (PCP) pathway. In Drosophila epithelia, a polarised subcellular accumulation of PCP core components, such as Dishevelled (Dvl) protein, is associated with PCP function. Dvl has long been thought to accumulate in the mediolateral protrusions in Xenopus chordamesoderm cells undergoing CE. Here we present a quantitative analysis of Dvl intracellular localisation in Xenopus chordamesoderm cells. We find that, surprisingly, accumulations previously observed at mediolateral protrusions of chordamesodermal cells are not protrusion-specific but reflect yolk-free cytoplasm and are quantitatively matched by the distribution of the cytoplasm-filling lineage marker dextran. However, separating cell cortex-associated from bulk Dvl signal reveals a statistical enrichment of Dvl in notochord-somite boundary-(NSB)-directed protrusions, which is dependent upon NSB proximity. Dvl puncta were also observed, but only upon elevated overexpression. These puncta showed no statistically significant spatial bias, in contrast to the strongly posteriorly-enriched GFP-Dvl puncta previously reported in zebrafish. We propose that Dvl distribution is more subtle and dynamic than previously appreciated and that in vertebrate mesoderm it reflects processes other than protrusion as such. PMID:23876427

  3. Pharyngeal mesoderm regulatory network controls cardiac and head muscle morphogenesis

    PubMed Central

    Harel, Itamar; Maezawa, Yoshiro; Avraham, Roi; Rinon, Ariel; Ma, Hsiao-Yen; Cross, Joe W.; Leviatan, Noam; Hegesh, Julius; Roy, Achira; Jacob-Hirsch, Jasmine; Rechavi, Gideon; Carvajal, Jaime; Tole, Shubha; Kioussi, Chrissa; Quaggin, Susan; Tzahor, Eldad

    2012-01-01

    The search for developmental mechanisms driving vertebrate organogenesis has paved the way toward a deeper understanding of birth defects. During embryogenesis, parts of the heart and craniofacial muscles arise from pharyngeal mesoderm (PM) progenitors. Here, we reveal a hierarchical regulatory network of a set of transcription factors expressed in the PM that initiates heart and craniofacial organogenesis. Genetic perturbation of this network in mice resulted in heart and craniofacial muscle defects, revealing robust cross-regulation between its members. We identified Lhx2 as a previously undescribed player during cardiac and pharyngeal muscle development. Lhx2 and Tcf21 genetically interact with Tbx1, the major determinant in the etiology of DiGeorge/velo-cardio-facial/22q11.2 deletion syndrome. Furthermore, knockout of these genes in the mouse recapitulates specific cardiac features of this syndrome. We suggest that PM-derived cardiogenesis and myogenesis are network properties rather than properties specific to individual PM members. These findings shed new light on the developmental underpinnings of congenital defects. PMID:23112163

  4. Generation of an expandable intermediate mesoderm restricted progenitor cell line from human pluripotent stem cells.

    PubMed

    Kumar, Nathan; Richter, Jenna; Cutts, Josh; Bush, Kevin T; Trujillo, Cleber; Nigam, Sanjay K; Gaasterland, Terry; Brafman, David; Willert, Karl

    2015-01-01

    The field of tissue engineering entered a new era with the development of human pluripotent stem cells (hPSCs), which are capable of unlimited expansion whilst retaining the potential to differentiate into all mature cell populations. However, these cells harbor significant risks, including tumor formation upon transplantation. One way to mitigate this risk is to develop expandable progenitor cell populations with restricted differentiation potential. Here, we used a cellular microarray technology to identify a defined and optimized culture condition that supports the derivation and propagation of a cell population with mesodermal properties. This cell population, referred to as intermediate mesodermal progenitor (IMP) cells, is capable of unlimited expansion, lacks tumor formation potential, and, upon appropriate stimulation, readily acquires properties of a sub-population of kidney cells. Interestingly, IMP cells fail to differentiate into other mesodermally-derived tissues, including blood and heart, suggesting that these cells are restricted to an intermediate mesodermal fate. PMID:26554899

  5. Mapping the Pairwise Choices Leading from Pluripotency to Human Bone, Heart, and Other Mesoderm Cell Types.

    PubMed

    Loh, Kyle M; Chen, Angela; Koh, Pang Wei; Deng, Tianda Z; Sinha, Rahul; Tsai, Jonathan M; Barkal, Amira A; Shen, Kimberle Y; Jain, Rajan; Morganti, Rachel M; Shyh-Chang, Ng; Fernhoff, Nathaniel B; George, Benson M; Wernig, Gerlinde; Salomon, Rachel E A; Chen, Zhenghao; Vogel, Hannes; Epstein, Jonathan A; Kundaje, Anshul; Talbot, William S; Beachy, Philip A; Ang, Lay Teng; Weissman, Irving L

    2016-07-14

    Stem-cell differentiation to desired lineages requires navigating alternating developmental paths that often lead to unwanted cell types. Hence, comprehensive developmental roadmaps are crucial to channel stem-cell differentiation toward desired fates. To this end, here, we map bifurcating lineage choices leading from pluripotency to 12 human mesodermal lineages, including bone, muscle, and heart. We defined the extrinsic signals controlling each binary lineage decision, enabling us to logically block differentiation toward unwanted fates and rapidly steer pluripotent stem cells toward 80%-99% pure human mesodermal lineages at most branchpoints. This strategy enabled the generation of human bone and heart progenitors that could engraft in respective in vivo models. Mapping stepwise chromatin and single-cell gene expression changes in mesoderm development uncovered somite segmentation, a previously unobservable human embryonic event transiently marked by HOPX expression. Collectively, this roadmap enables navigation of mesodermal development to produce transplantable human tissue progenitors and uncover developmental processes. VIDEO ABSTRACT. PMID:27419872

  6. Ectopic Expression Screen Identifies Genes Affecting Drosophila Mesoderm Development Including the HSPG Trol

    PubMed Central

    Trisnadi, Nathanie; Stathopoulos, Angelike

    2014-01-01

    Gastrulation of the embryo involves coordinate cell movements likely supported by multiple signaling pathways, adhesion molecules, and extracellular matrix components. Fibroblast growth factors (FGFs) have a major role in Drosophila melanogaster mesoderm migration; however, few other inputs are known and the mechanism supporting cell movement is unclear. To provide insight, we performed an ectopic expression screen to identify secreted or membrane-associated molecules that act to support mesoderm migration. Twenty-four UAS insertions were identified that cause lethality when expressed in either the mesoderm (Twi-Gal4) or the ectoderm (69B-Gal4). The list was narrowed to a subset of 10 genes that were shown to exhibit loss-of-function mutant phenotypes specifically affecting mesoderm migration. These include the FGF ligand Pyramus, α-integrins, E-cadherin, Cueball, EGFR, JAK/STAT signaling components, as well as the heparan sulfate proteoglycan (HSPG) Terribly reduced optic lobes (Trol). Trol encodes the ortholog of mammalian HSPG Perlecan, a demonstrated FGF signaling cofactor. Here, we examine the role of Trol in Drosophila mesoderm migration and compare and contrast its role with that of Syndecan (Sdc), another HSPG previously implicated in this process. Embryos mutant for Trol or Sdc were obtained and analyzed. Our data support the view that both HSPGs function to support FGF-dependent processes in the early embryo as they share phenotypes with FGF mutants: Trol in terms of effects on mesoderm migration and caudal visceral mesoderm (CVM) migration and Sdc in terms of dorsal mesoderm specification. The differential roles uncovered for these two HSPGs suggest that HSPG cofactor choice may modify FGF-signaling outputs. PMID:25538103

  7. Neural crest induction by paraxial mesoderm in Xenopus embryos requires FGF signals.

    PubMed

    Monsoro-Burq, Anne-Hélène; Fletcher, Russell B; Harland, Richard M

    2003-07-01

    At the border of the neural plate, the induction of the neural crest can be achieved by interactions with the epidermis, or with the underlying mesoderm. Wnt signals are required for the inducing activity of the epidermis in chick and amphibian embryos. Here, we analyze the molecular mechanisms of neural crest induction by the mesoderm in Xenopus embryos. Using a recombination assay, we show that prospective paraxial mesoderm induces a panel of neural crest markers (Slug, FoxD3, Zic5 and Sox9), whereas the future axial mesoderm only induces a subset of these genes. This induction is blocked by a dominant negative (dn) form of FGFR1. However, neither dnFGFR4a nor inhibition of Wnt signaling prevents neural crest induction in this system. Among the FGFs, FGF8 is strongly expressed by the paraxial mesoderm. FGF8 is sufficient to induce the neural crest markers FoxD3, Sox9 and Zic5 transiently in the animal cap assay. In vivo, FGF8 injections also expand the Slug expression domain. This suggests that FGF8 can initiate neural crest formation and cooperates with other DLMZ-derived factors to maintain and complete neural crest induction. In contrast to Wnts, eFGF or bFGF, FGF8 elicits neural crest induction in the absence of mesoderm induction and without a requirement for BMP antagonists. In vivo, it is difficult to dissociate the roles of FGF and WNT factors in mesoderm induction and neural patterning. We show that, in most cases, effects on neural crest formation were parallel to altered mesoderm or neural development. However, neural and neural crest patterning can be dissociated experimentally using different dominant-negative manipulations: while Nfz8 blocks both posterior neural plate formation and neural crest formation, dnFGFR4a blocks neural patterning without blocking neural crest formation. These results suggest that different signal transduction mechanisms may be used in neural crest induction, and anteroposterior neural patterning. PMID:12783784

  8. Cripto is required for mesoderm and endoderm cell allocation during mouse gastrulation

    PubMed Central

    Jin, Jiu-Zhen; Ding, Jixiang

    2013-01-01

    During mouse gastrulation, cells in the primitive streak undergo epithelial-mesenchymal transformation and the resulting mesenchymal cells migrate out laterally to form mesoderm and definitive endoderm across the entire embryonic cylinder. The mechanisms underlying mesoderm and endoderm specification, migration, and allocation is poorly understood. In this study, we focused on the function of mouse Cripto, a member of the EGF-CFC gene family that is highly expressed in the primitive streak and migrating mesoderm cells on embryonic day 6.5. Conditional inactivation of Cripto during gastrulation leads to varied defects in mesoderm and endoderm development. Mutant embryos display accumulation of mesenchymal cells around the shortened primitive streak indicating a functional requirement of Cripto during the formation of mesoderm layer in gastrulation. In addition, some mutant embryos showed poor formation and abnormal allocation of definitive endoderm cells on embryonic day 7.5. Consistently, many mutant embryos that survived to embryonic day 8.5 displayed defects in ventral closure of the gut endoderm causing cardia bifida. Detailed analyses revealed that both the Fgf8-Fgfr1 pathway and p38 MAP kinase activation are partially affected by the loss of Cripto function. These results demonstrate a critical role for Cripto during mouse gastrulation, especially in mesoderm and endoderm formation and allocation. PMID:23747598

  9. Mesodermal expression of integrin α5β1 regulates neural crest development and cardiovascular morphogenesis

    PubMed Central

    Liang, Dong; Wang, Xia; Mittal, Ashok; Dhiman, Sonam; Hou, Shuan-Yu; Degenhardt, Karl; Astrof, Sophie

    2014-01-01

    Integrin α5-null embryos die in mid-gestation from severe defects in cardiovascular morphogenesis, which stem from defective development of the neural crest, heart and vasculature. To investigate the role of integrin α5β1 in cardiovascular development, we used the Mesp1Cre knock-in strain of mice to ablate integrin α5 in the anterior mesoderm, which gives rise to all of the cardiac and many of the vascular and muscle lineages in the anterior portion of the embryo. Surprisingly, we found that mutant embryos displayed numerous defects related to the abnormal development of the neural crest such as cleft palate, ventricular septal defect, abnormal development of hypoglossal nerves, and defective remodeling of the aortic arch arteries. We found that defects in arch artery remodeling stem from the role of mesodermal integrin α5β1 in neural crest proliferation and differentiation into vascular smooth muscle cells, while proliferation of pharyngeal mesoderm and differentiation of mesodermal derivatives into vascular smooth muscle cells was not defective. Taken together our studies demonstrate a requisite role for mesodermal integrin α5β1 in signaling between the mesoderm and the neural crest, thereby regulating neural crest-dependent morphogenesis of essential embryonic structures. PMID:25242040

  10. biniou (FoxF), a central component in a regulatory network controlling visceral mesoderm development and midgut morphogenesis in Drosophila

    PubMed Central

    Zaffran, Stephane; Küchler, Axel; Lee, Hsiu-Hsiang; Frasch, Manfred

    2001-01-01

    The subdivision of the lateral mesoderm into a visceral (splanchnic) and a somatic layer is a crucial event during early mesoderm development in both arthropod and vertebrate embryos. In Drosophila, this subdivision leads to the differential development of gut musculature versus body wall musculature. Here we report that biniou, the sole Drosophila representative of the FoxF subfamily of forkhead domain genes, has a key role in the development of the visceral mesoderm and the derived gut musculature. biniou expression is activated in the trunk visceral mesoderm primordia downstream of dpp, tinman, and bagpipe and is maintained in all types of developing gut muscles. We show that biniou activity is essential for maintaining the distinction between splanchnic and somatic mesoderm and for differentiation of the splanchnic mesoderm into midgut musculature. biniou is required not only for the activation of differentiation genes that are expressed ubiquitously in the trunk visceral mesoderm but also for the expression of dpp in parasegment 7, which governs proper midgut morphogenesis. Activation of dpp is mediated by specific Biniou binding sites in a dpp enhancer element, which suggests that Biniou serves as a tissue-specific cofactor of homeotic gene products in visceral mesoderm patterning. Based upon these and other data, we propose that the splanchnic mesoderm layers in Drosophila and vertebrate embryos are homologous structures whose development into gut musculature and other visceral organs is critically dependent on FoxF genes. PMID:11691840

  11. The convergence of Notch and MAPK signaling specifies the blood progenitor fate in the Drosophila mesoderm

    PubMed Central

    Grigorian, Melina; Mandal, Lolitika; Hakimi, Manuel; Ortiz, Irma; Hartenstein, Volker

    2011-01-01

    Blood progenitors arise from a pool of pluripotential cells (“hemangioblasts”) within the Drosophila embryonic mesoderm. The fact that the cardiogenic mesoderm consists of only a small number of highly stereotypically patterned cells that can be queried individually regarding their gene expression in normal and mutant embryos, is one of the significant advantages that Drosophila offers to dissect the mechanism specifying the fate of these cells. We show in this paper that the expression of the Notch ligand Delta (Dl) reveals segmentally reiterated mesodermal clusters (“cardiogenic clusters”) that constitute the cardiogenic mesoderm. These clusters give rise to cardioblasts, blood progenitors and nephrocytes. Cardioblasts emerging from the cardiogenic clusters accumulate high levels of Dl, which is required to prevent more cells from adopting the cardioblast fate. In embryos lacking Dl function, all cells of the cardiogenic clusters become cardioblasts, and blood progenitors are lacking. Concomitant activation of the Mitogen Activated Protein Kinase (MAPK) pathway by Epidermal Growth Factor Receptor (EGFR) and Fibroblast Growth Factor Receptor (FGFR) is required for the specification and maintenance of the cardiogenic mesoderm; in addition, the spatially restricted localization of some of the FGFR ligands may be instrumental in controlling the spatial restriction of the Dl ligand to presumptive cardioblasts. PMID:21382367

  12. Temporal and spatial requirements for Nodal-induced anterior mesendoderm and mesoderm in anterior neurulation.

    PubMed

    Gonsar, Ngawang; Coughlin, Alicia; Clay-Wright, Jessica A; Borg, Bethanie R; Kindt, Lexy M; Liang, Jennifer O

    2016-01-01

    Zebrafish with defective Nodal signaling have a phenotype analogous to the fatal human birth defect anencephaly, which is caused by an open anterior neural tube. Previous work in our laboratory found that anterior open neural tube phenotypes in Nodal signaling mutants were caused by lack of mesendodermal/mesodermal tissues. Defects in these mutants are already apparent at neural plate stage, before the neuroepithelium starts to fold into a tube. Consistent with this, we found that the requirement for Nodal signaling maps to mid-late blastula stages. This timing correlates with the timing of prechordal plate mesendoderm and anterior mesoderm induction, suggesting these tissues act to promote neurulation. To further identify tissues important for neurulation, we took advantage of the variable phenotypes in Nodal signaling-deficient sqt mutant and Lefty1-overexpressing embryos. Statistical analysis indicated a strong, positive correlation between a closed neural tube and presence of several mesendoderm/mesoderm-derived tissues (hatching glands, cephalic paraxial mesoderm, notochord, and head muscles). However, the neural tube was closed in a subset of embryos that lacked any one of these tissues. This suggests that several types of Nodal-induced mesendodermal/mesodermal precursors are competent to promote neurulation. PMID:26528772

  13. The entire mesodermal mantle behaves as Spemann's organizer in dorsoanterior enhanced Xenopus laevis embryos.

    PubMed

    Kao, K R; Elinson, R P

    1988-05-01

    The body plan of Xenopus laevis can be respecified by briefly exposing early cleavage stage embryos to lithium. Such embryos develop exaggerated dorsoanterior structures such as a radial eye and cement gland (K.R. Kao, Y. Masui, and R.P. Elinson, 1986, Nature (London) 322, 371-373). In this paper, we demonstrate that the enhanced dorsoanterior phenotype results from an overcommitment of mesoderm to dorsoanterior mesoderm. Histological and immunohistochemical observations reveal that the embryos have a greatly enlarged notochord with very little muscle tissue. In addition, they develop a radial, beating heart, suggesting that lithium also specifies anterior mesoderm and pharyngeal endoderm. Randomly oriented diametrically opposed marginal zone grafts from lithium-treated embryos, when transplanted into ultraviolet (uv)-irradiated axis-deficient hosts, rescue dorsal axial structures. These transplantation experiments demonstrate that the entire marginal zone of the early gastrula consists of presumptive dorsal mesoderm. Vital dye marking experiments also indicate that the entire marginal zone maps to the prominent proboscis that is composed of chordamesoderm and represents the long axis of the embryo. These results suggest that lithium respecifies the mesoderm of Xenopus laevis embryos so that it differentiates into the Spemann organizer. We suggest that the origin of the dorsoanterior enhanced phenotypes generated by lithium and the dorsoanterior deficient phenotypes generated by uv irradiation are due to relative quantities of organizer. Our evidence demonstrates the existence of a continuum of body plan phenotypes based on this premise. PMID:3282938

  14. The convergence of Notch and MAPK signaling specifies the blood progenitor fate in the Drosophila mesoderm.

    PubMed

    Grigorian, Melina; Mandal, Lolitika; Hakimi, Manuel; Ortiz, Irma; Hartenstein, Volker

    2011-05-01

    Blood progenitors arise from a pool of pluripotential cells ("hemangioblasts") within the Drosophila embryonic mesoderm. The fact that the cardiogenic mesoderm consists of only a small number of highly stereotypically patterned cells that can be queried individually regarding their gene expression in normal and mutant embryos is one of the significant advantages that Drosophila offers to dissect the mechanism specifying the fate of these cells. We show in this paper that the expression of the Notch ligand Delta (Dl) reveals segmentally reiterated mesodermal clusters ("cardiogenic clusters") that constitute the cardiogenic mesoderm. These clusters give rise to cardioblasts, blood progenitors and nephrocytes. Cardioblasts emerging from the cardiogenic clusters accumulate high levels of Dl, which is required to prevent more cells from adopting the cardioblast fate. In embryos lacking Dl function, all cells of the cardiogenic clusters become cardioblasts, and blood progenitors are lacking. Concomitant activation of the Mitogen Activated Protein Kinase (MAPK) pathway by Epidermal Growth Factor Receptor (EGFR) and Fibroblast Growth Factor Receptor (FGFR) is required for the specification and maintenance of the cardiogenic mesoderm; in addition, the spatially restricted localization of some of the FGFR ligands may be instrumental in controlling the spatial restriction of the Dl ligand to presumptive cardioblasts. PMID:21382367

  15. Analysis of extraembryonic mesodermal structure formation in the absence of morphological primitive streak.

    PubMed

    Jin, Jiu-Zhen; Zhu, Yuanqi; Warner, Dennis; Ding, Jixiang

    2016-08-01

    During mouse gastrulation, the primitive streak is formed on the posterior side of the embryo. Cells migrate out of the primitive streak to form the future mesoderm and endoderm. Fate mapping studies revealed a group of cell migrate through the proximal end of the primitive streak and give rise to the extraembryonic mesoderm tissues such as the yolk sac blood islands and allantois. However, it is not clear whether the formation of a morphological primitive streak is required for the development of these extraembryonic mesodermal tissues. Loss of the Cripto gene in mice dramatically reduces, but does not completely abolish, Nodal activity leading to the absence of a morphological primitive streak. However, embryonic erythrocytes are still formed and assembled into the blood islands. In addition, Cripto mutant embryos form allantoic buds. However, Drap1 mutant embryos have excessive Nodal activity in the epiblast cells before gastrulation and form an expanded primitive streak, but no yolk sac blood islands or allantoic bud formation. Lefty2 embryos also have elevated levels of Nodal activity in the primitive streak during gastrulation, and undergo normal blood island and allantois formation. We therefore speculate that low level of Nodal activity disrupts the formation of morphological primitive streak on the posterior side, but still allows the formation of primitive streak cells on the proximal side, which give rise to the extraembryonic mesodermal tissues formation. Excessive Nodal activity in the epiblast at pre-gastrulation stage, but not in the primitive streak cells during gastrulation, disrupts extraembryonic mesoderm development. PMID:27273137

  16. Mesodermal Gene Expression in the Acoel Isodiametra pulchra Indicates a Low Number of Mesodermal Cell Types and the Endomesodermal Origin of the Gonads

    PubMed Central

    Chiodin, Marta; Børve, Aina; Berezikov, Eugene; Ladurner, Peter; Martinez, Pedro; Hejnol, Andreas

    2013-01-01

    Acoelomorphs are bilaterally symmetric small marine worms that lack a coelom and possess a digestive system with a single opening. Two alternative phylogenetic positions of this group within the animal tree are currently debated. In one view, Acoelomorpha is the sister group to all remaining Bilateria and as such, is a morphologically simple stepping stone in bilaterian evolution. In the other, the group is a lineage within the Deuterostomia, and therefore, has derived a simple morphology from a more complex ancestor. Acoels and the closely related Nemertodermatida and Xenoturbellida, which together form the Acoelomorpha, possess a very limited number of cell types. To further investigate the diversity and origin of mesodermal cell types we describe the expression pattern of 12 orthologs of bilaterian mesodermal markers including Six1/2, Twist, FoxC, GATA4/5/6, in the acoel Isodiametra pulchra. All the genes are expressed in stem cells (neoblasts), gonads, and at least subsets of the acoel musculature. Most are expressed in endomesodermal compartments of I. pulchra developing embryos similar to what has been described in cnidarians. Our molecular evidence indicates a very limited number of mesodermal cell types and suggests an endomesodermal origin of the gonads and the stem cell system. We discuss our results in light of the two prevailing phylogenetic positions of Acoelomorpha. PMID:23405161

  17. Mesoderm is required for coordinated cell movements within zebrafish neural plate in vivo

    PubMed Central

    2014-01-01

    Background Morphogenesis of the zebrafish neural tube requires the coordinated movement of many cells in both time and space. A good example of this is the movement of the cells in the zebrafish neural plate as they converge towards the dorsal midline before internalizing to form a neural keel. How these cells are regulated to ensure that they move together as a coherent tissue is unknown. Previous work in other systems has suggested that the underlying mesoderm may play a role in this process but this has not been shown directly in vivo. Results Here we analyze the roles of subjacent mesoderm in the coordination of neural cell movements during convergence of the zebrafish neural plate and neural keel formation. Live imaging demonstrates that the normal highly coordinated movements of neural plate cells are lost in the absence of underlying mesoderm and the movements of internalization and neural tube formation are severely disrupted. Despite this, neuroepithelial polarity develops in the abnormal neural primordium but the resulting tissue architecture is very disorganized. Conclusions We show that the movements of cells in the zebrafish neural plate are highly coordinated during the convergence and internalization movements of neurulation. Our results demonstrate that the underlying mesoderm is required for these coordinated cell movements in the zebrafish neural plate in vivo. PMID:24755297

  18. A mesoderm-derived precursor for mesenchymal stem and endothelial cells

    PubMed Central

    Vodyanik, Maxim A.; Yu, Junying; Zhang, Xin; Tian, Shulan; Stewart, Ron; Thomson, James A.; Slukvin, Igor I.

    2010-01-01

    Summary Among the three embryonic germ layers, the mesoderm is a major source of the mesenchymal precursors giving rise to skeletal and connective tissues. These precursors, however, have not been previously identified and characterized. Using human embryonic stem cells directed to mesendodermal differentiation, here we show that mesenchymal stem/stromal cells (MSCs) originate from a population of mesodermal cells identified by expression of the apelin receptor. In semisolid medium, these precursors form FGF2-dependent compact spheroid colonies containing mesenchymal cells with a transcriptional profile representative of mesoderm-derived embryonic mesenchyme. When transferred to the adherent cultures, individual colonies give rise to MSC lines with chondro-, osteo-, and adipogenenic differentiation potentials. Although the MSC lines lacked endothelial potential, endothelial cells could be derived from mesenchymal colonies, suggesting that, similar to hematopoietic cells, MSCs arise from precursors with angiogenic potential. Together, these studies identified a common precursor of mesenchymal and endothelial cells, mesenchymoangioblast, as the source of mesoderm-derived MSCs. PMID:21112566

  19. How the sea squirt nucleus tells mesoderm not to be endoderm.

    PubMed

    Parton, Richard M; Davis, Ilan

    2010-10-19

    Sea squirts are simple invertebrate chordates. In this issue of Developmental Cell, Takatori et al. show nuclear migration within ascidian mesendodermal cells enables polarized localization of Not mRNA, which encodes a homeobox protein that distinguishes mesoderm from endoderm fates. The link between nuclear migration and mRNA localization suggests exciting parallels with protostomes. PMID:20951340

  20. Hedgehog/Patched-associated rhabdomyosarcoma formation from delta1-expressing mesodermal cells.

    PubMed

    Nitzki, F; Cuvelier, N; Dräger, J; Schneider, A; Braun, T; Hahn, H

    2016-06-01

    Rhabdomyosarcoma (RMS) is the most common pediatric soft tissue sarcoma. In children, the 2 major RMS subtypes are alveolar and embryonal RMS. Aberrant Hedgehog/Patched1 (Hh/Ptch) signaling is a hallmark of embryonal RMS. We demonstrate that mice carrying a Ptch mutation in mesodermal Delta1-expressing cells develop embryonal-like RMS at a similar rate as mice harboring a Ptch mutation in the germline or the brachury-expressing mesoderm. The tumor incidence decreases dramatically when Ptch is mutated in Myf5- or Pax3-expressing cells. No RMS develop from Myogenin/Mef2c-expressing cells. This suggests that Hh/Ptch-associated RMS are derived from Delta1-positive, Myf5-negative, Myogenin-negative and Pax3-negative mesodermal progenitors that can undergo myogenic differentiation but lack stable lineage commitment. Additional preliminary genetic data and data on mesodermal progenitors further imply an interplay of Hh/Ptch and Delta/Notch signaling activity during RMS initiation. In contrast, Wnt signals supposedly suppress RMS formation because RMS multiplicity decreases after inactivation of the Wnt-inhibitor Wif1. Finally, our results strongly suggest that the tumor-initiating event determines the lineage of RMS origin. PMID:26387541

  1. How the sea squirt nucleus tells mesoderm Not to be endoderm

    PubMed Central

    Parton, Richard M.; Davis, Ilan

    2011-01-01

    Sea squirts are simple invertebrate chordates. In this issue, Takatori et al show nuclear migration within ascidian mesendodermal cells enables polarized localization of Not mRNA, which encodes a homeobox protein that distinguishes mesoderm from endoderm fates. The link between nuclear migration and mRNA localization suggests exciting parallels with protostomes. PMID:20951340

  2. Mesodermal origin of median fin mesenchyme and tail muscle in amphibian larvae.

    PubMed

    Taniguchi, Yuka; Kurth, Thomas; Medeiros, Daniel Meulemans; Tazaki, Akira; Ramm, Robert; Epperlein, Hans-Henning

    2015-01-01

    Mesenchyme is an embryonic precursor tissue that generates a range of structures in vertebrates including cartilage, bone, muscle, kidney, and the erythropoietic system. Mesenchyme originates from both mesoderm and the neural crest, an ectodermal cell population, via an epithelial to mesenchymal transition (EMT). Because ectodermal and mesodermal mesenchyme can form in close proximity and give rise to similar derivatives, the embryonic origin of many mesenchyme-derived tissues is still unclear. Recent work using genetic lineage tracing methods have upended classical ideas about the contributions of mesodermal mesenchyme and neural crest to particular structures. Using similar strategies in the Mexican axolotl (Ambystoma mexicanum), and the South African clawed toad (Xenopus laevis), we traced the origins of fin mesenchyme and tail muscle in amphibians. Here we present evidence that fin mesenchyme and striated tail muscle in both animals are derived solely from mesoderm and not from neural crest. In the context of recent work in zebrafish, our experiments suggest that trunk neural crest cells in the last common ancestor of tetrapods and ray-finned fish lacked the ability to form ectomesenchyme and its derivatives. PMID:26086331

  3. Mesodermal origin of median fin mesenchyme and tail muscle in amphibian larvae

    PubMed Central

    Taniguchi, Yuka; Kurth, Thomas; Medeiros, Daniel Meulemans; Tazaki, Akira; Ramm, Robert; Epperlein, Hans-Henning

    2015-01-01

    Mesenchyme is an embryonic precursor tissue that generates a range of structures in vertebrates including cartilage, bone, muscle, kidney, and the erythropoietic system. Mesenchyme originates from both mesoderm and the neural crest, an ectodermal cell population, via an epithelial to mesenchymal transition (EMT). Because ectodermal and mesodermal mesenchyme can form in close proximity and give rise to similar derivatives, the embryonic origin of many mesenchyme-derived tissues is still unclear. Recent work using genetic lineage tracing methods have upended classical ideas about the contributions of mesodermal mesenchyme and neural crest to particular structures. Using similar strategies in the Mexican axolotl (Ambystoma mexicanum), and the South African clawed toad (Xenopus laevis), we traced the origins of fin mesenchyme and tail muscle in amphibians. Here we present evidence that fin mesenchyme and striated tail muscle in both animals are derived solely from mesoderm and not from neural crest. In the context of recent work in zebrafish, our experiments suggest that trunk neural crest cells in the last common ancestor of tetrapods and ray-finned fish lacked the ability to form ectomesenchyme and its derivatives. PMID:26086331

  4. Generation of an expandable intermediate mesoderm restricted progenitor cell line from human pluripotent stem cells

    PubMed Central

    Kumar, Nathan; Richter, Jenna; Cutts, Josh; Bush, Kevin T; Trujillo, Cleber; Nigam, Sanjay K; Gaasterland, Terry; Brafman, David; Willert, Karl

    2015-01-01

    The field of tissue engineering entered a new era with the development of human pluripotent stem cells (hPSCs), which are capable of unlimited expansion whilst retaining the potential to differentiate into all mature cell populations. However, these cells harbor significant risks, including tumor formation upon transplantation. One way to mitigate this risk is to develop expandable progenitor cell populations with restricted differentiation potential. Here, we used a cellular microarray technology to identify a defined and optimized culture condition that supports the derivation and propagation of a cell population with mesodermal properties. This cell population, referred to as intermediate mesodermal progenitor (IMP) cells, is capable of unlimited expansion, lacks tumor formation potential, and, upon appropriate stimulation, readily acquires properties of a sub-population of kidney cells. Interestingly, IMP cells fail to differentiate into other mesodermally-derived tissues, including blood and heart, suggesting that these cells are restricted to an intermediate mesodermal fate. DOI: http://dx.doi.org/10.7554/eLife.08413.001 PMID:26554899

  5. Changes in glycosaminoglycan structure on differentiation of human embryonic stem cells towards mesoderm and endoderm lineages

    PubMed Central

    Gasimli, Leyla; Hickey, Anne M.; Yang, Bo; Li, Guoyun; Rosa, Mitche dela; Nairn, Alison V.; Kulik, Michael J.; Dordick, Jonathan S.; Moremen, Kelley W.; Dalton, Stephen; Linhardt, Robert J.

    2014-01-01

    Background Proteoglycans are found on the cell surface and in the extracellular matrix, and serve as prime sites for interaction with signaling molecules. Proteoglycans help regulate pathways that control stem cell fate, and therefore represent an excellent tool to manipulate these pathways. Despite their importance, there is a dearth of data linking glycosaminoglycan structure within proteoglycans with stem cell differentiation. Methods Human embryonic stem cell line WA09 (H9) was differentiated into early mesoderm and endoderm lineages, and the glycosaminoglycanomic changes accompanying these transitions were studied using transcript analysis, immunoblotting, immunofluorescence and disaccharide analysis. Results Pluripotent H9 cell lumican had no glycosaminoglycan chains whereas in splanchnic mesoderm lumican was glycosaminoglycanated. H9 cells have primarily non-sulfated heparan sulfate chains. On differentiation towards splanchnic mesoderm and hepatic lineages N-sulfo group content increases. Differences in transcript expression of NDST1, HS6ST2 and HS6ST3, three heparan sulfate biosynthetic enzymes, within splanchnic mesoderm cells compared to H9 cells correlate to changes in glycosaminoglycan structure. Conclusions Differentiation of embryonic stem cells markedly change the proteoglycanome General Significance The glycosaminoglycan biosynthetic pathway is complex and highly regulated, and therefore, understanding the details of this pathway should enable better control with the aim of directing stem cell differentiation. PMID:24412195

  6. Mesodermal Tbx1 is required for patterning the proximal mandible in mice

    PubMed Central

    Aggarwal, Vimla S.; Carpenter, Courtney; Freyer, Laina; Liao, Jun; Petti, Marilena; Morrow, Bernice E.

    2010-01-01

    Defects in the lower jaw, or mandible, occur commonly either as isolated malformations or in association with genetic syndromes. Understanding its formation and genetic pathways required for shaping its structure in mammalian model organisms will shed light into the pathogenesis of malformations in humans. The lower jaw is derived from the mandibular process of the first pharyngeal arch (MdPA1) during embryogenesis. Integral to the development of the mandible, is the signaling interplay between Fgf8 and Bmp4 in the rostral ectoderm and their downstream effector genes in the underlying neural crest derived mesenchyme. The non-neural crest MdPA1 core mesoderm is needed to form muscles of mastication, but its role in patterning the mandible is unknown. Here, we show that mesoderm specific deletion of Tbx1, a T- box transcription factor and gene for velo-cardio-facial/DiGeorge syndrome, results in defects in formation of the proximal mandible by shifting expression of Fgf8, Bmp4 and their downstream effector genes in mouse embryos at E10.5. This occurs without significant changes in cell proliferation or apoptosis at the same stage. Our results elucidate a new function for the non-neural crest core mesoderm and specifically, mesodermal Tbx1, in shaping the lower jaw. PMID:20501333

  7. Functional dissection of Pax3 in paraxial mesoderm development and myogenesis

    PubMed Central

    Magli, Alessandro; Schnettler, Erin; Rinaldi, Fabrizio; Bremer, Paul; Perlingeiro, Rita C. R.

    2012-01-01

    The paired box transcription factor Pax3 is well-known as a major regulator of embryonic myogenesis. Before Pax3 expression becomes restricted to the dermomyotome, this transcription factor is also expressed in the developing somites. The role of Pax3 at this early stage is unclear, in particular because of the scarce frequency of Pax3-positive cells in the early mouse embryo. Inducible gene expression in embryonic stem (ES) cells represents an excellent tool to overcome this limitation, since it can provide large quantities of otherwise rare embryonic populations expressing a factor of interest. Here we used engineered mouse ES cells to perform a functional analysis of Pax3 with the aim to identify the molecular determinants involved in the early functions of this transcription factor. We find that Pax3 induction during embryoid body (EB) differentiation results in the up-regulation of genes expressed in the presomitic and somitic mesoderm. Moreover, we show that paraxial mesoderm induced by transient expression of Pax3 is not irreversibly committed to myogenesis, rather requires sustained Pax3 expression. Using a series of deletion mutants of Pax3, which differentially affect its transcriptional activity, we map protein domains necessary for induction of paraxial mesoderm and induction of the myogenic program. The paired, homeo- and transcriptional activation domains were each required for both processes, however the paired-c-terminal RED domain showed a paraxial mesoderm-specific activity that was dispensable for myogenesis. These findings demonstrate and provide mechanistic insight into an early role for Pax3 in the generation of paraxial mesoderm. PMID:23081715

  8. Axial elongation in mouse embryos involves mediolateral cell intercalation behavior in the paraxial mesoderm

    NASA Astrophysics Data System (ADS)

    Yen, WeiWei; Burdsal, Carol; Periasamy, Ammasi; Sutherland, Ann E.

    2006-02-01

    The cell mechanical and signaling pathways involved in gastrulation have been studied extensively in invertebrates and amphibians, such as Xenopus, and more recently in non-mammalian vertebrates such as zebrafish and chick. However, because culturing mouse embryos extra-utero is very difficult, this fundamental process has been least characterized in the mouse. As the primary mammalian model for genetics, biochemistry, and the study of human disease and birth defects, it is important to investigate how gastrulation proceeds in murine embryos. We have developed a method of using 4D multiphoton excitation microscopy and extra-utero culture to visualize and characterize the morphogenetic movements in mouse embryos dissected at 8.5 days of gestation. Cells are labeled by expression of an X chromosome-linked enhanced green fluorescent protein (EGFP) transgene. This method has provided a unique approach, where, for the first time, patterns of cell behavior in the notochord and surrounding paraxial mesoderm can be visualized and traced quantitatively. Our observations of mouse embryos reveal both distinct differences as well as striking similarities in patterned cell motility relative to other vertebrate models such as Xenopus, where axial extension is driven primarily by mediolateral oriented cell behaviors in the notochord and paraxial somitic mesoderm. Unlike Xenopus, the width of the mouse notochord remains the same between 4-somite stage and 8-somite stage embryos. This implies the mouse notochord plays a lesser role in driving axial extension compared to Xenopus, although intercalation may occur where the anterior region of the node becomes notochordal plate. In contrast, the width of mouse paraxial mesoderm narrows significantly during this period and cells within the paraxial mesoderm are both elongated and aligned perpendicular to the midline. In addition, these cells are observed to intercalate, consistent with a role for paraxial mesoderm in driving convergence

  9. Diversification of oral and aboral mesodermal regulatory states in pregastrular sea urchin embryos.

    PubMed

    Materna, Stefan C; Ransick, Andrew; Li, Enhu; Davidson, Eric H

    2013-03-01

    Specification of the non-skeletogenic mesoderm (NSM) in sea urchin embryos depends on Delta signaling. Signal reception leads to expression of regulatory genes that later contribute to the aboral NSM regulatory state. In oral NSM, this is replaced by a distinct oral regulatory state in consequence of Nodal signaling. Through regulome wide analysis we identify the homeobox gene not as an immediate Nodal target. not expression in NSM causes extinction of the aboral regulatory state in the oral NSM, and expression of a new suite of regulatory genes. All NSM specific regulatory genes are henceforth expressed exclusively, in oral or aboral domains, presaging the mesodermal cell types that will emerge. We have analyzed the regulatory linkages within the aboral NSM gene regulatory network. A linchpin of this network is gataE which as we show is a direct Gcm target and part of a feedback loop locking down the aboral regulatory state. PMID:23261933

  10. Development of Bipotent Cardiac/Skeletal Myogenic Progenitors from MESP1+ Mesoderm

    PubMed Central

    Chan, Sunny Sun-Kin; Hagen, Hannah R.; Swanson, Scott A.; Stewart, Ron; Boll, Karly A.; Aho, Joy; Thomson, James A.; Kyba, Michael

    2016-01-01

    Summary The branchiomeric skeletal muscles co-evolved with new chambers of the heart to enable predatory feeding in chordates. These co-evolved tissues develop from a common population in anterior splanchnic mesoderm, referred to as cardiopharyngeal mesoderm (CPM). The regulation and development of CPM are poorly understood. We describe an embryonic stem cell-based system in which MESP1 drives a PDGFRA+ population with dual cardiac and skeletal muscle differentiation potential, and gene expression resembling CPM. Using this system, we investigate the regulation of these bipotent progenitors, and find that cardiac specification is governed by an antagonistic TGFβ-BMP axis, while skeletal muscle specification is enhanced by Rho kinase inhibition. We define transcriptional signatures of the first committed CPM-derived cardiac and skeletal myogenic progenitors, and discover surface markers to distinguish cardiac (PODXL+) from the skeletal muscle (CDH4+) CPM derivatives. These tools open an accessible window on this developmentally and evolutionarily important population. PMID:26771351

  11. Qualitative Dynamical Modelling Can Formally Explain Mesoderm Specification and Predict Novel Developmental Phenotypes.

    PubMed

    Mbodj, Abibatou; Gustafson, E Hilary; Ciglar, Lucia; Junion, Guillaume; Gonzalez, Aitor; Girardot, Charles; Perrin, Laurent; Furlong, Eileen E M; Thieffry, Denis

    2016-09-01

    Given the complexity of developmental networks, it is often difficult to predict the effect of genetic perturbations, even within coding genes. Regulatory factors generally have pleiotropic effects, exhibit partially redundant roles, and regulate highly interconnected pathways with ample cross-talk. Here, we delineate a logical model encompassing 48 components and 82 regulatory interactions involved in mesoderm specification during Drosophila development, thereby providing a formal integration of all available genetic information from the literature. The four main tissues derived from mesoderm correspond to alternative stable states. We demonstrate that the model can predict known mutant phenotypes and use it to systematically predict the effects of over 300 new, often non-intuitive, loss- and gain-of-function mutations, and combinations thereof. We further validated several novel predictions experimentally, thereby demonstrating the robustness of model. Logical modelling can thus contribute to formally explain and predict regulatory outcomes underlying cell fate decisions. PMID:27599298

  12. Identification and characterization of a twist ortholog in the polychaete annelid Platynereis dumerilii reveals mesodermal expression of Pdu-twist.

    PubMed

    Pfeifer, Kathrin; Schaub, Christoph; Wolfstetter, Georg; Dorresteijn, Adriaan

    2013-09-01

    The basic helix-loop-helix transcription factor twist plays a key role during mesoderm development in Bilateria. In this study, we identified a twist ortholog in the polychaete annelid Platynereis dumerilii and analyze its expression during larval development, postlarval growth up to the adult stage, and caudal regeneration after amputation of posterior segments. At late larval stages, Pdu-twist is expressed in the mesodermal anlagen and in developing muscles. During adulthood and caudal regeneration, Pdu-twist is expressed in the posterior growth zone, in mesodermal cells within the newly forming segments and budding parapodia. Our results indicate that Pdu-twist is involved in mesoderm formation during larval development, posterior growth, and caudal regeneration. PMID:23817621

  13. Stochastic specification of primordial germ cells from mesoderm precursors in axolotl embryos.

    PubMed

    Chatfield, Jodie; O'Reilly, Marie-Anne; Bachvarova, Rosemary F; Ferjentsik, Zoltan; Redwood, Catherine; Walmsley, Maggie; Patient, Roger; Loose, Mathew; Johnson, Andrew D

    2014-06-01

    A common feature of development in most vertebrate models is the early segregation of the germ line from the soma. For example, in Xenopus and zebrafish embryos primordial germ cells (PGCs) are specified by germ plasm that is inherited from the egg; in mice, Blimp1 expression in the epiblast mediates the commitment of cells to the germ line. How these disparate mechanisms of PGC specification evolved is unknown. Here, in order to identify the ancestral mechanism of PGC specification in vertebrates, we studied PGC specification in embryos from the axolotl (Mexican salamander), a model for the tetrapod ancestor. In the axolotl, PGCs develop within mesoderm, and classic studies have reported their induction from primitive ectoderm (animal cap). We used an axolotl animal cap system to demonstrate that signalling through FGF and BMP4 induces PGCs. The role of FGF was then confirmed in vivo. We also showed PGC induction by Brachyury, in the presence of BMP4. These conditions induced pluripotent mesodermal precursors that give rise to a variety of somatic cell types, in addition to PGCs. Irreversible restriction of the germ line did not occur until the mid-tailbud stage, days after the somatic germ layers are established. Before this, germline potential was maintained by MAP kinase signalling. We propose that this stochastic mechanism of PGC specification, from mesodermal precursors, is conserved in vertebrates. PMID:24917499

  14. A core transcriptional network for early mesoderm development in Drosophila melanogaster

    PubMed Central

    Sandmann, Thomas; Girardot, Charles; Brehme, Marc; Tongprasit, Waraporn; Stolc, Viktor; Furlong, Eileen E.M.

    2007-01-01

    Embryogenesis is controlled by large gene-regulatory networks, which generate spatially and temporally refined patterns of gene expression. Here, we report the characteristics of the regulatory network orchestrating early mesodermal development in the fruitfly Drosophila, where the transcription factor Twist is both necessary and sufficient to drive development. Through the integration of chromatin immunoprecipitation followed by microarray analysis (ChIP-on-chip) experiments during discrete time periods with computational approaches, we identified >2000 Twist-bound cis-regulatory modules (CRMs) and almost 500 direct target genes. Unexpectedly, Twist regulates an almost complete cassette of genes required for cell proliferation in addition to genes essential for morophogenesis and cell migration. Twist targets almost 25% of all annotated Drosophila transcription factors, which may represent the entire set of regulators necessary for the early development of this system. By combining in vivo binding data from Twist, Mef2, Tinman, and Dorsal we have constructed an initial transcriptional network of early mesoderm development. The network topology reveals extensive combinatorial binding, feed-forward regulation, and complex logical outputs as prevalent features. In addition to binary activation and repression, we suggest that Twist binds to almost all mesodermal CRMs to provide the competence to integrate inputs from more specialized transcription factors. PMID:17322403

  15. An NF-κB and Slug Regulatory Loop Active in Early Vertebrate Mesoderm

    PubMed Central

    Zhang, Chi; Carl, Timothy F.; Trudeau, Evan D.; Simmet, Thomas; Klymkowsky, Michael W.

    2006-01-01

    Background In both Drosophila and the mouse, the zinc finger transcription factor Snail is required for mesoderm formation; its vertebrate paralog Slug (Snai2) appears to be required for neural crest formation in the chick and the clawed frog Xenopus laevis. Both Slug and Snail act to induce epithelial to mesenchymal transition (EMT) and to suppress apoptosis. Methodology & Principle Findings Morpholino-based loss of function studies indicate that Slug is required for the normal expression of both mesodermal and neural crest markers in X. laevis. Both phenotypes are rescued by injection of RNA encoding the anti-apoptotic protein Bcl-xL; Bcl-xL's effects are dependent upon IκB kinase-mediated activation of the bipartite transcription factor NF-κB. NF-κB, in turn, directly up-regulates levels of Slug and Snail RNAs. Slug indirectly up-regulates levels of RNAs encoding the NF-κB subunit proteins RelA, Rel2, and Rel3, and directly down-regulates levels of the pro-apopotic Caspase-9 RNA. Conclusions/Significance These studies reveal a Slug/Snail–NF-κB regulatory circuit, analogous to that present in the early Drosophila embryo, active during mesodermal formation in Xenopus. This is a regulatory interaction of significance both in development and in the course of inflammatory and metastatic disease. PMID:17205110

  16. Claudin-10 is required for relay of left-right patterning cues from Hensen's node to the lateral plate mesoderm.

    PubMed

    Collins, Michelle M; Baumholtz, Amanda I; Simard, Annie; Gregory, Mary; Cyr, Daniel G; Ryan, Aimee K

    2015-05-15

    Species-specific symmetry-breaking events at the left-right organizer (LRO) drive an evolutionarily-conserved cascade of gene expression in the lateral plate mesoderm that is required for the asymmetric positioning of organs within the body cavity. The mechanisms underlying the transfer of the left and right laterality information from the LRO to the lateral plate mesoderm are poorly understood. Here, we investigate the role of Claudin-10, a tight junction protein, in facilitating the transfer of left-right identity from the LRO to the lateral plate mesoderm. Claudin-10 is asymmetrically expressed on the right side of the chick LRO, Hensen's node. Gain- and loss-of-function studies demonstrated that right-sided expression of Claudin-10 is essential for normal rightward heart tube looping, the first morphological asymmetry during organogenesis. Manipulation of Claudin-10 expression did not perturb asymmetric gene expression at Hensen's node, but did disrupt asymmetric gene expression in the lateral plate mesoderm. Bilateral expression of Claudin-10 at Hensen's node prevented expression of Nodal, Lefty-2 and Pitx2c in the left lateral plate mesoderm, while morpholino knockdown of Claudin-10 inhibited expression of Snail1 in the right lateral plate mesoderm. We also determined that amino acids that are predicted to affect ion selectivity and protein interactions that bridge Claudin-10 to the actin cytoskeleton were essential for its left-right patterning function. Collectively, our data demonstrate a novel role for Claudin-10 during the transmission of laterality information from Hensen's node to both the left and right sides of the embryo and demonstrate that tight junctions have a critical role during the relay of left-right patterning cues from Hensen's node to the lateral plate mesoderm. PMID:25744724

  17. Mesodermal expression of the C. elegans HMX homolog mls-2 requires the PBC homolog CEH-20

    PubMed Central

    Jiang, Yuan; Shi, Herong; Amin, Nirav M.; Sultan, Ibrahim; Liu, Jun

    2008-01-01

    Metazoan development proceeds primarily through the regulated expression of genes encoding transcription factors and components of cell signaling pathways. One way to decipher the complex developmental programs is to assemble the underlying gene regulatory networks by dissecting the cis-regulatory modules that direct temporal-spatial expression of developmental genes and identify corresponding trans-regulatory factors. Here, we focus on the regulation of a HMX homoebox gene called mls-2, which functions at the intersection of a network that regulates cleavage orientation, cell proliferation and fate specification in the C. elegans postembryonic mesoderm. In addition to its transient expression in the postembryonic mesodermal lineage, the M lineage, mls-2 expression is detected in a subset of embryonic cells, in three pairs of head neurons and transiently in the somatic gonad. Through mutational analysis of the mls-2 promoter, we identified two elements (E1 and E2) involved in regulating the temporal-spatial expression of mls-2. In particular, we showed that one of the elements (E1) required for mls-2 expression in the M lineage contains two critical putative PBC-Hox binding sites that are evolutionarily conserved in C. briggsae and C. remanei. Furthermore, the C. elegans PBC homolog CEH-20 is required for mls-2 expression in the M lineage. Our data suggests that mls-2 might be a direct target of CEH-20 in the M lineage and that the regulation of CEH-20 on mls-2 is likely Hox-independent. PMID:18316179

  18. Neurotrophin receptor homolog (NRH1) proteins regulate mesoderm formation and apoptosis during early Xenopus development.

    PubMed

    Knapp, Dunja; Messenger, Nigel; Ahmed Rana, Amer; Smith, James C

    2006-12-15

    Recent experiments suggest that Xenopus Neurotrophin Receptor Homolog 1 (NRH1) proteins act through the planar cell polarity pathway to regulate convergent extension movements during gastrulation and neurulation. We show in this paper that NRH1 proteins are also required for the proper expression of mesodermally expressed genes such as Xbra and Chordin, and to a lesser extent, of Xwnt11. Loss of NRH1 function is followed, during gastrula and neurula stages, by a dramatic increase in apoptosis. Apoptosis is delayed by injection of Xbra RNA, suggesting that cell death is a consequence, at least in part, of the down-regulation of this gene, and it is also delayed by expression of activated forms of Rho, Rac and Cdc42. These small GTPases have previously been implicated in the planar cell polarity pathway in Xenopus and, in other systems, in the regulation of apoptosis. We conclude that the effects of NRH1 proteins include the regulation of mesodermal gene expression and that the disruption of gastrulation that is caused by their loss of function is a consequence of the down-regulation of Xbra and other genes, in addition to direct interference with the planar cell polarity pathway. The apoptosis observed in embryos lacking NRH1 function is not an indirect consequence of the disruption of gastrulation, and indeed it may contribute to the observed morphological defects. PMID:17055478

  19. A critical role for the chromatin remodeller CHD7 in anterior mesoderm during cardiovascular development.

    PubMed

    Payne, Sophie; Burney, Matthew J; McCue, Karen; Popal, Nelo; Davidson, Sean M; Anderson, Robert H; Scambler, Peter J

    2015-09-01

    CHARGE syndrome is caused by spontaneous loss-of-function mutations to the ATP-dependant chromatin remodeller chromodomain-helicase-DNA-binding protein 7 (CHD7). It is characterised by a distinct pattern of congenital anomalies, including cardiovascular malformations. Disruption to the neural crest lineage has previously been emphasised in the aetiology of this developmental disorder. We present evidence for an additional requirement for CHD7 activity in the Mesp1-expressing anterior mesoderm during heart development. Conditional ablation of Chd7 in this lineage results in major structural cardiovascular defects akin to those seen in CHARGE patients, as well as a striking loss of cardiac innervation and embryonic lethality. Genome-wide transcriptional analysis identified aberrant expression of key components of the Class 3 Semaphorin and Slit-Robo signalling pathways in Chd7(fl/fl);Mesp1-Cre mutant hearts. CHD7 localises at the Sema3c promoter in vivo, with alteration of the local chromatin structure seen following Chd7 ablation, suggestive of direct transcriptional regulation. Furthermore, we uncover a novel role for CHD7 activity upstream of critical calcium handling genes, and demonstrate an associated functional defect in the ability of cardiomyocytes to undergo excitation-contraction coupling. This work therefore reveals the importance of CHD7 in the cardiogenic mesoderm for multiple processes during cardiovascular development. PMID:26102480

  20. Differential mesodermal expression of two amphioxus MyoD family members (AmphiMRF1 and AmphiMRF2)

    NASA Technical Reports Server (NTRS)

    Schubert, Michael; Meulemans, Daniel; Bronner-Fraser, Marianne; Holland, Linda Z.; Holland, Nicholas D.

    2003-01-01

    To explore the evolution of myogenic regulatory factors in chordates, we isolated two MyoD family genes (AmphiMRF1 and AmphiMRF2) from amphioxus. AmphiMRF1 is first expressed at the late gastrula in the paraxial mesoderm. As the first somites form, expression is restricted to their myotomal region. In the early larva, expression is strongest in the most anterior and most posterior somites. AmphiMRF2 transcription begins at mid/late gastrula in the paraxial mesoderm, but never spreads into its most anterior region. Through much of the neurula stage, AmphiMRF2 expression is strong in the myotomal region of all somites except the most anterior pair; by late neurula expression is downregulated except in the most posterior somites forming just rostral to the tail bud. These two MRF genes of amphioxus have partly overlapping patterns of mesodermal expression and evidently duplicated independent of the diversification of the vertebrate MRF family.

  1. Nodal signaling is required for mesodermal and ventral but not for dorsal fates in the indirect developing hemichordate, Ptychodera flava

    PubMed Central

    Röttinger, Eric; DuBuc, Timothy Q.; Amiel, Aldine R.; Martindale, Mark Q.

    2015-01-01

    ABSTRACT Nodal signaling plays crucial roles in vertebrate developmental processes such as endoderm and mesoderm formation, and axial patterning events along the anteroposterior, dorsoventral and left-right axes. In echinoderms, Nodal plays an essential role in the establishment of the dorsoventral axis and left-right asymmetry, but not in endoderm or mesoderm induction. In protostomes, Nodal signaling appears to be involved only in establishing left-right asymmetry. Hence, it is hypothesized that Nodal signaling has been co-opted to pattern the dorsoventral axis of deuterostomes and for endoderm, mesoderm formation as well as anteroposterior patterning in chordates. Hemichordata, together with echinoderms, represent the sister taxon to chordates. In this study, we analyze the role of Nodal signaling in the indirect developing hemichordate Ptychodera flava. In particular, we show that during gastrulation nodal transcripts are detected in a ring of cells at the vegetal pole that gives rise to endomesoderm and in the ventral ectoderm at later stages of development. Inhibition of Nodal function disrupts dorsoventral fates and also blocks formation of the larval mesoderm. Interestingly, molecular analysis reveals that only mesodermal, apical and ventral gene expression is affected while the dorsal side appears to be patterned correctly. Taken together, this study suggests that the co-option of Nodal signaling in mesoderm formation and potentially in anteroposterior patterning has occurred prior to the emergence of chordates and that Nodal signaling on the ventral side is uncoupled from BMP signaling on the dorsal side, representing a major difference from the molecular mechanisms of dorsoventral patterning events in echinoderms. PMID:25979707

  2. Nodal signaling is required for mesodermal and ventral but not for dorsal fates in the indirect developing hemichordate, Ptychodera flava.

    PubMed

    Röttinger, Eric; DuBuc, Timothy Q; Amiel, Aldine R; Martindale, Mark Q

    2015-01-01

    Nodal signaling plays crucial roles in vertebrate developmental processes such as endoderm and mesoderm formation, and axial patterning events along the anteroposterior, dorsoventral and left-right axes. In echinoderms, Nodal plays an essential role in the establishment of the dorsoventral axis and left-right asymmetry, but not in endoderm or mesoderm induction. In protostomes, Nodal signaling appears to be involved only in establishing left-right asymmetry. Hence, it is hypothesized that Nodal signaling has been co-opted to pattern the dorsoventral axis of deuterostomes and for endoderm, mesoderm formation as well as anteroposterior patterning in chordates. Hemichordata, together with echinoderms, represent the sister taxon to chordates. In this study, we analyze the role of Nodal signaling in the indirect developing hemichordate Ptychodera flava. In particular, we show that during gastrulation nodal transcripts are detected in a ring of cells at the vegetal pole that gives rise to endomesoderm and in the ventral ectoderm at later stages of development. Inhibition of Nodal function disrupts dorsoventral fates and also blocks formation of the larval mesoderm. Interestingly, molecular analysis reveals that only mesodermal, apical and ventral gene expression is affected while the dorsal side appears to be patterned correctly. Taken together, this study suggests that the co-option of Nodal signaling in mesoderm formation and potentially in anteroposterior patterning has occurred prior to the emergence of chordates and that Nodal signaling on the ventral side is uncoupled from BMP signaling on the dorsal side, representing a major difference from the molecular mechanisms of dorsoventral patterning events in echinoderms. PMID:25979707

  3. Collinear activation of Hoxb genes during gastrulation is linked to mesoderm cell ingression.

    PubMed

    Iimura, Tadahiro; Pourquié, Olivier

    2006-08-01

    The vertebral column exhibits segmentation and regionalization along the antero-posterior axis. During embryogenesis, the rhythmic production of the precursors of the vertebrae, the somites, imposes a segmented aspect to the spine, whereas the spine's regional differentiation is controlled by Hox genes. Here we show that in the paraxial mesoderm, Hoxb genes are first activated in a temporal collinear fashion in precursors located in the epiblast lateral to the primitive streak. Our data suggest that collinear activation of Hoxb genes regulates the flux of cells from the epiblast to the streak and thus directly controls the establishment of the genes' characteristic nested expression domains in the somites. This suggests that establishment of the spatial co-linearity in the embryo is directly controlled by the Hox genes themselves. PMID:16760928

  4. Hoxd11 specifies a program of metanephric kidney development within the intermediate mesoderm of the mouse embryo.

    PubMed

    Mugford, Joshua W; Sipilä, Petra; Kobayashi, Akio; Behringer, Richard R; McMahon, Andrew P

    2008-07-15

    The mammalian kidney consists of an array of tubules connected to a ductal system that collectively function to control water/salt balance and to remove waste from the organisms' circulatory system. During mammalian embryogenesis, three kidney structures form within the intermediate mesoderm. The two most anterior structures, the pronephros and the mesonephros, are transitory and largely non-functional, while the most posterior, the metanephros, persists as the adult kidney. We have explored the mechanisms underlying regional specific differentiation of the kidney forming mesoderm. Previous studies have shown a requirement for Hox11 paralogs (Hoxa11, Hoxc11 and Hoxd11) in metanephric development. Mice lacking all Hox11 activity fail to form metanephric kidney structures. We demonstrate that the Hox11 paralog expression is restricted in the intermediate mesoderm to the posterior, metanephric level. When Hoxd11 is ectopically activated in the anterior mesonephros, we observe a partial transformation to a metanephric program of development. Anterior Hoxd11(+) cells activate Six2, a transcription factor required for the maintenance of metanephric tubule progenitors. Additionally, Hoxd11(+) mesonephric tubules exhibit an altered morphology and activate several metanephric specific markers normally confined to distal portions of the functional nephron. Collectively, our data support a model where Hox11 paralogs specify a metanephric developmental program in responsive intermediate mesoderm. This program maintains tubule forming progenitors and instructs a metanephric specific pattern of nephron differentiation. PMID:18485340

  5. Hoxd11 specifies a program of metanephric kidney development within the intermediate mesoderm of the mouse embryo

    PubMed Central

    Mugford, Joshua W.; Sipilä, Petra; Kobayashi, Akio; Behringer, Richard R.; McMahon, Andrew P.

    2008-01-01

    The mammalian kidney consists of an array of tubules connected to a ductal system that collectively function to control water/salt balance and to remove waste from the organisms’ circulatory system. During mammalian embryogenesis, three kidney structures form within the intermediate mesoderm. The two most anterior structures, the pronephros and the mesonephros, are transitory and largely non-functional, while the most posterior, the metanephros, persists as the adult kidney. We have explored the mechanisms underlying regional specific differentiation of the kidney forming mesoderm. Previous studies have shown a requirement for Hox11 paralogs (Hoxa11, Hoxc11 and Hoxd11) in metanephric development. Mice lacking all Hox11 activity fail to form metanephric kidney structures. We demonstrate that the Hox11 paralog expression is restricted in the intermediate mesoderm to the posterior, metanephric level. When Hoxd11 is ectopically activated in the anterior mesonephros, we observe a partial transformation to a metanephric program of development. Anterior Hoxd11 + cells activate Six2, a transcription factor required for the maintenance of metanephric tubule progenitors. Additionally, Hoxd11 + mesonephric tubules exhibit an altered morphology and activate several metanephric specific markers normally confined to distal portions of the functional nephron. Collectively, our data support a model where Hox11 paralogs specify a metanephric developmental program in responsive intermediate mesoderm. This program maintains tubule forming progenitors and instructs a metanephric specific pattern of nephron differentiation. PMID:18485340

  6. Commitment of the teratocarcinoma-derived mesodermal clone C1 towards terminal osteogenic differentiation.

    PubMed

    Poliard, A; Lamblin, D; Marie, P J; Buc-Caron, M H; Kellermann, O

    1993-10-01

    The mesodermal clone C1 was derived from the multipotent embryonal carcinoma 1003 cell line transformed with the plasmid pK4 carrying SV40 oncogenes under the control of the adenovirus E1A promoter. We have shown that the C1 clone becomes committed to the osteogenic pathway when cultured in aggregates in the presence of mediators of the osteogenic differentiation. To further validate C1 as a model with which to study osteogenesis in vitro the kinetics of its differentiation was studied, focusing on the histology of the aggregates and on the expression of a set of genes corresponding to representative bone matrix proteins. The presence of ascorbic acid and beta- glycerophosphate specifically leads to mineralization in almost 100% of the aggregates. Transcription of the above genes, silent in exponentially growing cells, specifically occurred with the establishment of cell-cell contacts independently of the presence of ascorbic acid and inorganic phosphate. The latter, however, were absolutely required for matrix deposition and mineralization. In their presence, one observed an overall decline in type I collagen and alkaline phosphatase transcripts while osteocalcin and osteopontin transcripts preferentially accumulated in cells lining the mineralizing foci. Concomitantly, type I collagen and osteocalcin became extracellularly deposited. The osteogenic differentiation of C1 occurred while cells were still proliferating. The C1 clone thus behaves as a mesodermal stem cell, becoming committed to the osteogenic pathway upon: firstly, establishment of cellular contacts; and secondly, addition of ascorbate and beta-glycerophosphate. It therefore appears to be a promising in vitro system for deciphering the molecular basis of osteoblast ontogeny.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:8282757

  7. ProNodal acts via FGFR3 to govern duration of Shh expression in the prechordal mesoderm.

    PubMed

    Ellis, Pamela S; Burbridge, Sarah; Soubes, Sandrine; Ohyama, Kyoji; Ben-Haim, Nadav; Chen, Canhe; Dale, Kim; Shen, Michael M; Constam, Daniel; Placzek, Marysia

    2015-11-15

    The secreted glycoprotein sonic hedgehog (Shh) is expressed in the prechordal mesoderm, where it plays a crucial role in induction and patterning of the ventral forebrain. Currently little is known about how Shh is regulated in prechordal tissue. Here we show that in the embryonic chick, Shh is expressed transiently in prechordal mesoderm, and is governed by unprocessed Nodal. Exposure of prechordal mesoderm microcultures to Nodal-conditioned medium, the Nodal inhibitor CerS, or to an ALK4/5/7 inhibitor reveals that Nodal is required to maintain both Shh and Gsc expression, but whereas Gsc is largely maintained through canonical signalling, Nodal signals through a non-canonical route to maintain Shh. Further, Shh expression can be maintained by a recombinant Nodal cleavage mutant, proNodal, but not by purified mature Nodal. A number of lines of evidence suggest that proNodal acts via FGFR3. ProNodal and FGFR3 co-immunoprecipitate and proNodal increases FGFR3 tyrosine phosphorylation. In microcultures, soluble FGFR3 abolishes Shh without affecting Gsc expression. Further, prechordal mesoderm cells in which Fgfr3 expression is reduced by Fgfr3 siRNA fail to bind to proNodal. Finally, targeted electroporation of Fgfr3 siRNA to prechordal mesoderm in vivo results in premature Shh downregulation without affecting Gsc. We report an inverse correlation between proNodal-FGFR3 signalling and pSmad1/5/8, and show that proNodal-FGFR3 signalling antagonises BMP-mediated pSmad1/5/8 signalling, which is poised to downregulate Shh. Our studies suggest that proNodal/FGFR3 signalling governs Shh duration by repressing canonical BMP signalling, and that local BMPs rapidly silence Shh once endogenous Nodal-FGFR3 signalling is downregulated. PMID:26417042

  8. ProNodal acts via FGFR3 to govern duration of Shh expression in the prechordal mesoderm

    PubMed Central

    Ellis, Pamela S.; Burbridge, Sarah; Soubes, Sandrine; Ohyama, Kyoji; Ben-Haim, Nadav; Chen, Canhe; Dale, Kim; Shen, Michael M.; Constam, Daniel; Placzek, Marysia

    2015-01-01

    The secreted glycoprotein sonic hedgehog (Shh) is expressed in the prechordal mesoderm, where it plays a crucial role in induction and patterning of the ventral forebrain. Currently little is known about how Shh is regulated in prechordal tissue. Here we show that in the embryonic chick, Shh is expressed transiently in prechordal mesoderm, and is governed by unprocessed Nodal. Exposure of prechordal mesoderm microcultures to Nodal-conditioned medium, the Nodal inhibitor CerS, or to an ALK4/5/7 inhibitor reveals that Nodal is required to maintain both Shh and Gsc expression, but whereas Gsc is largely maintained through canonical signalling, Nodal signals through a non-canonical route to maintain Shh. Further, Shh expression can be maintained by a recombinant Nodal cleavage mutant, proNodal, but not by purified mature Nodal. A number of lines of evidence suggest that proNodal acts via FGFR3. ProNodal and FGFR3 co-immunoprecipitate and proNodal increases FGFR3 tyrosine phosphorylation. In microcultures, soluble FGFR3 abolishes Shh without affecting Gsc expression. Further, prechordal mesoderm cells in which Fgfr3 expression is reduced by Fgfr3 siRNA fail to bind to proNodal. Finally, targeted electroporation of Fgfr3 siRNA to prechordal mesoderm in vivo results in premature Shh downregulation without affecting Gsc. We report an inverse correlation between proNodal-FGFR3 signalling and pSmad1/5/8, and show that proNodal-FGFR3 signalling antagonises BMP-mediated pSmad1/5/8 signalling, which is poised to downregulate Shh. Our studies suggest that proNodal/FGFR3 signalling governs Shh duration by repressing canonical BMP signalling, and that local BMPs rapidly silence Shh once endogenous Nodal-FGFR3 signalling is downregulated. PMID:26417042

  9. Receptor tyrosine phosphatase psi is required for Delta/Notch signalling and cyclic gene expression in the presomitic mesoderm.

    PubMed

    Aerne, Birgit; Ish-Horowicz, David

    2004-07-01

    Segmentation in vertebrate embryos is controlled by a biochemical oscillator ('segmentation clock') intrinsic to the cells in the unsegmented presomitic mesoderm, and is manifested in cyclic transcription of genes involved in establishing somite polarity and boundaries. We show that the receptor protein tyrosine phosphatase psi (RPTPpsi) gene is essential for normal functioning of the somitogenesis clock in zebrafish. We show that reduction of RPTPpsi activity using morpholino antisense oligonucleotides results in severe disruption of the segmental pattern of the embryo, and loss of cyclic gene expression in the presomitic mesoderm. Analysis of cyclic genes in RPTPpsi morphant embryos indicates an important requirement for RPTPpsi in the control of the somitogenesis clock upstream of or in parallel with Delta/Notch signalling. Impairing RPTPpsi activity also interferes with convergent extension during gastrulation. We discuss this dual requirement for RPTPpsi in terms of potential functions in Notch and Wnt signalling. PMID:15226256

  10. Midline-derived Shh regulates mesonephric tubule formation through the paraxial mesoderm

    PubMed Central

    Murashima, Aki; Akita, Hiroki; Okazawa, Mika; Kishigami, Satoshi; Nakagata, Naomi; Nishinakamura, Ryuichi; Yamada, Gen

    2014-01-01

    During organogenesis, Sonic hedgehog (Shh) possesses dual functions: Shh emanating from midline structures regulates the positioning of bilateral structures at early stages, whereas organ-specific Shh locally regulates organ morphogenesis at later stages. The mesonephros is a transient embryonic kidney in amniote, whereas it becomes definitive adult kidney in some anamniotes. Thus, elucidating the regulation of mesonephros formation has important implications for our understanding of kidney development and evolution. In Shh knockout (KO) mutant mice, the mesonephros was displaced towards the midline and ectopic mesonephric tubules (MTs) were present in the caudal mesonephros. Mesonephros-specific ablation of Shh in Hoxb7-Cre;Shhflox/− and Sall1CreERT2/+;Shhflox/− mice embryos indicated that Shh expressed in the mesonephros was not required for either the development of the mesonephros or the differentiation of the male reproductive tract. Moreover, stage-specific ablation of Shh in ShhCreERT2/flox mice showed that notochord- and/or floor plate-derived Shh were essential for the regulation of the number and position of MTs. Lineage analysis of hedgehog (Hh)-responsive cells, and analysis of gene expression in Shh KO embryos suggested that Shh regulated nephrogenic gene expression indirectly, possibly through effects on the paraxial mesoderm. These data demonstrate the essential role of midline-derived Shh in local tissue morphogenesis and differentiation. PMID:24370450

  11. Scl binds to primed enhancers in mesoderm to regulate hematopoietic and cardiac fate divergence.

    PubMed

    Org, Tõnis; Duan, Dan; Ferrari, Roberto; Montel-Hagen, Amelie; Van Handel, Ben; Kerényi, Marc A; Sasidharan, Rajkumar; Rubbi, Liudmilla; Fujiwara, Yuko; Pellegrini, Matteo; Orkin, Stuart H; Kurdistani, Siavash K; Mikkola, Hanna Ka

    2015-03-12

    Scl/Tal1 confers hemogenic competence and prevents ectopic cardiomyogenesis in embryonic endothelium by unknown mechanisms. We discovered that Scl binds to hematopoietic and cardiac enhancers that become epigenetically primed in multipotent cardiovascular mesoderm, to regulate the divergence of hematopoietic and cardiac lineages. Scl does not act as a pioneer factor but rather exploits a pre-established epigenetic landscape. As the blood lineage emerges, Scl binding and active epigenetic modifications are sustained in hematopoietic enhancers, whereas cardiac enhancers are decommissioned by removal of active epigenetic marks. Our data suggest that, rather than recruiting corepressors to enhancers, Scl prevents ectopic cardiogenesis by occupying enhancers that cardiac factors, such as Gata4 and Hand1, use for gene activation. Although hematopoietic Gata factors bind with Scl to both activated and repressed genes, they are dispensable for cardiac repression, but necessary for activating genes that enable hematopoietic stem/progenitor cell development. These results suggest that a unique subset of enhancers in lineage-specific genes that are accessible for regulators of opposing fates during the time of the fate decision provide a platform where the divergence of mutually exclusive fates is orchestrated. PMID:25564442

  12. A Wide Spectrum of Axial Mesodermal Dysplasia Complex With Rhombencephalic Anomaly: A Case Report

    PubMed Central

    Kim, Kang-Won; Seo, Jeoung-Hwan; Ko, Myoung-Hwan; Won, Yu-Hui

    2016-01-01

    Axial mesodermal dysplasia complex (AMDC) arises in variable combinations of craniocaudal anomalies such as musculoskeletal deformities, neuroschisis, or rhombencephalic developmental disorders. To the best of our knowledge, the co-existence of AMDC with associated musculoskeletal anomalies, medullary neuroschisis with mirror movements, and cranial nerve anomalies has not yet been reported. Here, we report the case of a 4-year-old boy whose clinical features were suggestive of Goldenhar syndrome and Poland syndrome with Sprengel deformity. Moreover, he showed mirror movements in his hands suspected of rhombencephalic malformation, and infranuclear-type facial nerve palsy of the left side of his face, the opposite side to the facial anomalies of Goldenhar syndrome. After conducting radiological studies, he was diagnosed with medullary neuroschisis without pontine malformations and Klippel-Feil syndrome with rib anomalies. Based on these findings, we propose that clinical AMDC can be accompanied by a wide variety of musculoskeletal defects and variable degrees of central nervous system malformations. Therefore, in addition to detailed physical and neurological examinations, imaging studies should be considered in AMDC. PMID:26949683

  13. Transcription factor KLF7 regulates differentiation of neuroectodermal and mesodermal cell lineages

    SciTech Connect

    Caiazzo, Massimiliano; Colucci-D'Amato, Luca; Esposito, Maria T.; Parisi, Silvia; Stifani, Stefano; Ramirez, Francesco; Porzio, Umberto di

    2010-08-15

    Previous gene targeting studies in mice have implicated the nuclear protein Krueppel-like factor 7 (KLF7) in nervous system development while cell culture assays have documented its involvement in cell cycle regulation. By employing short hairpin RNA (shRNA)-mediated gene silencing, here we demonstrate that murine Klf7 gene expression is required for in vitro differentiation of neuroectodermal and mesodermal cells. Specifically, we show a correlation of Klf7 silencing with down-regulation of the neuronal marker microtubule-associated protein 2 (Map2) and the nerve growth factor (NGF) tyrosine kinase receptor A (TrkA) using the PC12 neuronal cell line. Similarly, KLF7 inactivation in Klf7-null mice decreases the expression of the neurogenic marker brain lipid-binding protein/fatty acid-binding protein 7 (BLBP/FABP7) in neural stem cells (NSCs). We also report that Klf7 silencing is detrimental to neuronal and cardiomyocytic differentiation of embryonic stem cells (ESCs), in addition to altering the adipogenic and osteogenic potential of mouse embryonic fibroblasts (MEFs). Finally, our results suggest that genes that are key for self-renewal of undifferentiated ESCs repress Klf7 expression in ESCs. Together with previous findings, these results provide evidence that KLF7 has a broad spectrum of regulatory functions, which reflect the discrete cellular and molecular contexts in which this transcription factor operates.

  14. The mesoderm determinant snail collaborates with related zinc-finger proteins to control Drosophila neurogenesis.

    PubMed

    Ashraf, S I; Hu, X; Roote, J; Ip, Y T

    1999-11-15

    The Snail protein functions as a transcriptional regulator to establish early mesodermal cell fate. Later, in germ band-extended embryos, Snail is also expressed in most neuroblasts. Here we present evidence that this expression of Snail is required for central nervous system (CNS) development. The neural function of snail is masked by two closely linked genes, escargot and worniu. Both Escargot and Worniu contain zinc-finger domains that are highly homologous to that of Snail. Although not affecting expression of early neuroblast markers, the deletion of the region containing all three genes correlates with loss of expression of CNS determinants including fushi tarazu, pdm-2 and even-skipped. Transgenic expression of each of the three Snail family proteins can rescue efficiently the fushi tarazu defects, and partially the pdm-2 and even-skipped CNS patterns. These results demonstrate that the Snail family proteins have essential functions during embryonic CNS development, around the time of ganglion mother cell formation. PMID:10562554

  15. Mesodermal iPSC–derived progenitor cells functionally regenerate cardiac and skeletal muscle

    PubMed Central

    Quattrocelli, Mattia; Swinnen, Melissa; Giacomazzi, Giorgia; Camps, Jordi; Barthélemy, Ines; Ceccarelli, Gabriele; Caluwé, Ellen; Grosemans, Hanne; Thorrez, Lieven; Pelizzo, Gloria; Muijtjens, Manja; Verfaillie, Catherine M.; Blot, Stephane; Janssens, Stefan; Sampaolesi, Maurilio

    2015-01-01

    Conditions such as muscular dystrophies (MDs) that affect both cardiac and skeletal muscles would benefit from therapeutic strategies that enable regeneration of both of these striated muscle types. Protocols have been developed to promote induced pluripotent stem cells (iPSCs) to differentiate toward cardiac or skeletal muscle; however, there are currently no strategies to simultaneously target both muscle types. Tissues exhibit specific epigenetic alterations; therefore, source-related lineage biases have the potential to improve iPSC-driven multilineage differentiation. Here, we determined that differential myogenic propensity influences the commitment of isogenic iPSCs and a specifically isolated pool of mesodermal iPSC-derived progenitors (MiPs) toward the striated muscle lineages. Differential myogenic propensity did not influence pluripotency, but did selectively enhance chimerism of MiP-derived tissue in both fetal and adult skeletal muscle. When injected into dystrophic mice, MiPs engrafted and repaired both skeletal and cardiac muscle, reducing functional defects. Similarly, engraftment into dystrophic mice of canine MiPs from dystrophic dogs that had undergone TALEN-mediated correction of the MD-associated mutation also resulted in functional striatal muscle regeneration. Moreover, human MiPs exhibited the same capacity for the dual differentiation observed in murine and canine MiPs. The findings of this study suggest that MiPs should be further explored for combined therapy of cardiac and skeletal muscles. PMID:26571398

  16. Pronephric tubule morphogenesis in zebrafish depends on Mnx mediated repression of irx1b within the intermediate mesoderm.

    PubMed

    Ott, Elisabeth; Wendik, Björn; Srivastava, Monika; Pacho, Frederic; Töchterle, Sonja; Salvenmoser, Willi; Meyer, Dirk

    2016-03-01

    Mutations in the homeobox transcription factor MNX1 are the major cause of dominantly inherited sacral agenesis. Studies in model organisms revealed conserved mnx gene requirements in neuronal and pancreatic development while Mnx activities that could explain the caudal mesoderm specific agenesis phenotype remain elusive. Here we use the zebrafish pronephros as a simple yet genetically conserved model for kidney formation to uncover a novel role of Mnx factors in nephron morphogenesis. Pronephros formation can formally be divided in four stages, the specification of nephric mesoderm from the intermediate mesoderm (IM), growth and epithelialisation, segmentation and formation of the glomerular capillary tuft. Two of the three mnx genes in zebrafish are dynamically transcribed in caudal IM in a time window that proceeds segmentation. We show that expression of one mnx gene, mnx2b, is restricted to the pronephric lineage and that mnx2b knock-down causes proximal pronephric tubule dilation and impaired pronephric excretion. Using expression profiling of embryos transgenic for conditional activation and repression of Mnx regulated genes, we further identified irx1b as a direct target of Mnx factors. Consistent with a repression of irx1b by Mnx factors, the transcripts of irx1b and mnx genes are found in mutual exclusive regions in the IM, and blocking of Mnx functions results in a caudal expansion of the IM-specific irx1b expression. Finally, we find that knock-down of irx1b is sufficient to rescue proximal pronephric tubule dilation and impaired nephron function in mnx-morpholino injected embryos. Our data revealed a first caudal mesoderm specific requirement of Mnx factors in a non-human system and they demonstrate that Mnx-dependent restriction of IM-specific irx1b activation is required for the morphogenesis and function of the zebrafish pronephros. PMID:26472045

  17. Transplacental delivery of the Wnt antagonist Frzb1 inhibits development of caudal paraxial mesoderm and skeletal myogenesis in mouse embryos.

    PubMed

    Borello, U; Coletta, M; Tajbakhsh, S; Leyns, L; De Robertis, E M; Buckingham, M; Cossu, G

    1999-10-01

    Axial structures (neural tube/notochord) and surface ectoderm activate myogenesis in the mouse embryo; their action can be reproduced, at least in part, by several molecules such as Sonic hedgehog and Wnts. Recently, soluble Wnt antagonists have been identified. Among those examined only Frzb1 was found to be expressed in the presomitic mesoderm and newly formed somites and thus its possible role in regulating myogenesis was investigated in detail. When presomitic mesoderm or newly formed somites were cultured with axial structures and surface ectoderm on a feeder layer of C3H10T1/2 cells expressing Frzb1, myogenesis was abolished or severely reduced in presomitic mesoderm and the three most recently formed somites. In contrast, no effect was observed on more mature somites. Inhibition of myogenesis did not appear to be associated with increased cell death since the final number of cells in the explants grown in the presence of Frzb1 was only slightly reduced in comparison with controls. In order to examine the possible function of Frzb1 in vivo, we developed a method based on the overexpression of the soluble antagonist by transient transfection of WOP cells with a Frzb1 expression vector and injection of transfected cells into the placenta of pregnant females before the onset of maternofoetal circulation. Frzb1, secreted by WOP cells, accumulated in the embryo and caused a marked reduction in size of caudal structures. Myogenesis was strongly reduced and, in the most severe cases, abolished. This was not due to a generalized toxic effect since only several genes downstream of the Wnt signaling pathway such as En1, Noggin and Myf5 were downregulated; in contrast, Pax3 and Mox1 expression levels were not affected even in embryos exhibiting the most severe phenotypes. Taken together, these results suggest that Wnt signals may act by regulating both myogenic commitment and expansion of committed cells in the mouse mesoderm. PMID:10477293

  18. Chordin forms a self-organizing morphogen gradient in the extracellular space between ectoderm and mesoderm in the Xenopus embryo

    PubMed Central

    Plouhinec, Jean-Louis; Zakin, Lise; Moriyama, Yuki; De Robertis, Edward M.

    2013-01-01

    The vertebrate body plan follows stereotypical dorsal–ventral (D-V) tissue differentiation controlled by bone morphogenetic proteins (BMPs) and secreted BMP antagonists, such as Chordin. The three germ layers—ectoderm, mesoderm, and endoderm—are affected coordinately by the Chordin–BMP morphogen system. However, extracellular morphogen gradients of endogenous proteins have not been directly visualized in vertebrate embryos to date. In this study, we improved immunolocalization methods in Xenopus embryos and analyzed the distribution of endogenous Chordin using a specific antibody. Chordin protein secreted by the dorsal Spemann organizer was found to diffuse along a narrow region that separates the ectoderm from the anterior endoderm and mesoderm. This Fibronectin-rich extracellular matrix is called “Brachet’s cleft” in the Xenopus gastrula and is present in all vertebrate embryos. Chordin protein formed a smooth gradient that encircled the embryo, reaching the ventral-most Brachet cleft. Depletion with morpholino oligos showed that this extracellular gradient was regulated by the Chordin protease Tolloid and its inhibitor Sizzled. The Chordin gradient, as well as the BMP signaling gradient, was self-regulating and, importantly, was able to rescale in dorsal half-embryos. Transplantation of Spemann organizer tissue showed that Chordin diffused over long distances along this signaling highway between the ectoderm and mesoderm. Chordin protein must reach very high concentrations in this narrow region. We suggest that as ectoderm and mesoderm undergo morphogenetic movements during gastrulation, cells in both germ layers read their positional information coordinately from a single morphogen gradient located in Brachet’s cleft. PMID:24284174

  19. A conserved Six-Eya cassette acts downstream of Wnt signaling to direct non-myogenic versus myogenic fates in the C. elegans postembryonic mesoderm.

    PubMed

    Amin, Nirav M; Lim, Sung-Eun; Shi, Herong; Chan, Tiffany L; Liu, Jun

    2009-07-15

    The subdivision of mesodermal cells into muscle and non-muscle cells is crucial to animal development. In the C. elegans postembryonic mesoderm, this subdivision is a result of an asymmetric cell division that leads to the formation of striated body wall muscles and non-muscle coelomocytes. Here we report that the Six homeodomain protein CEH-34 and its cofactor Eyes Absent, EYA-1, function synergistically to promote the non-muscle fate in cells also competent to form muscles. We further show that the asymmetric expression of ceh-34 and eya-1 is regulated by a combination of 1) mesodermal intrinsic factors MAB-5, HLH-1 and FOZI-1, 2) differential POP-1 (TCF/LEF) transcriptional activity along the anterior-posterior axis, and 3) coelomocyte competence factor(s). These factors are conserved in both vertebrates and invertebrates, suggesting a conserved paradigm for mesoderm development in metazoans. PMID:19427847

  20. Knockout of the PKN Family of Rho Effector Kinases Reveals a Non-redundant Role for PKN2 in Developmental Mesoderm Expansion.

    PubMed

    Quétier, Ivan; Marshall, Jacqueline J T; Spencer-Dene, Bradley; Lachmann, Sylvie; Casamassima, Adele; Franco, Claudio; Escuin, Sarah; Worrall, Joseph T; Baskaran, Priththivika; Rajeeve, Vinothini; Howell, Michael; Copp, Andrew J; Stamp, Gordon; Rosewell, Ian; Cutillas, Pedro; Gerhardt, Holger; Parker, Peter J; Cameron, Angus J M

    2016-01-26

    In animals, the protein kinase C (PKC) family has expanded into diversely regulated subgroups, including the Rho family-responsive PKN kinases. Here, we describe knockouts of all three mouse PKN isoforms and reveal that PKN2 loss results in lethality at embryonic day 10 (E10), with associated cardiovascular and morphogenetic defects. The cardiovascular phenotype was not recapitulated by conditional deletion of PKN2 in endothelial cells or the developing heart. In contrast, inducible systemic deletion of PKN2 after E7 provoked collapse of the embryonic mesoderm. Furthermore, mouse embryonic fibroblasts, which arise from the embryonic mesoderm, depend on PKN2 for proliferation and motility. These cellular defects are reflected in vivo as dependence on PKN2 for mesoderm proliferation and neural crest migration. We conclude that failure of the mesoderm to expand in the absence of PKN2 compromises cardiovascular integrity and development, resulting in lethality. PMID:26774483

  1. Knockout of the PKN Family of Rho Effector Kinases Reveals a Non-redundant Role for PKN2 in Developmental Mesoderm Expansion

    PubMed Central

    Quétier, Ivan; Marshall, Jacqueline J.T.; Spencer-Dene, Bradley; Lachmann, Sylvie; Casamassima, Adele; Franco, Claudio; Escuin, Sarah; Worrall, Joseph T.; Baskaran, Priththivika; Rajeeve, Vinothini; Howell, Michael; Copp, Andrew J.; Stamp, Gordon; Rosewell, Ian; Cutillas, Pedro; Gerhardt, Holger; Parker, Peter J.; Cameron, Angus J.M.

    2016-01-01

    Summary In animals, the protein kinase C (PKC) family has expanded into diversely regulated subgroups, including the Rho family-responsive PKN kinases. Here, we describe knockouts of all three mouse PKN isoforms and reveal that PKN2 loss results in lethality at embryonic day 10 (E10), with associated cardiovascular and morphogenetic defects. The cardiovascular phenotype was not recapitulated by conditional deletion of PKN2 in endothelial cells or the developing heart. In contrast, inducible systemic deletion of PKN2 after E7 provoked collapse of the embryonic mesoderm. Furthermore, mouse embryonic fibroblasts, which arise from the embryonic mesoderm, depend on PKN2 for proliferation and motility. These cellular defects are reflected in vivo as dependence on PKN2 for mesoderm proliferation and neural crest migration. We conclude that failure of the mesoderm to expand in the absence of PKN2 compromises cardiovascular integrity and development, resulting in lethality. PMID:26774483

  2. HoxBlinc RNA Recruits Set1/MLL Complexes to Activate Hox Gene Expression Patterns and Mesoderm Lineage Development.

    PubMed

    Deng, Changwang; Li, Ying; Zhou, Lei; Cho, Joonseok; Patel, Bhavita; Terada, Naohiro; Li, Yangqiu; Bungert, Jörg; Qiu, Yi; Huang, Suming

    2016-01-01

    Trithorax proteins and long-intergenic noncoding RNAs are critical regulators of embryonic stem cell pluripotency; however, how they cooperatively regulate germ layer mesoderm specification remains elusive. We report here that HoxBlinc RNA first specifies Flk1(+) mesoderm and then promotes hematopoietic differentiation through regulation of hoxb pathways. HoxBlinc binds to the hoxb genes, recruits Setd1a/MLL1 complexes, and mediates long-range chromatin interactions to activate transcription of the hoxb genes. Depletion of HoxBlinc by shRNA-mediated knockdown or CRISPR-Cas9-mediated genetic deletion inhibits expression of hoxb genes and other factors regulating cardiac/hematopoietic differentiation. Reduced hoxb expression is accompanied by decreased recruitment of Set1/MLL1 and H3K4me3 modification, as well as by reduced chromatin loop formation. Re-expression of hoxb2-b4 genes in HoxBlinc-depleted embryoid bodies rescues Flk1(+) precursors that undergo hematopoietic differentiation. Thus, HoxBlinc plays an important role in controlling hoxb transcription networks that mediate specification of mesoderm-derived Flk1(+) precursors and differentiation of Flk1(+) cells into hematopoietic lineages. PMID:26725110

  3. The transforming growth factor beta type II receptor can replace the activin type II receptor in inducing mesoderm.

    PubMed Central

    Bhushan, A; Lin, H Y; Lodish, H F; Kintner, C R

    1994-01-01

    The type II receptors for the polypeptide growth factors transforming growth factor beta (TGF-beta) and activin belong to a new family of predicted serine/threonine protein kinases. In Xenopus embryos, the biological effects of activin and TGF-beta 1 are strikingly different; activin induces a full range of mesodermal cell types in the animal cap assay, while TGF-beta 1 has no effects, presumably because of the lack of functional TGF-beta receptors. In order to assess the biological activities of exogenously added TGF-beta 1, RNA encoding the TGF-beta type II receptor was introduced into Xenopus embryos. In animal caps from these embryos, TGF-beta 1 and activin show similar potencies for induction of mesoderm-specific mRNAs, and both elicit the same types of mesodermal tissues. In addition, the response of animal caps to TGF-beta 1, as well as to activin, is blocked by a dominant inhibitory ras mutant, p21(Asn-17)Ha-ras. These results indicate that the activin and TGF-beta type II receptors can couple to similar signalling pathways and that the biological specificities of these growth factors lie in their different ligand-binding domains and in different competences of the responding cells. Images PMID:8196664

  4. HoxBlinc RNA recruits Set1/MLL complexes to activate Hox gene expression patterns and mesoderm lineage development

    PubMed Central

    Deng, Changwang; Li, Ying; Zhou, Lei; Cho, Joonseok; Patel, Bhavita; Terada, Nao; Li, Yangqiu; Bungert, Jörg; Qiu, Yi; Huang, Suming

    2015-01-01

    Summary Trithorax proteins and long-intergenic noncoding RNAs are critical regulators of embryonic stem cell pluripotency; however, how they cooperatively regulate germ layer mesoderm specification remains elusive. We report here that HoxBlinc RNA first specifies Flk1+ mesoderm and then promotes hematopoietic differentiation through regulating hoxb gene pathways. HoxBlinc binds to the hoxb genes, recruits Setd1a/MLL1 complexes, and mediates long-range chromatin interactions to activate transcription of the hoxb genes. Depletion of HoxBlinc by shRNA-mediated KD or CRISPR-Cas9-mediated genetic deletion inhibits expression of hoxb genes and other factors regulating cardiac/hematopoietic differentiation. Reduced hoxb gene expression is accompanied by decreased recruitment of Set1/MLL1 and H3K4me3 modification, as well as by reduced chromatin loop formation. Re-expression of hoxb2-b4 genes in HoxBlinc-depleted embryoid bodies rescues Flk1+ precursors that undergo hematopoietic differentiation. Thus, HoxBlinc plays an important role in controlling hoxb transcription networks that mediate specification of mesoderm-derived Flk1+ precursors and differentiation of Flk1+ cells into hematopoietic lineages. PMID:26725110

  5. Hhex and Cer1 Mediate the Sox17 Pathway for Cardiac Mesoderm Formation in Embryonic Stem Cells

    PubMed Central

    Liu, Yu; Kaneda, Ruri; Leja, Thomas W; Subkhankulova, Tatiana; Tolmachov, Oleg; Minchiotti, Gabriella; Schwartz, Robert J; Barahona, Mauricio; Schneider, Michael D

    2014-01-01

    Abstract Cardiac muscle differentiation in vivo is guided by sequential growth factor signals, including endoderm-derived diffusible factors, impinging on cardiogenic genes in the developing mesoderm. Previously, by RNA interference in AB2.2 mouse embryonic stem cells (mESCs), we identified the endodermal transcription factor Sox17 as essential for Mesp1 induction in primitive mesoderm and subsequent cardiac muscle differentiation. However, downstream effectors of Sox17 remained to be proven functionally. In this study, we used genome-wide profiling of Sox17-dependent genes in AB2.2 cells, RNA interference, chromatin immunoprecipitation, and luciferase reporter genes to dissect this pathway. Sox17 was required not only for Hhex (a second endodermal transcription factor) but also for Cer1, a growth factor inhibitor from endoderm that, like Hhex, controls mesoderm patterning in Xenopus toward a cardiac fate. Suppressing Hhex or Cer1 blocked cardiac myogenesis, although at a later stage than induction of Mesp1/2. Hhex was required but not sufficient for Cer1 expression. Over-expression of Sox17 induced endogenous Cer1 and sequence-specific transcription of a Cer1 reporter gene. Forced expression of Cer1 was sufficient to rescue cardiac differentiation in Hhex-deficient cells. Thus, Hhex and Cer1 are indispensable components of the Sox17 pathway for cardiopoiesis in mESCs, acting at a stage downstream from Mesp1/2. Stem Cells 2014;32:1515–1526 PMID:24585688

  6. Mixl1 and Flk1 Are Key Players of Wnt/TGF-β Signaling During DMSO-Induced Mesodermal Specification in P19 cells.

    PubMed

    Choi, Seung-Cheol; Choi, Ji-Hyun; Cui, Long-Hui; Seo, Ha-Rim; Kim, Jong-Ho; Park, Chi-Yeon; Joo, Hyung-Joon; Park, Jae-Hyoung; Hong, Soon-Jun; Yu, Cheol-Woong; Lim, Do-Sun

    2015-08-01

    Dimethyl sulfoxide (DMSO) is widely used to induce multilineage differentiation of embryonic and adult progenitor cells. To date, little is known about the mechanisms underlying DMSO-induced mesodermal specification. In this study, we investigated the signaling pathways and lineage-determining genes involved in DMSO-induced mesodermal specification in P19 cells. Wnt/β-catenin and TGF-β superfamily signaling pathways such as BMP, TGF-β and GDF1 signaling were significantly activated during DMSO-induced mesodermal specification. In contrast, Nodal/Cripto signaling pathway molecules, required for endoderm specification, were severely downregulated. DMSO significantly upregulated the expression of cardiac mesoderm markers but inhibited the expression of endodermal and hematopoietic lineage markers. Among the DMSO-activated cell lineage markers, the expression of Mixl1 and Flk1 was dramatically upregulated at both the transcript and protein levels, and the populations of Mixl1+, Flk1+ and Mixl1+/Flk1+ cells also increased significantly. DMSO modulated cell cycle molecules and induced cell apoptosis, resulting in significant cell death during EB formation of P19 cells. An inhibitor of Flk1, SU5416 significantly blocked expressions of TGF-β superfamily members, mesodermal cell lineage markers and cell cycle molecules but it did not affect Wnt molecules. These results demonstrate that Mixl1 and Flk1 play roles as key downstream or interacting effectors of Wnt/TGF-β signaling pathway during DMSO-induced mesodermal specification in P19 cells. PMID:25521758

  7. Growth factor-mediated mesodermal cell guidance and skeletogenesis during sea urchin gastrulation.

    PubMed

    Adomako-Ankomah, Ashrifia; Ettensohn, Charles A

    2013-10-01

    Growth factor signaling pathways provide essential cues to mesoderm cells during gastrulation in many metazoans. Recent studies have implicated the VEGF and FGF pathways in providing guidance and differentiation cues to primary mesenchyme cells (PMCs) during sea urchin gastrulation, although the relative contributions of these pathways and the cell behaviors they regulate are not fully understood. Here, we show that FGF and VEGF ligands are expressed in distinct domains in the embryonic ectoderm of Lytechinus variegatus. We find that PMC guidance is specifically disrupted in Lv-vegf3 morphants and these embryos fail to form skeletal elements. By contrast, PMC migration is unaffected in Lv-fgfa morphants, and well-patterned but shortened skeletal elements form. We use a VEGFR inhibitor, axitinib, to show that VEGF signaling is essential not only for the initial phase of PMC migration (subequatorial ring formation), but also for the second phase (migration towards the animal pole). VEGF signaling is not required, however, for PMC fusion. Inhibition of VEGF signaling after the completion of PMC migration causes significant defects in skeletogenesis, selectively blocking the elongation of skeletal rods that support the larval arms, but not rods that form in the dorsal region of the embryo. Nanostring nCounter analysis of ∼100 genes in the PMC gene regulatory network shows a decrease in the expression of many genes with proven or predicted roles in biomineralization in vegf3 morphants. Our studies lead to a better understanding of the roles played by growth factors in sea urchin gastrulation and skeletogenesis. PMID:24026121

  8. Visualizing Late Insect Embryogenesis: Extraembryonic and Mesodermal Enhancer Trap Expression in the Beetle Tribolium castaneum

    PubMed Central

    Koelzer, Stefan; Kölsch, Yvonne; Panfilio, Kristen A.

    2014-01-01

    The beetle Tribolium castaneum has increasingly become a powerful model for comparative research on insect development. One recent resource is a collection of piggyBac transposon-based enhancer trap lines. Here, we provide a detailed analysis of three selected lines and demonstrate their value for investigations in the second half of embryogenesis, which has thus far lagged behind research on early stages. Two lines, G12424 and KT650, show enhanced green fluorescent protein (EGFP) expression throughout the extraembryonic serosal tissue and in a few discrete embryonic domains. Intriguingly, both lines show for the first time a degree of regionalization within the mature serosa. However, their expression profiles illuminate distinct aspects of serosal biology: G12424 tracks the tissue’s rapid maturation while KT650 expression likely reflects ongoing physiological processes. The third line, G04609, is stably expressed in mesodermal domains, including segmental muscles and the heart. Genomic mapping followed by in situ hybridization for genes near to the G04609 insertion site suggests that the transposon has trapped enhancer information for the Tribolium orthologue of midline (Tc-mid). Altogether, our analyses provide the first live imaging, long-term characterizations of enhancer traps from this collection. We show that EGFP expression is readily detected, including in heterozygote crosses that permit the simultaneous visualization of multiple tissue types. The tissue specificity provides live, endogenous marker gene expression at key developmental stages that are inaccessible for whole mount staining. Furthermore, the nonlocalized EGFP in these lines illuminates both the nucleus and cytoplasm, providing cellular resolution for morphogenesis research on processes such as dorsal closure and heart formation. In future work, identification of regulatory regions driving these enhancer traps will deepen our understanding of late developmental control, including in the

  9. From Dynamic Expression Patterns to Boundary Formation in the Presomitic Mesoderm

    PubMed Central

    Tiedemann, Hendrik B.; Schneltzer, Elida; Zeiser, Stefan; Hoesel, Bastian; Beckers, Johannes; Przemeck, Gerhard K. H.; de Angelis, Martin Hrabě

    2012-01-01

    The segmentation of the vertebrate body is laid down during early embryogenesis. The formation of signaling gradients, the periodic expression of genes of the Notch-, Fgf- and Wnt-pathways and their interplay in the unsegmented presomitic mesoderm (PSM) precedes the rhythmic budding of nascent somites at its anterior end, which later develops into epithelialized structures, the somites. Although many in silico models describing partial aspects of somitogenesis already exist, simulations of a complete causal chain from gene expression in the growth zone via the interaction of multiple cells to segmentation are rare. Here, we present an enhanced gene regulatory network (GRN) for mice in a simulation program that models the growing PSM by many virtual cells and integrates WNT3A and FGF8 gradient formation, periodic gene expression and Delta/Notch signaling. Assuming Hes7 as core of the somitogenesis clock and LFNG as modulator, we postulate a negative feedback of HES7 on Dll1 leading to an oscillating Dll1 expression as seen in vivo. Furthermore, we are able to simulate the experimentally observed wave of activated NOTCH (NICD) as a result of the interactions in the GRN. We esteem our model as robust for a wide range of parameter values with the Hes7 mRNA and protein decays exerting a strong influence on the core oscillator. Moreover, our model predicts interference between Hes1 and HES7 oscillators when their intrinsic frequencies differ. In conclusion, we have built a comprehensive model of somitogenesis with HES7 as core oscillator that is able to reproduce many experimentally observed data in mice. PMID:22761566

  10. Non-canonical Wnt5a/Ror2 signaling regulates kidney morphogenesis by controlling intermediate mesoderm extension.

    PubMed

    Yun, Kangsun; Ajima, Rieko; Sharma, Nirmala; Costantini, Frank; Mackem, Susan; Lewandoski, Mark; Yamaguchi, Terry P; Perantoni, Alan O

    2014-12-20

    Congenital anomalies of the kidney and urinary tract (CAKUT) affect about 1 in 500 births and are a major cause of morbidity in infants. Duplex collecting systems rank among the most common abnormalities of CAKUT, but the molecular basis for this defect is poorly understood. In mice, conditional deletion of Wnt5a in mesoderm results in bilateral duplex kidney and ureter formation. The ureteric buds (UBs) in mutants emerge as doublets from the intermediate mesoderm (IM)-derived nephric duct (ND) without anterior expansion of the glial cell line-derived neurotrophic factor (Gdnf) expression domain in the surrounding mesenchyme. Wnt5a is normally expressed in a graded manner at the posterior end of the IM, but its expression is down-regulated prior to UB outgrowth at E10.5. Furthermore, ablation of Wnt5a in the mesoderm with an inducible Cre at E7.5 results in duplex UBs, whereas ablation at E8.5 yields normal UB outgrowth, demonstrating that Wnt5a functions in IM development well before the formation of the metanephros. In mutants, the posterior ND is duplicated and surrounding Pax2-positive mesenchymal cells persist in the nephric cord, suggesting that disruption of normal ND patterning prompts the formation of duplex ureters and kidneys. Ror2 homozygous mutants, which infrequently yield duplex collecting systems, show a dramatic increase in incidence with the additional deletion of one copy of Wnt5a, implicating this receptor in non-canonical Wnt5a signaling during IM development. This work provides the first evidence of a role of Wnt5a/Ror2 signaling in IM extension and offers new insights into the etiology of CAKUT and possible involvement of Wnt5a/Ror2 mutations. PMID:25082826

  11. BMP and retinoic acid regulate anterior–posterior patterning of the non-axial mesoderm across the dorsal–ventral axis

    PubMed Central

    Naylor, Richard W.; Skvarca, Lauren Brilli; Thisse, Christine; Thisse, Bernard; Hukriede, Neil A.; Davidson, Alan J.

    2016-01-01

    Despite the fundamental importance of patterning along the dorsal–ventral (DV) and anterior–posterior (AP) axes during embryogenesis, uncertainty exists in the orientation of these axes for the mesoderm. Here we examine the origin and formation of the zebrafish kidney, a ventrolateral mesoderm derivative, and show that AP patterning of the non-axial mesoderm occurs across the classic gastrula stage DV axis while DV patterning aligns along the animal–vegetal pole. We find that BMP signalling acts early to establish broad anterior and posterior territories in the non-axial mesoderm while retinoic acid (RA) functions later, but also across the classic DV axis. Our data support a model in which RA on the dorsal side of the embryo induces anterior kidney fates while posterior kidney progenitors are protected ventrally by the RA-catabolizing enzyme Cyp26a1. This work clarifies our understanding of vertebrate axis orientation and establishes a new paradigm for how the kidney and other mesodermal derivatives arise during embryogenesis. PMID:27406002

  12. BMP and retinoic acid regulate anterior-posterior patterning of the non-axial mesoderm across the dorsal-ventral axis.

    PubMed

    Naylor, Richard W; Skvarca, Lauren Brilli; Thisse, Christine; Thisse, Bernard; Hukriede, Neil A; Davidson, Alan J

    2016-01-01

    Despite the fundamental importance of patterning along the dorsal-ventral (DV) and anterior-posterior (AP) axes during embryogenesis, uncertainty exists in the orientation of these axes for the mesoderm. Here we examine the origin and formation of the zebrafish kidney, a ventrolateral mesoderm derivative, and show that AP patterning of the non-axial mesoderm occurs across the classic gastrula stage DV axis while DV patterning aligns along the animal-vegetal pole. We find that BMP signalling acts early to establish broad anterior and posterior territories in the non-axial mesoderm while retinoic acid (RA) functions later, but also across the classic DV axis. Our data support a model in which RA on the dorsal side of the embryo induces anterior kidney fates while posterior kidney progenitors are protected ventrally by the RA-catabolizing enzyme Cyp26a1. This work clarifies our understanding of vertebrate axis orientation and establishes a new paradigm for how the kidney and other mesodermal derivatives arise during embryogenesis. PMID:27406002

  13. An amphioxus nodal gene (AmphiNodal) with early symmetrical expression in the organizer and mesoderm and later asymmetrical expression associated with left-right axis formation

    NASA Technical Reports Server (NTRS)

    Yu, Jr-Kai; Holland, Linda Z.; Holland, Nicholas D.

    2002-01-01

    The full-length sequence and zygotic expression of an amphioxus nodal gene are described. Expression is first detected in the early gastrula just within the dorsal lip of the blastopore in a region of hypoblast that is probably comparable with the vertebrate Spemann's organizer. In the late gastrula and early neurula, expression remains bilaterally symmetrical, limited to paraxial mesoderm and immediately overlying regions of the neural plate. Later in the neurula stage, all neural expression disappears, and mesodermal expression disappears from the right side. All along the left side of the neurula, mesodermal expression spreads into the left side of the gut endoderm. Soon thereafter, all expression is down-regulated except near the anterior and posterior ends of the animal, where transcripts are still found in the mesoderm and endoderm on the left side. At this time, expression also begins in the ectoderm on the left side of the head, in the region where the mouth later forms. These results suggest that amphioxus and vertebrate nodal genes play evolutionarily conserved roles in establishing Spemann's organizer, patterning the mesoderm rostrocaudally and setting up the asymmetrical left-right axis of the body.

  14. In vivo genetic ablation of the periotic mesoderm affects cell proliferation survival and differentiation in the cochlea

    PubMed Central

    Xu, Huansheng; Chen, Li; Baldini, Antonio

    2007-01-01

    Tbx1 is required for ear development in humans and mice. Gene manipulation in the mouse has discovered multiple consequences of loss of function on early development of the inner ear, some of which are attributable to a cell autonomous role in maintaining cell proliferation of epithelial progenitors of the cochlear and vestibular apparata. However, ablation of the mesodermal domain of the gene also results in severe but more restricted abnormalities. Here we show that Tbx1 has a dynamic expression during late development of the ear, in particular, is expressed in the sensory epithelium of the vestibular organs but not of the cochlea. Vice versa, it is expressed in the condensed mesenchyme that surrounds the cochlea but not in the one that surrounds the vestibule. Loss of Tbx1 in the mesoderm disrupts this peri-cochlear capsule by strongly reducing the proliferation of mesenchymal cells. The organogenesis of the cochlea, which normally occurs inside the capsule, was dramatically affected in terms of growth of the organ, as well as proliferation, differentiation and survival of its epithelial cells. This model provides a striking demonstration of the essential role played by the periotic mesenchyme in the organogenesis of the cochlea. PMID:17825816

  15. CD13 and ROR2 Permit Isolation of Highly Enriched Cardiac Mesoderm from Differentiating Human Embryonic Stem Cells

    PubMed Central

    Skelton, Rhys J.P.; Brady, Bevin; Khoja, Suhail; Sahoo, Debashis; Engel, James; Arasaratnam, Deevina; Saleh, Kholoud K.; Abilez, Oscar J.; Zhao, Peng; Stanley, Edouard G.; Elefanty, Andrew G.; Kwon, Murray; Elliott, David A.; Ardehali, Reza

    2016-01-01

    Summary The generation of tissue-specific cell types from human embryonic stem cells (hESCs) is critical for the development of future stem cell-based regenerative therapies. Here, we identify CD13 and ROR2 as cell-surface markers capable of selecting early cardiac mesoderm emerging during hESC differentiation. We demonstrate that the CD13+/ROR2+ population encompasses pre-cardiac mesoderm, which efficiently differentiates to all major cardiovascular lineages. We determined the engraftment potential of CD13+/ROR2+ in small (murine) and large (porcine) animal models, and demonstrated that CD13+/ROR2+ progenitors have the capacity to differentiate toward cardiomyocytes, fibroblasts, smooth muscle, and endothelial cells in vivo. Collectively, our data show that CD13 and ROR2 identify a cardiac lineage precursor pool that is capable of successful engraftment into the porcine heart. These markers represent valuable tools for further dissection of early human cardiac differentiation, and will enable a detailed assessment of human pluripotent stem cell-derived cardiac lineage cells for potential clinical applications. PMID:26771355

  16. Metabolic suppression during mesodermal differentiation of embryonic stem cells identified by single-cell comprehensive gene expression analysis.

    PubMed

    Zhou, Yuanshu; Fujisawa, Ikuma; Ino, Kosuke; Matsue, Tomokazu; Shiku, Hitoshi

    2015-09-01

    Flk-1 (VEGF receptor 2) is a well-defined mesodermal progenitor marker and the Flk-1-positive (Flk-1(+)) cells derived from embryonic stem cells (ESCs) have been known to generate hemangioblasts and cardiovascular progenitor cells, which are formed in the early and late stages of differentiation, respectively. In this study, we separated Flk-1(+) and Flk-1(-) cells from spontaneously differentiating embryoid bodies (EBs) of mouse ESCs. We found that cell aggregates derived from late stage Flk-1(+) cells had a relatively small size and a low oxygen consumption rate (OCR) compared with those derived from Flk-1(-) cells. Furthermore, using single-cell comprehensive gene expression analysis, we found that both Flk-1(+) and Flk-1(-) cells could be categorized into subgroups with either low or high glucose metabolic activity. We observed that metabolic suppression occurs in cells expressing an intermediate level of both Nanog and Pou5f1. Taken together, our data suggested that the temporary metabolic suppression is an intrinsic feature of mesodermal differentiation. PMID:26211925

  17. De novo expression of a type IV collagen gene in Drosophila embryos is restricted to mesodermal derivatives and occurs at germ band shortening.

    PubMed

    Mirre, C; Cecchini, J P; Le Parco, Y; Knibiehler, B

    1988-02-01

    We have examined directly the expression of one collagen gene (DCg1) during Drosophila melanogaster embryogenesis by means of in situ hybridization. Transcripts of this gene, which were demonstrated to encode a basement membrane type IV collagen chain, began to accumulate specifically in mesodermal derivatives at stages 12-13 of embryogenesis, and not before. Cells expressing this gene overlap, or are closely intermingled with, somatic and visceral mesoderm in stages 12-14. In stages 15-17, in addition to the strongly positive fat bodies, highly labelled cell spots are found scattered around all the parts of the gut and symmetrically on each side of the ventral nerve cord. They correspond to circulating mesodermal cells which we consider to be haemocytes or mesoblasts. PMID:3138101

  18. Gata4 expression in lateral mesoderm is downstream of BMP4 and isactivated directly by Forkhead and GATA transcription factors through adistal enhancer element

    SciTech Connect

    Rojas, Anabel; De Val, Sarah; Heidt, Analeah B.; Xu, Shan-Mei; Bristow, James; Black, Brian L.

    2005-05-20

    The GATA family of zinc-finger transcription factors plays key roles in the specification and differentiation of multiple cell types during development. GATA4 is an early regulator of gene expression during the development of endoderm and mesoderm, and genetic studies in mice have demonstrated that GATA4 is required for embryonic development.Despite the importance of GATA4 in tissue specification and differentiation, the mechanisms by which Gata4 expression is activated and the transcription factor pathways upstream of GATA4 remain largely undefined. To identify transcriptional regulators of Gata4 in the mouse,we screened conserved noncoding sequences from the mouse Gata4 gene for enhancer activity in transgenic embryos. Here, we define the regulation of a distal enhancer element from Gata4 that is sufficient to direct expression throughout the lateral mesoderm, beginning at 7.5 days of mouse embryonic development. The activity of this enhancer is initially broad but eventually becomes restricted to the mesenchyme surrounding the liver. We demonstrate that the function of this enhancer in transgenic embryos is dependent upon highly conserved Forkhead and GATA transcription factor binding sites, which are bound by FOXF1 and GATA4,respectively. Furthermore, the activity of the Gata4 lateral mesoderm enhancer is attenuated by the BMP antagonist Noggin, and the enhancer is not activated in Bmp4-null embryos. Thus, these studies establish that Gata4 is a direct transcriptional target of Forkhead and GATA transcription factors in the lateral mesoderm, and demonstrate that Gata4lateral mesoderm enhancer activation requires BMP4, supporting a model in which GATA4 serves as a downstream effector of BMP signaling in the lateral mesoderm.

  19. Primordial germ cells in an oligochaete annelid are specified according to the birth rank order in the mesodermal teloblast lineage.

    PubMed

    Kato, Yukie; Nakamoto, Ayaki; Shiomi, Inori; Nakao, Hajime; Shimizu, Takashi

    2013-07-15

    The primordial germ cells (PGCs) in the oligochaete annelid Tubifex tubifex are descentants of the mesodermal (M) teloblast and are located in the two midbody segments X and XI in which they serve as germline precursors forming the testicular gonad and the ovarian gonad, respectively. During embryogenesis, vasa-expressing cells (termed presumptive PGCs or pre-PGCs) emerge in a variable set of midbody segments including the genital segments (X and XI); at the end of embryogenesis, pre-PGCs are confined to the genital segments, where they become PGCs in the juvenile. Here, using live imaging of pre-PGCs, we have demonstrated that during Tubifex embryogenesis, pre-PGCs (defined by Vasa expression) stay in segments where they have emerged, suggesting that it is unlikely that pre-PGCs move intersegmentally during embryogenesis. Thus, it is apparent that pre-PGCs derived from the 10th and 11th M teloblast-derived primary m blast cells (designated m10 and m11) that give rise, respectively, to segments X and XI are specified in situ as PGCs and that those born in other segments become undetectable at the end of embryogenesis. To address the mechanisms for this segment-specific development of PGCs, we have performed a set of cell-transplantation experiments as well as cell-ablation experiments. When m10 and m11 that are normally located in the mid region of the embryo were placed in positions near the anterior end of the host embryo, these cells formed two consecutive segments, which exhibited Vasa-positive PGC-like cells at early juvenile stage. This suggests that in terms of PGC generation, the fates of m10 and m11 remain unchanged even if they are placed in ectopic positions along the anteroposterior axis. Nor was the fate of m10 and m11 changed even if mesodermal blast cell chains preceding or succeeding m10 and m11 were absent. In a previous study, it was shown that PGC development in segments X and XI occurs normally in the absence of the overlying ectoderm. All this

  20. Effect of angiotensin II on proliferation and differentiation of mouse induced pluripotent stem cells into mesodermal progenitor cells

    SciTech Connect

    Ishizuka, Toshiaki; Goshima, Hazuki; Ozawa, Ayako; Watanabe, Yasuhiro

    2012-03-30

    Highlights: Black-Right-Pointing-Pointer Treatment with angiotensin II enhanced LIF-induced DNA synthesis of mouse iPS cells. Black-Right-Pointing-Pointer Angiotensin II may enhance the DNA synthesis via induction of superoxide. Black-Right-Pointing-Pointer Treatment with angiotensin II significantly increased JAK/STAT3 phosphorylation. Black-Right-Pointing-Pointer Angiotensin II enhanced differentiation into mesodermal progenitor cells. Black-Right-Pointing-Pointer Angiotensin II may enhance the differentiation via activation of p38 MAPK. -- Abstract: Previous studies suggest that angiotensin receptor stimulation may enhance not only proliferation but also differentiation of undifferentiated stem/progenitor cells. Therefore, in the present study, we determined the involvement of the angiotensin receptor in the proliferation and differentiation of mouse induced pluripotent stem (iPS) cells. Stimulation with angiotensin II (Ang II) significantly increased DNA synthesis in mouse iPS cells cultured in a medium with leukemia inhibitory factor (LIF). Pretreatment of the cells with either candesartan (a selective Ang II type 1 receptor [AT{sub 1}R] antagonist) or Tempol (a cell-permeable superoxide scavenger) significantly inhibited Ang II-induced DNA synthesis. Treatment with Ang II significantly increased JAK/STAT3 phosphorylation. Pretreatment with candesartan significantly inhibited Ang II- induced JAK/STAT3 phosphorylation. In contrast, induction of mouse iPS cell differentiation into Flk-1-positive mesodermal progenitor cells was performed in type IV collagen (Col IV)- coated dishes in a differentiation medium without LIF. When Col IV-exposed iPS cells were treated with Ang II for 5 days, the expression of Flk-1 was significantly increased compared with that in the cells treated with the vehicle alone. Pretreatment of the cells with both candesartan and SB203580 (a p38 MAPK inhibitor) significantly inhibited the Ang II- induced increase in Flk-1 expression

  1. Csrp1 regulates dynamic cell movements of the mesendoderm and cardiac mesoderm through interactions with Dishevelled and Diversin

    PubMed Central

    Miyasaka, Kota Y.; Kida, Yasuyuki S.; Sato, Takayuki; Minami, Mari; Ogura, Toshihiko

    2007-01-01

    Zebrafish Csrp1 is a member of the cysteine- and glycine-rich protein (CSRP) family and is expressed in the mesendoderm and its derivatives. Csrp1 interacts with Dishevelled 2 (Dvl2) and Diversin (Div), which control cell morphology and other dynamic cell behaviors via the noncanonical Wnt and JNK pathways. When csrp1 message is knocked down, abnormal convergent extension cell movement is induced, resulting in severe deformities in midline structures. In addition, cardiac bifida is induced as a consequence of defects in cardiac mesoderm cell migration. Our data highlight Csrp1 as a key molecule of the noncanonical Wnt pathway, which orchestrates cell behaviors during dynamic morphogenetic movements of tissues and organs. PMID:17592114

  2. GRP94 is essential for mesoderm induction and muscle development because it regulates insulin-like growth factor secretion.

    PubMed

    Wanderling, Sherry; Simen, Birgitte B; Ostrovsky, Olga; Ahmed, Noreen T; Vogen, Shawn M; Gidalevitz, Tali; Argon, Yair

    2007-10-01

    Because only few of its client proteins are known, the physiological roles of the endoplasmic reticulum chaperone glucose-regulated protein 94 (GRP94) are poorly understood. Using targeted disruption of the murine GRP94 gene, we show that it has essential functions in embryonic development. grp94-/- embryos die on day 7 of gestation, fail to develop mesoderm, primitive streak, or proamniotic cavity. grp94-/- ES cells grow in culture and are capable of differentiation into cells representing all three germ layers. However, these cells do not differentiate into cardiac, smooth, or skeletal muscle. Differentiation cultures of mutant ES cells are deficient in secretion of insulin-like growth factor II and their defect can be complemented with exogenous insulin-like growth factors I or II. The data identify insulin-like growth factor II as one developmentally important protein whose production depends on the activity of GRP94. PMID:17634284

  3. Hedgehog–BMP signalling establishes dorsoventral patterning in lateral plate mesoderm to trigger gonadogenesis in chicken embryos

    PubMed Central

    Yoshino, Takashi; Murai, Hidetaka; Saito, Daisuke

    2016-01-01

    The gonad appears in the early embryo after several events: cells at the lateral plate mesoderm (LPM) undergo ingression, begin gonadal differentiation and then retain primordial germ cells (PGCs). Here we show that in the chicken embryo, these events are triggered on the basis of dorsoventral patterning at the medial LPM. Gonadal progenitor cells (GPCs) at the ventromedial LPM initiate gonadogenesis by undergoing ingression, whereas mesonephric capsule progenitor cells (MCPCs) at the dorsomedial LPM do not. These contrasting behaviours are caused by Hedgehog signalling, which is activated in GPCs but not in MCPCs. Inhibiting Hedgehog signalling prevents GPCs from forming gonadal structures and collecting PGCs. When activated by Hedgehog signalling, MCPCs form an ectopic gonad. This Hedgehog signalling is mediated by BMP4. These findings provide insight into embryonic patterning and gonadal initiation in the chicken embryo. PMID:27558761

  4. Normal development of the tomato clownfish Amphiprion frenatus: live imaging and in situ hybridization analyses of mesodermal and neurectodermal development.

    PubMed

    Ghosh, J; Wilson, R W; Kudoh, T

    2009-12-01

    The normal embryonic development of the tomato clownfish Amphiprion frenatus was analysed using live imaging and by in situ hybridization for detection of mesodermal and neurectodermal development. Both morphology of live embryos and tissue-specific staining revealed significant differences in the gross developmental programme of A. frenatus compared with better-known teleost fish models, in particular, initiation of somitogenesis before complete epiboly, initiation of narrowing of the neurectoderm (neurulation) before somitogenesis, relatively early pigmentation of melanophores at the 10-15 somite stage and a distinctive pattern of melanophore distribution. These results suggest evolutionary adaptability of the teleost developmental programme. The ease of obtaining eggs, in vitro culture of the embryo, in situ staining analyses and these reported characteristics make A. frenatus a potentially important model marine fish species for studying embryonic development, physiology, ecology and evolution. PMID:20738687

  5. The control of rostrocaudal pattern in the developing spinal cord: specification of motor neuron subtype identity is initiated by signals from paraxial mesoderm.

    PubMed

    Ensini, M; Tsuchida, T N; Belting, H G; Jessell, T M

    1998-03-01

    The generation of distinct classes of motor neurons is an early step in the control of vertebrate motor behavior. To study the interactions that control the generation of motor neuron subclasses in the developing avian spinal cord we performed in vivo grafting studies in which either the neural tube or flanking mesoderm were displaced between thoracic and brachial levels. The positional identity of neural tube cells and motor neuron subtype identity was assessed by Hox and LIM homeodomain protein expression. Our results show that the rostrocaudal identity of neural cells is plastic at the time of neural tube closure and is sensitive to positionally restricted signals from the paraxial mesoderm. Such paraxial mesodermal signals appear to control the rostrocaudal identity of neural tube cells and the columnar subtype identity of motor neurons. These results suggest that the generation of motor neuron subtypes in the developing spinal cord involves the integration of distinct rostrocaudal and dorsoventral patterning signals that derive, respectively, from paraxial and axial mesodermal cell groups. PMID:9463344

  6. Light and electron microscopic studies of the intestinal epithelium in Notoplana humilis (Platyhelminthes, Polycladida): the contribution of mesodermal/gastrodermal neoblasts to intestinal regeneration.

    PubMed

    Okano, Daisuke; Ishida, Sachiko; Ishiguro, Sei-ichi; Kobayashi, Kazuya

    2015-12-01

    Some free-living flatworms in the phylum Platyhelminthes possess strong regenerative capability that depends on putative pluripotent stem cells known as neoblasts. These neoblasts are defined based on several criteria, including their proliferative capacity and the presence of cellular components known as chromatoid bodies. Polyclads, which are marine flatworms, have the potential to be a good model system for stem cell research, yet little information is available regarding neoblasts and regeneration. In this study, transmission electron microscopy and immunostaining analyses, using antibodies against phospho-histone H3 and BrdU, were used to identify two populations of neoblasts in the polyclad Notoplana humilis: mesodermal neoblasts (located in the mesenchymal space) and gastrodermal neoblasts (located within the intestine, where granular club cells and phagocytic cells are also located). Light and electron microscopic analyses also suggested that phagocytic cells and mesodermal/gastrodermal neoblasts, but not granular club cells, migrated into blastemas and remodeled the intestine during regeneration. Therefore, we suggest that, in polyclads, intestinal regeneration is accomplished by mechanisms underlying both morphallaxis (remodeling of pre-existing tissues) and epimorphosis (de novo tissue formation derived from mesodermal/gastrodermal neoblasts). Based on the assumption that gastrodermal neoblasts, which are derived from mesodermal neoblasts, are intestinal stem cells, we propose a model to study intestinal regeneration. PMID:26104134

  7. Xwnt-8 and lithium can act upon either dorsal mesodermal or neurectodermal cells to cause a loss of forebrain in Xenopus embryos

    NASA Technical Reports Server (NTRS)

    Fredieu, J. R.; Cui, Y.; Maier, D.; Danilchik, M. V.; Christian, J. L.

    1997-01-01

    When Xenopus gastrulae are made to misexpress Xwnt-8, or are exposed to lithium ions, they develop with a loss of anterior structures. In the current study, we have characterized the neural defects produced by either Xwnt-8 or lithium and have examined potential cellular mechanisms underlying this anterior truncation. We find that the primary defect in embryos exposed to lithium at successively earlier stages during gastrulation is a progressive rostral to caudal deletion of the forebrain, while hindbrain and spinal regions of the CNS remain intact. Misexpression of Xwnt-8 during gastrulation produces an identical loss of forebrain. Our results demonstrate that lithium and Wnts can act upon either prospective neural ectodermal cells, or upon dorsal mesodermal cells, to cause a loss of anterior pattern. Specifically, ectodermal cells isolated from lithium- or Wnt-exposed embryos are unable to form anterior neural tissue in response to inductive signals from normal dorsal mesoderm. In addition, although dorsal mesodermal cells from lithium- or Wnt-exposed embryos are specified properly, and produce normal levels of the anterior neural inducing molecules noggin and chordin, they show a greatly reduced capacity to induce anterior neural tissue in conjugated ectoderm. Taken together, our results are consistent with a model in which Wnt- or lithium-mediated signals can induce either mesodermal or ectodermal cells to produce a dominant posteriorizing morphogen which respecifies anterior neural tissue as posterior.

  8. The myogenic repressor gene Holes in muscles is a direct transcriptional target of Twist and Tinman in the Drosophila embryonic mesoderm

    PubMed Central

    Elwell, Jennifer A.; Lovato, TyAnna L.; Adams, Melanie M.; Baca, Erica M.; Lee, Thai; Cripps, Richard M.

    2015-01-01

    Understanding the regulatory circuitry controlling myogenesis is critical to understanding developmental mechanisms and developmentally-derived diseases. We analyzed the transcriptional regulation of a Drosophila myogenic repressor gene, Holes in muscles (Him). Previously, Him was shown to inhibit Myocyte enhancer factor-2 (MEF2) activity, and is expressed in myoblasts but not differentiating myotubes. We demonstrate that different phases of Him embryonic expression arise through the actions of different enhancers, and we characterize the enhancer required for its early mesoderm expression. This Him early mesoderm enhancer contains two conserved binding sites for the basic helix-loop-helix regulator Twist, and one binding site for the NK homeodomain protein Tinman. The sites for both proteins are required for enhancer activity in early embryos. Twist and Tinman activate the enhancer in tissue culture assays, and ectopic expression of either factor is sufficient to direct ectopic expression of a Him-lacZ reporter, or of the endogenous Him gene. Moreover, sustained expression of twist expression in the mesoderm up-regulates mesodermal Him expression in late embryos. Our findings provide a model to define mechanistically how Twist can both promotes myogenesis through direct activation of Mef2, and can place a brake on myogenesis, through direct activation of Him. PMID:25704510

  9. Positional identity of murine mesenchymal stem cells resident in different organs is determined in the postsegmentation mesoderm.

    PubMed

    Sági, Bernadett; Maraghechi, Pouneh; Urbán, Veronika S; Hegyi, Beáta; Szigeti, Anna; Fajka-Boja, Roberta; Kudlik, Gyöngyi; Német, Katalin; Monostori, Eva; Gócza, Elen; Uher, Ferenc

    2012-03-20

    Although mesenchymal stem cells (MSCs) of distinct tissue origin have a large number of similarities and differences, it has not been determined so far whether tissue-resident MSCs are the progenies of one ancestor cell lineage or the results of parallel cell developmental events. Here we compared the expression levels of 177 genes in murine MSCs derived from adult and juvenile bone marrow and adult adipose tissue, as well as juvenile spleen, thymus, and aorta wall by quantitative real-time polymerase chain reaction and the results were partially validated at protein level. All MSC lines uniformly expressed a large set of genes including well-known mesenchymal markers, such as α-smooth muscle actin, collagen type I α-chain, GATA6, Mohawk, and vimentin. In contrast, pluripotency genes and the early mesodermal marker T-gene were not expressed. On the other hand, different MSC lines consistently expressed distinct patterns of Hox genes determining the positional identity of a given cell population. Moreover, MSCs of different origin expressed a few other transcription factors also reflecting their topological identity and so the body segment or organ to which they normally contributed in vivo: (1) thymus-derived cells specifically expressed Tbx5 and Pitx2; (2) spleen-derived MSCs were characterized with Tlx1 and Nkx2.5; (3) Pitx1 designated femoral bone marrow cells and (4) En2 appeared in aorta wall-derived MSCs. Thus, MSCs exhibited topographic identity and memory even after long-term cultivation in vitro. On the basis of these results, we suggest that postnatal MSCs isolated from different anatomical sites descend from precursor cells developing in the postsegmentation mesoderm. PMID:22149974

  10. Dvr1 transfers left-right asymmetric signals from Kupffer's vesicle to lateral plate mesoderm in zebrafish.

    PubMed

    Peterson, Annita G; Wang, Xinghao; Yost, H Joseph

    2013-10-01

    An early step in establishing left-right (LR) symmetry in zebrafish is the generation of asymmetric fluid flow by Kupffer's vesicle (KV). As a result of fluid flow, a signal is generated and propagated from the KV to the left lateral plate mesoderm, activating a transcriptional response of Nodal expression in the left lateral plate mesoderm (LPM). The mechanisms and molecules that aid in this transfer of information from the KV to the left LPM are still not clear. Here we provide several lines of evidence demonstrating a role for a member of the TGFβ family member, Dvr1, a zebrafish Vg1 ortholog. Dvr1 is expressed bilaterally between the KV and the LPM. Knockdown of Dvr1 by morpholino causes dramatically reduced or absent expression of southpaw (spaw, a Nodal homolog), in LPM, and corresponding loss of downstream Lefty (lft1 and lft) expression, and aberrant brain and heart LR patterning. Dvr1 morphant embryos have normal KV morphology and function, normal expression of southpaw (spaw) and charon (cha) in the peri-KV region and normal expression of a variety of LPM markers in LPM. Additionally, Dvr1 knockdown does not alter the capability of LPM to respond to signals that initiate and propagate spaw expression. Co-injection experiments in Xenopus and zebrafish indicate that Dvr1 and Spaw can enhance each other's ability to activate the Nodal response pathway and co-immunoprecipitation experiments reveal differential relationships among activators and inhibitors in this pathway. These results indicate that Dvr1 is responsible for enabling the transfer of a left-right signal from KV to the LPM. PMID:23791819

  11. Loss of Abdominal Muscle in Pitx2 Mutants Associated with Altered Axial Specification of Lateral Plate Mesoderm

    PubMed Central

    Eng, Diana; Ma, Hsiao-Yen; Xu, Jun; Shih, Hung-Ping; Gross, Michael K.; Kiouss, Chrissa

    2012-01-01

    Sequence specific transcription factors (SSTFs) combinatorially define cell types during development by forming recursively linked network kernels. Pitx2 expression begins during gastrulation, together with Hox genes, and becomes localized to the abdominal lateral plate mesoderm (LPM) before the onset of myogenesis in somites. The somatopleure of Pitx2 null embryos begins to grow abnormally outward before muscle regulatory factors (MRFs) or Pitx2 begin expression in the dermomyotome/myotome. Abdominal somites become deformed and stunted as they elongate into the mutant body wall, but maintain normal MRF expression domains. Subsequent loss of abdominal muscles is therefore not due to defects in specification, determination, or commitment of the myogenic lineage. Microarray analysis was used to identify SSTF families whose expression levels change in E10.5 interlimb body wall biopsies. All Hox9-11 paralogs had lower RNA levels in mutants, whereas genes expressed selectively in the hypaxial dermomyotome/myotome and sclerotome had higher RNA levels in mutants. In situ hybridization analyses indicate that Hox gene expression was reduced in parts of the LPM and intermediate mesoderm of mutants. Chromatin occupancy studies conducted on E10.5 interlimb body wall biopsies showed that Pitx2 protein occupied chromatin sites containing conserved bicoid core motifs in the vicinity of Hox 9-11 and MRF genes. Taken together, the data indicate that Pitx2 protein in LPM cells acts, presumably in combination with other SSTFs, to repress gene expression, that are normally expressed in physically adjoining cell types. Pitx2 thereby prevents cells in the interlimb LPM from adopting the stable network kernels that define sclerotomal, dermomyotomal, or myotomal mesenchymal cell types. This mechanism may be viewed either as lineage restriction or specification. PMID:22860089

  12. The migration of paraxial and lateral plate mesoderm cells emerging from the late primitive streak is controlled by different Wnt signals

    PubMed Central

    Sweetman, Dylan; Wagstaff, Laura; Cooper, Oliver; Weijer, Cornelis; Münsterberg, Andrea

    2008-01-01

    Background Co-ordinated cell movement is a fundamental feature of developing embryos. Massive cell movements occur during vertebrate gastrulation and during the subsequent extension of the embryonic body axis. These are controlled by cell-cell signalling and a number of pathways have been implicated. Here we use long-term video microscopy in chicken embryos to visualize the migration routes and movement behaviour of mesoderm progenitor cells as they emerge from the primitive streak (PS) between HH stages 7 and 10. Results We observed distinct cell movement behaviours along the length of the streak and determined that this is position dependent with cells responding to environmental cues. The behaviour of cells was altered by exposing embryos or primitive streak explants to cell pellets expressing Wnt3a and Wnt5a, without affecting cell fates, thus implicating these ligands in the regulation of cell movement behaviour. Interestingly younger embryos were not responsive, suggesting that Wnt3a and Wnt5a are specifically involved in the generation of posterior mesoderm, consistent with existing mouse and zebrafish mutants. To investigate which downstream components are involved mutant forms of dishevelled (dsh) and prickle1 (pk1) were electroporated into the primitive streak. These had differential effects on the behaviour of mesoderm progenitors emerging from anterior or posterior regions of the streak, suggesting that multiple Wnt pathways are involved in controlling cell migration during extension of the body axis in amniote embryos. Conclusion We suggest that the distinct behaviours of paraxial and lateral mesoderm precursors are regulated by the opposing actions of Wnt5a and Wnt3a as they leave the primitive streak in neurula stage embryos. Our data suggests that Wnt5a acts via prickle to cause migration of cells from the posterior streak. In the anterior streak, this is antagonised by Wnt3a to generate non-migratory medial mesoderm. PMID:18541012

  13. Plant Homeo Domain Finger Protein 8 Regulates Mesodermal and Cardiac Differentiation of Embryonic Stem Cells Through Mediating the Histone Demethylation of pmaip1.

    PubMed

    Tang, Yan; Hong, Ya-Zhen; Bai, Hua-Jun; Wu, Qiang; Chen, Charlie Degui; Lang, Jing-Yu; Boheler, Kenneth R; Yang, Huang-Tian

    2016-06-01

    Histone demethylases have emerged as key regulators of biological processes. The H3K9me2 demethylase plant homeo domain finger protein 8(PHF8), for example, is involved in neuronal differentiation, but its potential function in the differentiation of embryonic stem cells (ESCs) to cardiomyocytes is poorly understood. Here, we explored the role of PHF8 during mesodermal and cardiac lineage commitment of mouse ESCs (mESCs). Using a phf8 knockout (ph8(-/Y) ) model, we found that deletion of phf8 in ESCs did not affect self-renewal, proliferation or early ectodermal/endodermal differentiation, but it did promote the mesodermal lineage commitment with the enhanced cardiomyocyte differentiation. The effects were accompanied by a reduction in apoptosis through a caspase 3-independent pathway during early ESC differentiation, without significant differences between differentiating wide-type (ph8(+/Y) ) and ph8(-/Y) ESCs in cell cycle progression or proliferation. Functionally, PHF8 promoted the loss of a repressive mark H3K9me2 from the transcription start site of a proapoptotic gene pmaip1 and activated its transcription. Furthermore, knockdown of pmaip1 mimicked the phenotype of ph8(-/Y) by showing the decreased apoptosis during early differentiation of ESCs and promoted mesodermal and cardiac commitment, while overexpression of pmaip1 or phf8 rescued the phenotype of ph8(-/Y) ESCs by increasing the apoptosis and weakening the mesodermal and cardiac differentiation. These results reveal that the histone demethylase PHF8 regulates mesodermal lineage and cell fate decisions in differentiating mESCs through epigenetic control of the gene critical to programmed cell death pathways. Stem Cells 2016;34:1527-1540. PMID:26866517

  14. Morphological comparison of the craniofacial phenotypes of mouse models expressing the Apert FGFR2 S252W mutation in neural crest- or mesoderm-derived tissues

    PubMed Central

    Heuzé, Yann; Singh, Nandini; Basilico, Claudio; Jabs, Ethylin Wang; Holmes, Greg; Richtsmeier, Joan T

    2014-01-01

    Bones of the craniofacial skeleton are derived from two distinct cell lineages, cranial neural crest and mesoderm, and articulate at sutures and synchondroses which represent major bone growth sites. Premature fusion of cranial suture(s) is associated with craniofacial dysmorphogenesis caused in part by alteration in the growth potential at sutures and can occur as an isolated birth defect or as part of a syndrome, such as Apert syndrome. Conditional expression of the Apert FGFR2 S252W mutation in mesoderm was previously shown to be necessary and sufficient to cause coronal craniosynostosis. Here we used micro computed tomography images of mice expressing the Apert mutation constitutively in either mesoderm or neural crest to quantify craniofacial shape variation and suture fusion patterns, and to identify shape changes in craniofacial bones derived from the lineage not expressing the mutation, referred to here as secondary shape changes. Our results show that at postnatal day 0: (i) conditional expression of the FGFR2 S252W mutation in neural crest-derived tissues causes a more severe craniofacial phenotype than when expressed in mesoderm-derived tissues; and (ii) both mesoderm- and neural crest-specific mouse models display secondary shape changes. We also show that premature suture fusion is not necessarily dependent on the expression of the FGFR2 S252W mutation in the sutural mesenchyme. More specifically, it appears that suture fusion patterns in both mouse models are suture-specific resulting from a complex combination of the influence of primary abnormalities of biogenesis or signaling within the sutures, and timing. PMID:24632501

  15. Sonic hedgehog enhances somite cell viability and formation of primary slow muscle fibers in avian segmented mesoderm.

    PubMed

    Cann, G M; Lee, J W; Stockdale, F E

    1999-09-01

    Primary skeletal muscle fibers first form in the segmented portions of paraxial mesoderm called somites. Although the neural tube and notochord are recognized as crucial in patterning myogenic cell lineages during avian and mammalian somitic myogenesis, the source, identities, and actions of the signals governing this process remain controversial. It has been shown that signals emanating from the ventral neural tube and/or notochord alone or Shh alone serve to activate MyoD expression in somites. However, beyond a role in initiating MyoD expression, little is known about the effects of Shh on primary muscle fiber formation in somites of higher vertebrates. The studies reported here investigate how the ventral neural tube promotes myogenesis and compare the effects of the ventral neural tube with those of purified Shh protein on fiber formation in somites. We show that purified Shh protein mimics actions of the ventral neural tube on somites including initiation of muscle fiber formation, enhancement of numbers of primary muscle fibers, and particularly, the formation of primary fibers that express slow myosin. There is a marked increase in slow myosin expression in fibers in response to Shh as somites mature. The effects of ventral neural tube on fiber formation can be blocked by disrupting the Shh signaling pathway by increasing the activity of somitic cyclic AMP-dependent protein kinase A. Furthermore, it was demonstrated that apoptosis is a dominant fate of somite cells, but not somitic muscle fibers, when cultured in the absence of the neural tube, and that application of Shh protein to somites reduced apoptosis. The block to apoptosis by Shh is a manifestation of the maturity of the somite with a progressive increase in the block as somites are displaced rostrally from somite III forward. We conclude that purified Shh protein in mimicking the effects of the ventral neural tube on segmented mesoderm can exert pleiotropic effects during primary myogenesis

  16. Two Hox cofactors, the Meis/Hth homolog UNC-62 and the Pbx/Exd homolog CEH-20, function together during C. elegans postembryonic mesodermal development

    PubMed Central

    Jiang, Yuan; Shi, Herong; Liu, Jun

    2009-01-01

    The TALE homeodomain-containing PBC and MEIS proteins play multiple roles during metazoan development. Mutations in these proteins can cause various disorders, including cancer. In this study, we examined the roles of MEIS proteins in mesoderm development in C. elegans using the postembryonic mesodermal M lineage as a model system. We found that the MEIS protein UNC-62 plays essential roles in regulating cell fate specification and differentiation in the M lineage. Furthermore, UNC-62 appears to function together with the PBC protein CEH-20 in regulating these processes. Both unc-62 and ceh-20 have overlapping expression patterns within and outside of the M lineage, and they share physical and regulatory interactions. In particular, we found that ceh-20 is genetically required for the promoter activity of unc-62, providing evidence for another layer of regulatory interactions between MEIS and PBC proteins. PMID:19643105

  17. Left-right asymmetry and kinesin superfamily protein KIF3A: new insights in determination of laterality and mesoderm induction by kif3A-/- mice analysis.

    PubMed

    Takeda, S; Yonekawa, Y; Tanaka, Y; Okada, Y; Nonaka, S; Hirokawa, N

    1999-05-17

    KIF3A is a classical member of the kinesin superfamily proteins (KIFs), ubiquitously expressed although predominantly in neural tissues, and which forms a heterotrimeric KIF3 complex with KIF3B or KIF3C and an associated protein, KAP3. To elucidate the function of the kif3A gene in vivo, we made kif3A knockout mice. kif3A-/- embryos displayed severe developmental abnormalities characterized by neural tube degeneration and mesodermal and caudal dysgenesis and died during the midgestational period at approximately 10.5 dpc (days post coitum), possibly resulting from cardiovascular insufficiency. Whole mount in situ hybridization of Pax6 revealed a normal pattern while staining by sonic hedgehog (shh) and Brachyury (T) exhibited abnormal patterns in the anterior-posterior (A-P) direction at both mesencephalic and thoracic levels. These results suggest that KIF3A might be involved in mesodermal patterning and in turn neurogenesis. PMID:10330409

  18. Atmospheric-pressure plasma-irradiation inhibits mouse embryonic stem cell differentiation to mesoderm and endoderm but promotes ectoderm differentiation

    NASA Astrophysics Data System (ADS)

    Miura, Taichi; Hamaguchi, Satoshi; Nishihara, Shoko

    2016-04-01

    Recently, various effects of low-temperature atmospheric-pressure plasma irradiation on living cells have been demonstrated, such as tissue sterilization, blood coagulation, angiogenesis, wound healing, and tumor elimination. However, the effect of plasma-irradiation on the differentiation of mouse embryonic stem cells (mESCs) has not yet been clarified. A large number of reactive species are generated by plasma-irradiation in medium, of which hydrogen peroxide (H2O2) is one of the main species generated. Here, we investigated the effect of plasma-irradiation on the differentiation of mESCs using an embryoid body (EB) formation assay with plasma-irradiated medium or H2O2-supplemented non-irradiated medium. Our findings demonstrated that plasma-irradiated medium potently inhibits the differentiation from mESCs to mesoderm and endoderm by inhibiting Wnt signaling as determined by quantitative polymerase chain reaction and immunoblotting analyses. In contrast, both the plasma-irradiated medium and H2O2-supplemented non-irradiated medium enhanced the differentiation to epiblastoid, ectodermal, and neuronal lineages by activation of fibroblast growth factor 4 (FGF4) signaling, suggesting that these effects are caused by the H2O2 generated by plasma-irradiation in medium. However, in each case, the differentiation to glial cells remained unaffected. This study is the first demonstration that plasma-irradiation affects the differentiation of mESCs by the regulation of Wnt and FGF4 signaling pathways.

  19. Organiser role of the stem cell of the mesoderm in prototroch patterning in Patella vulgata (Mollusca, Gastropoda).

    PubMed

    Damen, P; Dictus, W J

    1996-05-01

    During early development of the gastropod mollusc Patella vulgata, the stem cell of the mesoderm (3D-macromere) is induced. As a result of this induction, the embryo becomes dorsoventrally organised. At about the same time in development, ciliated cells, so-called trochoblasts, are formed. Later in development, some trochoblasts deciliate and, together with the ciliated trochoblast, form the dorsoventrally organised prototroch, the locomotory organ of the larva. In order to study the role of the 3D-macromere in the specification of trochoblasts and in the induction of the dorsoventral organisation of the prototroch, induction of 3D has been prevented in various ways. it is shown that preventing 3D-induction results in the formation of a radially symmetrical prototroch. The trochoblasts of all four quadrants developed like corresponding trochoblasts of the A-quadrant. Somewhere between 30 and 120 min after fifth cleavage the 3D-macromere induces the formation of specific trochoblasts and organises the dorsoventral pattern of the prototroch. Besides a role of the 3D-macromere, a role of other cells has been demonstrated in the conditionally specified deciliation of trochoblasts. PMID:8798146

  20. Tbx1 is required autonomously for cell survival and fate in the pharyngeal core mesoderm to form the muscles of mastication.

    PubMed

    Kong, Ping; Racedo, Silvia E; Macchiarulo, Stephania; Hu, Zunju; Carpenter, Courtney; Guo, Tingwei; Wang, Tao; Zheng, Deyou; Morrow, Bernice E

    2014-08-15

    Velo-cardio-facial/DiGeorge syndrome, also known as 22q11.2 deletion syndrome, is a congenital anomaly disorder characterized by craniofacial anomalies including velo-pharyngeal insufficiency, facial muscle hypotonia and feeding difficulties, in part due to hypoplasia of the branchiomeric muscles. Inactivation of both alleles of mouse Tbx1, encoding a T-box transcription factor, deleted on chromosome 22q11.2, results in reduction or loss of branchiomeric muscles. To identify downstream pathways, we performed gene profiling of microdissected pharyngeal arch one (PA1) from Tbx1(+/+) and Tbx1(-/-) embryos at stages E9.5 (somites 20-25) and E10.5 (somites 30-35). Basic helix-loop-helix (bHLH) transcription factors were reduced, while secondary heart field genes were increased in expression early and were replaced by an increase in expression of cellular stress response genes later, suggesting a change in gene expression patterns or cell populations. Lineage tracing studies using Mesp1(Cre) and T-Cre drivers showed that core mesoderm cells within PA1 were present at E9.5 but were greatly reduced by E10.5 in Tbx1(-/-) embryos. Using Tbx1(Cre) knock-in mice, we found that cells are lost due to apoptosis, consistent with increase in expression of cellular stress response genes at E10.5. To determine whether Tbx1 is required autonomously in the core mesoderm, we used Mesp1(Cre) and T-Cre mesodermal drivers in combination with inactivate Tbx1 and found reduction or loss of branchiomeric muscles from PA1. These mechanistic studies inform us that Tbx1 is required upstream of key myogenic genes needed for core mesoderm cell survival and fate, between E9.5 and E10.5, resulting in formation of the branchiomeric muscles. PMID:24705356

  1. Tbx1 is required autonomously for cell survival and fate in the pharyngeal core mesoderm to form the muscles of mastication

    PubMed Central

    Kong, Ping; Racedo, Silvia E.; Macchiarulo, Stephania; Hu, Zunju; Carpenter, Courtney; Guo, Tingwei; Wang, Tao; Zheng, Deyou; Morrow, Bernice E.

    2014-01-01

    Velo-cardio-facial/DiGeorge syndrome, also known as 22q11.2 deletion syndrome, is a congenital anomaly disorder characterized by craniofacial anomalies including velo-pharyngeal insufficiency, facial muscle hypotonia and feeding difficulties, in part due to hypoplasia of the branchiomeric muscles. Inactivation of both alleles of mouse Tbx1, encoding a T-box transcription factor, deleted on chromosome 22q11.2, results in reduction or loss of branchiomeric muscles. To identify downstream pathways, we performed gene profiling of microdissected pharyngeal arch one (PA1) from Tbx1+/+ and Tbx1−/− embryos at stages E9.5 (somites 20–25) and E10.5 (somites 30–35). Basic helix–loop–helix (bHLH) transcription factors were reduced, while secondary heart field genes were increased in expression early and were replaced by an increase in expression of cellular stress response genes later, suggesting a change in gene expression patterns or cell populations. Lineage tracing studies using Mesp1Cre and T-Cre drivers showed that core mesoderm cells within PA1 were present at E9.5 but were greatly reduced by E10.5 in Tbx1−/− embryos. Using Tbx1Cre knock-in mice, we found that cells are lost due to apoptosis, consistent with increase in expression of cellular stress response genes at E10.5. To determine whether Tbx1 is required autonomously in the core mesoderm, we used Mesp1Cre and T-Cre mesodermal drivers in combination with inactivate Tbx1 and found reduction or loss of branchiomeric muscles from PA1. These mechanistic studies inform us that Tbx1 is required upstream of key myogenic genes needed for core mesoderm cell survival and fate, between E9.5 and E10.5, resulting in formation of the branchiomeric muscles. PMID:24705356

  2. The meso-angioblast: a multipotent, self-renewing cell that originates from the dorsal aorta and differentiates into most mesodermal tissues.

    PubMed

    Minasi, Maria G; Riminucci, Mara; De Angelis, Luciana; Borello, Ugo; Berarducci, Barbara; Innocenzi, Anna; Caprioli, Arianna; Sirabella, Dario; Baiocchi, Marta; De Maria, Ruggero; Boratto, Renata; Jaffredo, Thierry; Broccoli, Vania; Bianco, Paolo; Cossu, Giulio

    2002-06-01

    We have previously reported the origin of a class of skeletal myogenic cells from explants of dorsal aorta. This finding disagrees with the known origin of all skeletal muscle from somites and has therefore led us to investigate the in vivo origin of these cells and, moreover, whether their fate is restricted to skeletal muscle, as observed in vitro under the experimental conditions used. To address these issues, we grafted quail or mouse embryonic aorta into host chick embryos. Donor cells, initially incorporated into the host vessels, were later integrated into mesodermal tissues, including blood, cartilage, bone, smooth, skeletal and cardiac muscle. When expanded on a feeder layer of embryonic fibroblasts, the clonal progeny of a single cell from the mouse dorsal aorta acquired unlimited lifespan, expressed hemo-angioblastic markers (CD34, Flk1 and Kit) at both early and late passages, and maintained multipotency in culture or when transplanted into a chick embryo. We conclude that these newly identified vessel-associated stem cells, the meso-angioblasts, participate in postembryonic development of the mesoderm, and we speculate that postnatal mesodermal stem cells may be derived from a vascular developmental origin. PMID:12015303

  3. FoxA4 Favours Notochord Formation by Inhibiting Contiguous Mesodermal Fates and Restricts Anterior Neural Development in Xenopus Embryos

    PubMed Central

    Murgan, Sabrina; Castro Colabianchi, Aitana Manuela; Monti, Renato José; Boyadjián López, Laura Elena; Aguirre, Cecilia E.; Stivala, Ernesto González; López, Silvia L.

    2014-01-01

    In vertebrates, the embryonic dorsal midline is a crucial signalling centre that patterns the surrounding tissues during development. Members of the FoxA subfamily of transcription factors are expressed in the structures that compose this centre. Foxa2 is essential for dorsal midline development in mammals, since knock-out mouse embryos lack a definitive node, notochord and floor plate. The related gene foxA4 is only present in amphibians. Expression begins in the blastula –chordin and –noggin expressing centre (BCNE) and is later restricted to the dorsal midline derivatives of the Spemann's organiser. It was suggested that the early functions of mammalian foxa2 are carried out by foxA4 in frogs, but functional experiments were needed to test this hypothesis. Here, we show that some important dorsal midline functions of mammalian foxa2 are exerted by foxA4 in Xenopus. We provide new evidence that the latter prevents the respecification of dorsal midline precursors towards contiguous fates, inhibiting prechordal and paraxial mesoderm development in favour of the notochord. In addition, we show that foxA4 is required for the correct regionalisation and maintenance of the central nervous system. FoxA4 participates in constraining the prospective rostral forebrain territory during neural specification and is necessary for the correct segregation of the most anterior ectodermal derivatives, such as the cement gland and the pituitary anlagen. Moreover, the early expression of foxA4 in the BCNE (which contains precursors of the whole forebrain and most of the midbrain and hindbrain) is directly required to restrict anterior neural development. PMID:25343614

  4. Functional analysis of grimp, a novel gene required for mesodermal cell proliferation at an initial stage of regeneration in Enchytraeus japonensis (Enchytraeidae, Oligochaete).

    PubMed

    Takeo, Makoto; Yoshida-Noro, Chikako; Tochinai, Shin

    2010-01-01

    Enchytraeus japonensis is a small oligochaete species, which has a remarkable regeneration capacity. It has been proposed as a new model animal for the study of regeneration, and some histological studies of this species have been carried out. On the other hand, the molecular biological mechanism of regeneration is almost unknown in this species. To clarify the molecular biological mechanism operating at an initial stage of regeneration in E. japonensis, we isolated by the cDNA subtraction method five genes whose expression levels changed in the regeneration process occurring between growing and early regenerating worms. One of the isolated genes (a novel gene named grimp) was expressed transiently from 3 to 12 h post amputation only in neoblasts and a population of mesodermal cells (the non-neoblast grimp-expressing cells) incorporating BrdU simultaneously showed mitotic activity. We succeeded in inhibiting grimp expression by RNA interference (RNAi), thus applying this technique for the first time in Oligochaeta. In knock-down worms, the number of BrdU-positive neoblasts and the non-neoblast grimp-expressing cells in the coelom drastically decreased. Moreover, the elongation and the segmentation of blastemas were inhibited, while no statistically significant inhibitory effect was observed in epidermal and intestinal cells. These results suggest that grimp is required for initial proliferation of neoblasts and some mesodermal cells for regeneration. PMID:19876829

  5. Primary Bovine Extra-Embryonic Cultured Cells: A New Resource for the Study of In Vivo Peri-Implanting Phenotypes and Mesoderm Formation

    PubMed Central

    Hue, Isabelle; Evain-Brion, Danièle; Fournier, Thierry; Degrelle, Séverine A.

    2015-01-01

    In addition to nourishing the embryo, extra-embryonic tissues (EETs) contribute to early embryonic patterning, primitive hematopoiesis, and fetal health. These tissues are of major importance for human medicine, as well as for efforts to improve livestock efficiency, but they remain incompletely understood. In bovines, EETs are accessible easily, in large amounts, and prior to implantation. We took advantage of this system to describe, in vitro and in vivo, the cell types present in bovine EETs at Day 18 of development. Specifically, we characterized the gene expression patterns and phenotypes of bovine extra-embryonic ectoderm (or trophoblast; bTC), endoderm (bXEC), and mesoderm (bXMC) cells in culture and compared them to their respective in vivo micro-dissected cells. After a week of culture, certain characteristics (e.g., gene expression) of the in vitro cells were altered with respect to the in vivo cells, but we were able to identify “cores” of cell-type-specific (and substrate-independent) genes that were shared between in vitro and in vivo samples. In addition, many cellular phenotypes were cell-type-specific with regard to extracellular adhesion. We evaluated the ability of individual bXMCs to migrate and spread on micro-patterns, and observed that they easily adapted to diverse environments, similar to in vivo EE mesoderm cells, which encounter different EE epithelia to form chorion, yolk sac, and allantois. With these tissue interactions, different functions arose that were detected in silico and corroborated in vivo at D21–D25. Moreover, analysis of bXMCs allowed us to identify the EE cell ring surrounding the embryonic disc (ED) at D14-15 as mesoderm cells, which had been hypothesized but not shown prior to this study. We envision these data will serve as a major resource for the future in the analysis of peri-implanting phenotypes in response to the maternal metabolism and contribute to subsequent studies of placental/fetal development in

  6. Expression cloning of Xantivin, a Xenopus lefty/antivin-related gene, involved in the regulation of activin signaling during mesoderm induction.

    PubMed

    Tanegashima, K; Yokota, C; Takahashi, S; Asashima, M

    2000-12-01

    In a screening for activin-responsive genes, we isolated a Xenopus lefty/antivin-related gene, called Xantivin (Xatv). In the animal cap assay, the expression of Xatv was induced by activin signaling, and in the embryo, by nodal-related genes. Overexpression of Xatv in the marginal zone caused suppression of mesoderm formation and gastrulation defects, and inhibited the secondary axis formation induced by Xnr1 and Xactivin, suggesting that Xatv acted as a feedback inhibitor of activin signaling. However, in the animal cap, Xatv failed to antagonize Xnr1 and Xactivin. This result suggested that Xatv has different responses in the marginal zone and in the animal region, and antagonizes to a higher degree activin signaling in the marginal zone. PMID:11091069

  7. A comprehensive gene expression analysis at sequential stages of in vitro cardiac differentiation from isolated MESP1-expressing-mesoderm progenitors.

    PubMed

    den Hartogh, Sabine C; Wolstencroft, Katherine; Mummery, Christine L; Passier, Robert

    2016-01-01

    In vitro cardiac differentiation of human pluripotent stem cells (hPSCs) closely recapitulates in vivo embryonic heart development, and therefore, provides an excellent model to study human cardiac development. We recently generated the dual cardiac fluorescent reporter MESP1(mCherry/w)NKX2-5(eGFP/w) line in human embryonic stem cells (hESCs), allowing the visualization of pre-cardiac MESP1+ mesoderm and their further commitment towards the cardiac lineage, marked by activation of the cardiac transcription factor NKX2-5. Here, we performed a comprehensive whole genome based transcriptome analysis of MESP1-mCherry derived cardiac-committed cells. In addition to previously described cardiac-inducing signalling pathways, we identified novel transcriptional and signalling networks indicated by transient activation and interactive network analysis. Furthermore, we found a highly dynamic regulation of extracellular matrix components, suggesting the importance to create a versatile niche, adjusting to various stages of cardiac differentiation. Finally, we identified cell surface markers for cardiac progenitors, such as the Leucine-rich repeat-containing G-protein coupled receptor 4 (LGR4), belonging to the same subfamily of LGR5, and LGR6, established tissue/cancer stem cells markers. We provide a comprehensive gene expression analysis of cardiac derivatives from pre-cardiac MESP1-progenitors that will contribute to a better understanding of the key regulators, pathways and markers involved in human cardiac differentiation and development. PMID:26783251

  8. Cytostatic Effect of Repeated Exposure to Simvastatin: A Mechanism for Chronic Myotoxicity Revealed by the Use of Mesodermal Progenitors Derived from Human Pluripotent Stem Cells.

    PubMed

    Peric, Delphine; Barragan, Isabel; Giraud-Triboult, Karine; Egesipe, Anne-Laure; Meyniel-Schicklin, Laurène; Cousin, Christelle; Lotteau, Vincent; Petit, Vincent; Touhami, Jawida; Battini, Jean-Luc; Sitbon, Marc; Pinset, Christian; Ingelman-Sundberg, Magnus; Laustriat, Delphine; Peschanski, Marc

    2015-10-01

    Statin treatment of hypercholesterolemia can lead to chronic myotoxicity which is, in most cases, alleviated by drug withdrawal. Cellular and molecular mechanisms of this adverse effect have been elusive, in particular because of the lack of in vitro models suitable for long-term exposures. We have taken advantage of the properties of human pluripotent stem cell-derived mesodermal precursors, that can be maintained unaltered in vitro for a long period of time, to develop a model of repeated exposures to simvastatin during more than 2 weeks. This approach unveiled major differences, both in functional and molecular terms, in response to single versus repeated-dose exposures to simvastatin. The main functional effect of the in vitro simvastatin-induced long-term toxicity was a loss of proliferative capacity in the absence of concomitant cell death, revealing that cytostatic effect could be a major contributor to statin-induced myotoxicity. Comparative analysis of molecular modifications induced by simvastatin short-term versus prolonged exposures demonstrated powerful adaptive cell responses, as illustrated by the dramatic decrease in the number of differentially expressed genes, distinct biological pathway enrichments, and distinct patterns of nutrient transporters expressed at the cell surface. This study underlines the potential of derivatives of human pluripotent stem cells for developing new approaches in toxicology, in particular for chronic toxicity testing. PMID:26184566

  9. Osr1 expression demarcates a multi-potent population of intermediate mesoderm that undergoes progressive restriction to an Osr1-dependent nephron progenitor compartment within the mammalian kidney

    PubMed Central

    Mugford, Joshua W.; Sipilä, Petra; McMahon, Jill A.; McMahon, Andrew P.

    2008-01-01

    The mammalian metanephric kidney is derived from the intermediate mesoderm. In this report, we use molecular fate mapping to demonstrate that the majority of cell types within the metanephric kidney arise from an Osr1+ population of metanephric progenitor cells. These include the ureteric epithelium of the collecting duct network, the cap mesenchyme and its nephron epithelia derivatives, the interstitial mesenchyme, vasculature and smooth muscle. Temporal fate mapping shows a progressive restriction of Osr1+ cell fates such that at the onset of active nephrogenesis, Osr1 activity is restricted to the Six2+ cap mesenchyme nephron progenitors. However, low-level labeling of Osr1+ cells suggests that the specification of interstitial mesenchyme and cap mesenchyme progenitors occurs within the Osr1+ population prior to the onset of metanephric development. Furthermore, although Osr1+ progenitors give rise to much of the kidney, Osr1 function is only essential for the development of the nephron progenitor compartment. These studies provide new insights into the cellular origins of metanephric kidney structures and lend support to a model where Osr1 function is limited to establishing the nephron progenitor pool. PMID:18835385

  10. Osr1 expression demarcates a multi-potent population of intermediate mesoderm that undergoes progressive restriction to an Osr1-dependent nephron progenitor compartment within the mammalian kidney.

    PubMed

    Mugford, Joshua W; Sipilä, Petra; McMahon, Jill A; McMahon, Andrew P

    2008-12-01

    The mammalian metanephric kidney is derived from the intermediate mesoderm. In this report, we use molecular fate mapping to demonstrate that the majority of cell types within the metanephric kidney arise from an Osr1(+) population of metanephric progenitor cells. These include the ureteric epithelium of the collecting duct network, the cap mesenchyme and its nephron epithelia derivatives, the interstitial mesenchyme, vasculature and smooth muscle. Temporal fate mapping shows a progressive restriction of Osr1(+) cell fates such that at the onset of active nephrogenesis, Osr1 activity is restricted to the Six2(+) cap mesenchyme nephron progenitors. However, low-level labeling of Osr1(+) cells suggests that the specification of interstitial mesenchyme and cap mesenchyme progenitors occurs within the Osr1(+) population prior to the onset of metanephric development. Furthermore, although Osr1(+) progenitors give rise to much of the kidney, Osr1 function is only essential for the development of the nephron progenitor compartment. These studies provide new insights into the cellular origins of metanephric kidney structures and lend support to a model where Osr1 function is limited to establishing the nephron progenitor pool. PMID:18835385

  11. A comprehensive gene expression analysis at sequential stages of in vitro cardiac differentiation from isolated MESP1-expressing-mesoderm progenitors

    PubMed Central

    den Hartogh, Sabine C.; Wolstencroft, Katherine; Mummery, Christine L.; Passier, Robert

    2016-01-01

    In vitro cardiac differentiation of human pluripotent stem cells (hPSCs) closely recapitulates in vivo embryonic heart development, and therefore, provides an excellent model to study human cardiac development. We recently generated the dual cardiac fluorescent reporter MESP1mCherry/wNKX2-5eGFP/w line in human embryonic stem cells (hESCs), allowing the visualization of pre-cardiac MESP1+ mesoderm and their further commitment towards the cardiac lineage, marked by activation of the cardiac transcription factor NKX2-5. Here, we performed a comprehensive whole genome based transcriptome analysis of MESP1-mCherry derived cardiac-committed cells. In addition to previously described cardiac-inducing signalling pathways, we identified novel transcriptional and signalling networks indicated by transient activation and interactive network analysis. Furthermore, we found a highly dynamic regulation of extracellular matrix components, suggesting the importance to create a versatile niche, adjusting to various stages of cardiac differentiation. Finally, we identified cell surface markers for cardiac progenitors, such as the Leucine-rich repeat-containing G-protein coupled receptor 4 (LGR4), belonging to the same subfamily of LGR5, and LGR6, established tissue/cancer stem cells markers. We provide a comprehensive gene expression analysis of cardiac derivatives from pre-cardiac MESP1-progenitors that will contribute to a better understanding of the key regulators, pathways and markers involved in human cardiac differentiation and development. PMID:26783251

  12. Hidden in the crowd: primordial germ cells and somatic stem cells in the mesodermal posterior growth zone of the polychaete Platynereis dumerillii are two distinct cell populations

    PubMed Central

    2012-01-01

    Background In the polychaete Platynereis, the primordial germ cells (PGCs) emerge from the vasa, piwi, and PL10 expressing mesodermal posterior growth zone (MPGZ) at the end of larval development, suggesting a post-embryonic formation from stem cells. Methods In order to verify this hypothesis, embryos and larvae were pulse labeled with the proliferation marker 5-ethynyl-2'-deoxyuridine (EdU) at different stages of development. Subsequently, the PGCs were visualized in 7-day-old young worms using antibodies against the Vasa protein. Results Surprisingly, the primordial germ cells of Platynereis incorporate EdU only shortly before gastrulation (6-8 hours post fertilization (hpf)), which coincides with the emergence of four small blastomeres from the mesoblast lineage. We conclude that these so-called 'secondary mesoblast cells' constitute the definitive PGCs in Platynereis. In contrast, the cells of the MPGZ incorporate EdU only from the pre-trochophore stage onward (14 hpf). Conclusion While PGCs and the cells of the MPGZ in Platynereis are indistinguishable in morphology and both express the germline markers vasa, nanos, and piwi, a distinct cluster of PGCs is detectable anterior of the MPGZ following EdU pulse-labeling. Indeed the PGCs form independently from the stem cells of the MPGZ prior to gastrulation. Our data suggest an early PGC formation in the polychaete by preformation rather than by epigenesis. PMID:22512981

  13. Tissue Reactivity of the 14F7 Mab Raised against N-Glycolyl GM3 Ganglioside in Tumors of Neuroectodermal, Mesodermal, and Epithelial Origin

    PubMed Central

    Blanco, Rancés; Quintana, Yisel; Blanco, Damián; Cedeño, Mercedes; Rengifo, Charles E.; Frómeta, Milagros; Ríos, Martha; Rengifo, Enrique; Carr, Adriana

    2013-01-01

    The expression of N-glycolylneuraminic acid forming the structure of gangliosides and/or other glycoconjugates (Hanganutziu-Deicher antigen) in human has been considered as a tumor-associated antigen. Specifically, some reports of 14F7 Mab (a highly specific Mab raised against N-glycolyl GM3 ganglioside) reactivity in human tumors have been recently published. Nevertheless, tumors of epithelial origin have been mostly evaluated. The goal of the present paper was to evaluate the immunohistochemical recognition of 14F7 Mab in different human tumors of neuroectodermal, mesodermal, and epithelial origins using an immunoperoxidase staining method. Samples of fetal, normal, and reactive astrocytosis of the brain were also included in the study. In general, nontumoral tissues, as well as, low-grade brain tumors showed no or a limited immunoreaction with 14F7 Mab. Nevertheless, high-grade astrocytomas (III-IV) and neuroblastomas, as well as, sarcomas and thyroid carcinomas were mostly reactive with 14F7. No reaction was evidenced in medulloblastomas and ependymoblastomas. Our data suggest that the expression of N-glycolyl GM3 ganglioside could be related to the aggressive behavior of malignant cells, without depending on the tumor origin. Our data could also support the possible use of N-glycolyl GM3 as a target for both active and passive immunotherapies of malignancies expressing this molecule. PMID:26317019

  14. Free-form-fabricated commercially pure Ti and Ti6Al4V porous scaffolds support the growth of human embryonic stem cell-derived mesodermal progenitors.

    PubMed

    de Peppo, G M; Palmquist, A; Borchardt, P; Lennerås, M; Hyllner, J; Snis, A; Lausmaa, J; Thomsen, P; Karlsson, C

    2012-01-01

    Commercially-pure titanium (cp-Ti) and the titanium-aluminum-vanadium alloy (Ti6Al4V) are widely used as reconstructive implants for skeletal engineering applications, due to their good mechanical properties, biocompatibility and ability to integrate with the surrounding bone. Electron beam melting technology (EBM) allows the fabrication of customized implants with tailored mechanical properties and high potential in the clinical practice. In order to augment the interaction with the biological tissue, stem cells have recently been combined with metallic scaffolds for skeletal engineering applications. We previously demonstrated that human embryonic stem cell-derived mesodermal progenitors (hES-MPs) hold a great potential to provide a homogeneous and unlimited supply of cells for bone engineering applications. This study demonstrates the effect of EBM-fabricated cp-Ti and Ti6Al4V porous scaffolds on hES-MPs behavior, in terms of cell attachment, growth and osteogenic differentiation. Displaying different chemical composition but similar surface properties, EBM-fabricated cp-Ti and Ti6Al4V scaffolds supported cell attachment and growth, and did not seem to alter the expression of genes involved in osteogenic differentiation and affect the alkaline phosphatase activity. In conclusion, interfacing hES-MPs to EBM-fabricated scaffolds may represent an interesting strategy for design of third-generation biomaterials, with the potential to promote implant integration in clinical conditions characterized by poor bone quality. PMID:22262956

  15. Differential activation of Myf5 and MyoD by different Wnts in explants of mouse paraxial mesoderm and the later activation of myogenesis in the absence of Myf5.

    PubMed

    Tajbakhsh, S; Borello, U; Vivarelli, E; Kelly, R; Papkoff, J; Duprez, D; Buckingham, M; Cossu, G

    1998-11-01

    Activation of myogenesis in newly formed somites is dependent upon signals derived from neighboring tissues, namely axial structures (neural tube and notochord) and dorsal ectoderm. In explants of paraxial mesoderm from mouse embryos, axial structures preferentially activate myogenesis through a Myf5-dependent pathway and dorsal ectoderm preferentially through a MyoD-dependent pathway. Here we report that cells expressing Wnt1 will preferentially activate Myf5 while cells expressing Wnt7a will preferentially activate MyoD. Wnt1 is expressed in the dorsal neural tube and Wnt7a in dorsal ectoderm in the early embryo, therefore both can potentially act in vivo to activate Myf5 and MyoD, respectively. Wnt4, Wnt5a and Wnt6 exert an intermediate effect activating both Myf5 and MyoD equivalently in paraxial mesoderm. Sonic Hedgehog synergises with both Wnt1 and Wnt7a in explants from E8.5 paraxial mesoderm but not in explants from E9.5 embryos. Signaling through different myogenic pathways may explain the rescue of muscle formation in Myf5 null embryos, which do not form an early myotome but later develop both epaxial and hypaxial musculature. Explants of unsegmented paraxial mesoderm contain myogenic precursors capable of expressing MyoD in response to signaling from a neural tube isolated from E10.5 embryos, the developmental stage when MyoD is present throughout the embryo. Myogenic cells cannot activate MyoD in response to signaling from a less mature neural tube. Together these data suggest that different Wnt molecules can activate myogenesis through different pathways such that commitment of myogenic precursors is precisely regulated in space and time to achieve the correct pattern of skeletal muscle development. PMID:9753670

  16. Transcriptome profiles of Penaeus (Marsupenaeus) japonicus animal and vegetal half-embryos: identification of sex determination, germ line, mesoderm, and other developmental genes.

    PubMed

    Sellars, Melony J; Trewin, Carolyn; McWilliam, Sean M; Glaves, R S E; Hertzler, Philip L

    2015-06-01

    There is virtually no knowledge of the molecular events controlling early embryogenesis in Penaeid shrimp. A combination of controlled spawning environment, shrimp embryo micro-dissection techniques, and next-generation sequencing was used to produce transcriptome EST datasets of Penaeus japonicus animal and vegetal half-embryos. Embryos were collected immediately after spawning, and then blastomeres were separated at the two-cell stage and allowed to develop to late gastrulation, then pooled for RNA isolation and cDNA synthesis. Ion Torrent sequencing of cDNA from approximately 500 pooled animal and vegetal half-embryos from multiple spawnings resulted in 560,516 and 493,703 reads, respectively. Reads from each library were assembled and Gene Ontogeny analysis produced 3479 annotated animal contigs and 4173 annotated vegetal contigs, with 159/139 hits for developmental processes in the animal/vegetal contigs, respectively. Contigs were subject to BLAST for selected developmental toolbox genes. Some of the genes found included the sex determination genes sex-lethal and transformer; the germ line genes argonaute 1, boule, germ cell-less, gustavus, maelstrom, mex-3, par-1, pumilio, SmB, staufen, and tudor; the mesoderm genes brachyury, mef2, snail, and twist; the axis determination/segmentation genes β-catenin, deformed, distal-less, engrailed, giant, hairy, hunchback, kruppel, orthodenticle, patched, tailless, and wingless/wnt-8c; and a number of cell-cycle regulators. Animal and vegetal contigs were computationally subtracted from each other to produce sets unique to either half-embryo library. Genes expressed only in the animal half included bmp1, kruppel, maelstrom, and orthodenticle. Genes expressed only in the vegetal half included boule, brachyury, deformed, dorsal, engrailed, hunchback, spalt, twist, and wingless/wnt-8c. PMID:25634056

  17. Posterior-anterior gradient of zebrafish hes6 expression in the presomitic mesoderm is established by the combinatorial functions of the downstream enhancer and 3'UTR.

    PubMed

    Kawamura, Akinori; Ovara, Hiroki; Ooka, Yuko; Kinoshita, Hirofumi; Hoshikawa, Miki; Nakajo, Kenji; Yokota, Daisuke; Fujino, Yuuri; Higashijima, Shin-ichi; Takada, Shinji; Yamasu, Kyo

    2016-01-15

    In vertebrates, the periodic formation of somites from the presomitic mesoderm (PSM) is driven by the molecular oscillator known as the segmentation clock. The hairy-related gene, hes6/her13.2, functions as a hub by dimerizing with other oscillators of the segmentation clock in zebrafish embryos. Although hes6 exhibits a posterior-anterior expression gradient in the posterior PSM with a peak at the tailbud, the detailed mechanisms underlying this unique expression pattern have not yet been clarified. By establishing several transgenic lines, we found that the transcriptional regulatory region downstream of hes6 in combination with the hes6 3'UTR recapitulates the endogenous gradient of hes6 mRNA expression. This downstream region, which we termed the PT enhancer, possessed several putative binding sites for the T-box and Ets transcription factors that were required for the regulatory activity. Indeed, the T-box transcription factor (Tbx16) and Ets transcription factor (Pea3) bound specifically to the putative binding sites and regulated the enhancer activity in zebrafish embryos. In addition, the 3'UTR of hes6 is required for recapitulation of the endogenous mRNA expression. Furthermore, the PT enhancer with the 3'UTR of hes6 responded to the inhibition of retinoic acid synthesis and fibroblast growth factor signaling in a manner similar to endogenous hes6. The results showed that transcriptional regulation by the T-box and Ets transcription factors, combined with the mRNA stability given by the 3'UTR, is responsible for the unique expression gradient of hes6 mRNA in the posterior PSM of zebrafish embryos. PMID:26596999

  18. CXCR2 Inhibition in Human Pluripotent Stem Cells Induces Predominant Differentiation to Mesoderm and Endoderm Through Repression of mTOR, β-Catenin, and hTERT Activities

    PubMed Central

    Jung, Ji-Hye; Kang, Ka-Won; Kim, Jihea; Hong, Soon-Chul; Park, Yong

    2016-01-01

    On the basis of our previous report verifying that chemokine (C-X-C motif) receptor 2 (CXCR2) ligands in human placenta-derived cell conditioned medium (hPCCM) support human pluripotent stem cell (hPSC) propagation without exogenous basic fibroblast growth factor (bFGF), this study was designed to identify the effect of CXCR2 manipulation on the fate of hPSCs and the underlying mechanism, which had not been previously determined. We observed that CXCR2 inhibition in hPSCs induces predominant differentiation to mesoderm and endoderm with concomitant loss of hPSC characteristics and accompanying decreased expression of mammalian target of rapamycin (mTOR), β-catenin, and human telomerase reverse transcriptase (hTERT). These phenomena are recapitulated in hPSCs propagated in conventional culture conditions, including bFGF as well as those in hPCCM without exogenous bFGF, suggesting that the action of CXCR2 on hPSCs might not be associated with a bFGF-related mechanism. In addition, the specific CXCR2 ligand growth-related oncogene α (GROα) markedly increased the expression of ectodermal markers in differentiation-committed embryoid bodies derived from hPSCs. This finding suggests that CXCR2 inhibition in hPSCs prohibits the propagation of hPSCs and leads to predominant differentiation to mesoderm and endoderm owing to the blockage of ectodermal differentiation. Taken together, our results indicate that CXCR2 preferentially supports the maintenance of hPSC characteristics as well as facilitates ectodermal differentiation after the commitment to differentiation, and the mechanism might be associated with mTOR, β-catenin, and hTERT activities. PMID:27188501

  19. CXCR2 Inhibition in Human Pluripotent Stem Cells Induces Predominant Differentiation to Mesoderm and Endoderm Through Repression of mTOR, β-Catenin, and hTERT Activities.

    PubMed

    Jung, Ji-Hye; Kang, Ka-Won; Kim, Jihea; Hong, Soon-Chul; Park, Yong; Kim, Byung Soo

    2016-07-01

    On the basis of our previous report verifying that chemokine (C-X-C motif) receptor 2 (CXCR2) ligands in human placenta-derived cell conditioned medium (hPCCM) support human pluripotent stem cell (hPSC) propagation without exogenous basic fibroblast growth factor (bFGF), this study was designed to identify the effect of CXCR2 manipulation on the fate of hPSCs and the underlying mechanism, which had not been previously determined. We observed that CXCR2 inhibition in hPSCs induces predominant differentiation to mesoderm and endoderm with concomitant loss of hPSC characteristics and accompanying decreased expression of mammalian target of rapamycin (mTOR), β-catenin, and human telomerase reverse transcriptase (hTERT). These phenomena are recapitulated in hPSCs propagated in conventional culture conditions, including bFGF as well as those in hPCCM without exogenous bFGF, suggesting that the action of CXCR2 on hPSCs might not be associated with a bFGF-related mechanism. In addition, the specific CXCR2 ligand growth-related oncogene α (GROα) markedly increased the expression of ectodermal markers in differentiation-committed embryoid bodies derived from hPSCs. This finding suggests that CXCR2 inhibition in hPSCs prohibits the propagation of hPSCs and leads to predominant differentiation to mesoderm and endoderm owing to the blockage of ectodermal differentiation. Taken together, our results indicate that CXCR2 preferentially supports the maintenance of hPSC characteristics as well as facilitates ectodermal differentiation after the commitment to differentiation, and the mechanism might be associated with mTOR, β-catenin, and hTERT activities. PMID:27188501

  20. Fusion of the SUMO/Sentrin-specific protease 1 gene SENP1 and the embryonic polarity-related mesoderm development gene MESDC2 in a patient with an infantile teratoma and a constitutional t(12;15)(q13;q25).

    PubMed

    Veltman, Imke M; Vreede, Lilian A; Cheng, Jinke; Looijenga, Leendert H J; Janssen, Bert; Schoenmakers, Eric F P M; Yeh, Edward T H; van Kessel, Ad Geurts

    2005-07-15

    Recently, we identified a patient with an infantile sacrococcygeal teratoma and a constitutional t(12;15)(q13;q25). Here, we show that, as a result of this chromosomal translocation, the SUMO/Sentrin-specific protease 1 gene (SENP1) on chromosome 12 and the embryonic polarity-related mesoderm development gene (MESDC2) on chromosome 15 are disrupted and fused. Both reciprocal SENP1-MESDC2 (SEME) and MESDC2-SENP1 (MESE) fusion genes are transcribed in tumor-derived cells and their open reading frames encode aberrant proteins. As a consequence of this, and in contrast to wild-type (WT) MESDC2, the translocation-associated SEME protein is no longer targeted to the endoplasmatic reticulum, leading to a presumed loss-of-function as a chaperone for the WNT co-receptors LRP5 and/or LRP6. Ultimately, this might lead to abnormal development and/or routing of germ cell tumor precursor cells. SUMO, a post-translational modifier, plays an important role in several cellular key processes and is cleaved from its substrates by WT SENP1. Using a PML desumoylation assay, we found that translocation-associated MESE proteins exhibit desumoylation capacities similar to those observed for WT SENP1. We speculate that spatio-temporal disturbances in desumoylating activities during critical stages of embryonic development might have predisposed the patient. Together, the constitutional t(12;15)(q13;q25) translocation revealed two novel candidate genes for neonatal/infantile GCT development: MESDC2 and SENP1. PMID:15917269

  1. Human Myocardial Pericytes: Multipotent Mesodermal Precursors Exhibiting Cardiac Specificity

    PubMed Central

    Chen, William C.W.; Baily, James E.; Corselli, Mirko; Diaz, Mary; Sun, Bin; Xiang, Guosheng; Gray, Gillian A.; Huard, Johnny; Péault, Bruno

    2015-01-01

    Perivascular mesenchymal precursor cells (i.e. pericytes) reside in skeletal muscle where they contribute to myofiber regeneration; however, the existence of similar microvessel-associated regenerative precursor cells in cardiac muscle has not yet been documented. We tested whether microvascular pericytes within human myocardium exhibit phenotypes and multipotency similar to their anatomically and developmentally distinct counterparts. Fetal and adult human heart pericytes (hHPs) express canonical pericyte markers in situ, including CD146, NG2, PDGFRβ, PDGFRα, αSMA, and SM-MHC, but not CD117, CD133 and desmin, nor endothelial cell (EC) markers. hHPs were prospectively purified to homogeneity from ventricular myocardium by flow cytometry, based on a combination of positive- (CD146) and negative-selection (CD34, CD45, CD56, and CD117) cell lineage markers. Purified hHPs expanded in vitro were phenotypically similar to human skeletal muscle-derived pericytes (hSkMPs). hHPs express MSC markers in situ and exhibited osteo- chondro-, and adipogenic potentials but, importantly, no ability for skeletal myogenesis, diverging from pericytes of all other origins. hHPs supported network formation with/without ECs in Matrigel cultures; hHPs further stimulated angiogenic responses under hypoxia, markedly different from hSkMPs. The cardiomyogenic potential of hHPs was examined following 5-azacytidine treatment and neonatal cardiomyocyte co-culture in vitro, and intramyocardial transplantation in vivo. Results indicated cardiomyocytic differentiation in a small fraction of hHPs. In conclusion, human myocardial pericytes share certain phenotypic and developmental similarities with their skeletal muscle homologs, yet exhibit different antigenic, myogenic, and angiogenic properties. This is the first example of an anatomical restriction in the developmental potential of pericytes as native mesenchymal stem cells. PMID:25336400

  2. Regulation of UNC-130/FOXD-mediated mesodermal patterning in C. elegans.

    PubMed

    Kersey, Rossio K; Brodigan, Thomas M; Fukushige, Tetsunari; Krause, Michael W

    2016-08-15

    Spatial polarity cues in animals are used repeatedly during development for many processes, including cell fate determination, cell migration, and axon guidance. In Caenorhabditis elegans, the body wall muscle extends the length of the animal in four distinct quadrants and generates an UNC-129/TGF-β-related signal that is much higher in the dorsal two muscle quadrants compared to their ventral counterparts. This pattern of unc-129 expression requires the activity of the proposed transcriptional repressor UNC-130/FOXD whose body wall muscle activity is restricted to the ventral two body wall muscle quadrants. To understand how these dorsal-ventral differences in UNC-130 activity are established and maintained, we have analyzed the regulation of unc-130 expression and the distribution of UNC-130 protein. We have identified widespread, cis-acting elements in the unc-130 promoter that function to positively regulate ventral body wall muscle expression and negatively regulate dorsal body wall muscle expression. We have defined the temporal distribution of UNC-130 protein in body wall muscle cells during embryogenesis, demonstrated that this pattern is required to establish the dorsal-ventral polarity of UNC-129/TGF-β, and shown that UNC-130 is not required post-embryonically to maintain the asymmetry of body wall muscle unc-129 expression. Finally, we have tested the impact of the depletion of a variety of transcription factors, repressors, and signaling molecules to identify additional regulators of body wall muscle UNC-130 polarity. Our results confirm and extend earlier studies to clarify the mechanisms by which UNC-130 is controlled and affects the pattern of unc-129 expression in body wall muscle. These results further our understanding of the transcriptional logic behind the generation of polarity cues involving this poorly understood subclass of Forkhead factors. PMID:27341757

  3. HOX-mediated LMO2 expression in embryonic mesoderm is recapitulated in acute leukaemias.

    PubMed

    Calero-Nieto, F J; Joshi, A; Bonadies, N; Kinston, S; Chan, W-I; Gudgin, E; Pridans, C; Landry, J-R; Kikuchi, J; Huntly, B J; Gottgens, B

    2013-11-28

    The Lim Domain Only 2 (LMO2) leukaemia oncogene encodes an LIM domain transcriptional cofactor required for early haematopoiesis. During embryogenesis, LMO2 is also expressed in developing tail and limb buds, an expression pattern we now show to be recapitulated in transgenic mice by an enhancer in LMO2 intron 4. Limb bud expression depended on a cluster of HOX binding sites, while posterior tail expression required the HOX sites and two E-boxes. Given the importance of both LMO2 and HOX genes in acute leukaemias, we further demonstrated that the regulatory hierarchy of HOX control of LMO2 is activated in leukaemia mouse models as well as in patient samples. Moreover, Lmo2 knock-down impaired the growth of leukaemic cells, and high LMO2 expression at diagnosis correlated with poor survival in cytogenetically normal AML patients. Taken together, these results establish a regulatory hierarchy of HOX control of LMO2 in normal development, which can be resurrected during leukaemia development. Redeployment of embryonic regulatory hierarchies in an aberrant context is likely to be relevant in human pathologies beyond the specific example of ectopic activation of LMO2. PMID:23708655

  4. Tissue-Specific Cultured Human Pericytes: Perivascular Cells from Smooth Muscle Tissue Have Restricted Mesodermal Differentiation Ability.

    PubMed

    Pierantozzi, Enrico; Vezzani, Bianca; Badin, Margherita; Curina, Carlo; Severi, Filiberto Maria; Petraglia, Felice; Randazzo, Davide; Rossi, Daniela; Sorrentino, Vincenzo

    2016-05-01

    Microvascular pericytes (PCs) are considered the adult counterpart of the embryonic mesoangioblasts, which represent a multipotent cell population that resides in the dorsal aorta of the developing embryo. Although PCs have been isolated from several adult organs and tissues, it is still controversial whether PCs from different tissues exhibit distinct differentiation potentials. To address this point, we investigated the differentiation potentials of isogenic human cultured PCs isolated from skeletal (sk-hPCs) and smooth muscle tissues (sm-hPCs). We found that both sk-hPCs and sm-hPCs expressed known pericytic markers and did not express endothelial, hematopoietic, and myogenic markers. Both sk-hPCs and sm-hPCs were able to differentiate into smooth muscle cells. In contrast, sk-hPCs, but not sm-hPCs, differentiated in skeletal muscle cells and osteocytes. Given the reported ability of the Notch pathway to regulate skeletal muscle and osteogenic differentiation, sk-hPCs and sm-hPCs were treated with N-[N-(3,5- difluorophenacetyl)-L-alanyl]-S-phenylglycine t-butyl ester (DAPT), a known inhibitor of Notch signaling. DAPT treatment, as assessed by histological and molecular analysis, enhanced myogenic differentiation and abolished osteogenic potential of sk-hPCs. In contrast, DAPT treatment did not affect either myogenic or osteogenic differentiation of sm-hPCs. In summary, these results indicate that, despite being isolated from the same anatomical niche, cultured PCs from skeletal muscle and smooth muscle tissues display distinct differentiation abilities. PMID:26956507

  5. Zic3 is required in the extra-cardiac perinodal region of the lateral plate mesoderm for left-right patterning and heart development

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Mutations in ZIC3 cause human X-linked heterotaxy and isolated cardiovascular malformations. A mouse model with targeted deletion of Zic3 demonstrates an early role for Zic3 in gastrulation, CNS, cardiac and left-right axial development. The observation of multiple malformations in Zic3(null) mice a...

  6. Fibronectin mediates mesendodermal cell fate decisions

    PubMed Central

    Cheng, Paul; Andersen, Peter; Hassel, David; Kaynak, Bogac L.; Limphong, Pattraranee; Juergensen, Lonny; Kwon, Chulan; Srivastava, Deepak

    2013-01-01

    Non-cell-autonomous signals often play crucial roles in cell fate decisions during animal development. Reciprocal signaling between endoderm and mesoderm is vital for embryonic development, yet the key signals and mechanisms remain unclear. Here, we show that endodermal cells efficiently promote the emergence of mesodermal cells in the neighboring population through signals containing an essential short-range component. The endoderm-mesoderm interaction promoted precardiac mesoderm formation in mouse embryonic stem cells and involved endodermal production of fibronectin. In vivo, fibronectin deficiency resulted in a dramatic reduction of mesoderm accompanied by endodermal expansion in zebrafish embryos. This event was mediated by regulation of Wnt signaling in mesodermal cells through activation of integrin-β1. Our findings highlight the importance of the extracellular matrix in mediating short-range signals and reveal a novel function of endoderm, involving fibronectin and its downstream signaling cascades, in promoting the emergence of mesoderm. PMID:23715551

  7. Inhibition of β-catenin signaling respecifies anterior-like endothelium into beating human cardiomyocytes.

    PubMed

    Palpant, Nathan J; Pabon, Lil; Roberts, Meredith; Hadland, Brandon; Jones, Daniel; Jones, Christina; Moon, Randall T; Ruzzo, Walter L; Bernstein, Irwin; Zheng, Ying; Murry, Charles E

    2015-09-15

    During vertebrate development, mesodermal fate choices are regulated by interactions between morphogens such as activin/nodal, BMPs and Wnt/β-catenin that define anterior-posterior patterning and specify downstream derivatives including cardiomyocyte, endothelial and hematopoietic cells. We used human embryonic stem cells to explore how these pathways control mesodermal fate choices in vitro. Varying doses of activin A and BMP4 to mimic cytokine gradient polarization in the anterior-posterior axis of the embryo led to differential activity of Wnt/β-catenin signaling and specified distinct anterior-like (high activin/low BMP) and posterior-like (low activin/high BMP) mesodermal populations. Cardiogenic mesoderm was generated under conditions specifying anterior-like mesoderm, whereas blood-forming endothelium was generated from posterior-like mesoderm, and vessel-forming CD31(+) endothelial cells were generated from all mesoderm origins. Surprisingly, inhibition of β-catenin signaling led to the highly efficient respecification of anterior-like endothelium into beating cardiomyocytes. Cardiac respecification was not observed in posterior-derived endothelial cells. Thus, activin/BMP gradients specify distinct mesodermal subpopulations that generate cell derivatives with unique angiogenic, hemogenic and cardiogenic properties that should be useful for understanding embryogenesis and developing therapeutics. PMID:26153229

  8. Dual embryonic origin and patterning of the pharyngeal skeleton in the axolotl (Ambystoma mexicanum).

    PubMed

    Sefton, Elizabeth M; Piekarski, Nadine; Hanken, James

    2015-01-01

    The impressive morphological diversification of vertebrates was achieved in part by innovation and modification of the pharyngeal skeleton. Extensive fate mapping in amniote models has revealed a primarily cranial neural crest derivation of the pharyngeal skeleton. Although comparable fate maps of amphibians produced over several decades have failed to document a neural crest derivation of ventromedial elements in these vertebrates, a recent report provides evidence of a mesodermal origin of one of these elements, basibranchial 2, in the axolotl. We used a transgenic labeling protocol and grafts of labeled cells between GFP+ and white embryos to derive a fate map that describes contributions of both cranial neural crest and mesoderm to the axolotl pharyngeal skeleton, and we conducted additional experiments that probe the mechanisms that underlie mesodermal patterning. Our fate map confirms a dual embryonic origin of the pharyngeal skeleton in urodeles, including derivation of basibranchial 2 from mesoderm closely associated with the second heart field. Additionally, heterotopic transplantation experiments reveal lineage restriction of mesodermal cells that contribute to pharyngeal cartilage. The mesoderm-derived component of the pharyngeal skeleton appears to be particularly sensitive to retinoic acid (RA): administration of exogenous RA leads to loss of the second basibranchial, but not the first. Neural crest was undoubtedly critical in the evolution of the vertebrate pharyngeal skeleton, but mesoderm may have played a central role in forming ventromedial elements, in particular. When and how many times during vertebrate phylogeny a mesodermal contribution to the pharyngeal skeleton evolved remain to be resolved. PMID:25963195

  9. Experimental analysis of blood vessel development in the avian wing bud.

    PubMed

    Feinberg, R N; Noden, D M

    1991-09-01

    Shortly after its appearance, the avian limb bud becomes populated by a rich plexus of vascular channels. Formation of this plexus occurs by angiogenesis, specifically the ingrowth of branches from the dorsal aorta or cardinal veins, and by differentiation of endogenous angioblasts within limb mesoderm. However, mesenchyme located immediately beneath the surface ectoderm of the limb is devoid of patent blood vessels. The objective of this research is to ascertain whether peripheral limb mesoderm lacks angioblasts at all stages or becomes avascular secondarily during limb development. Grafts of core or peripheral wing mesoderm, identified by the presence or absence of patent channels following systemic infusion with ink, were grafted from quail embryos at stages 16-26 into the head region of chick embryos at stages 9-10. Hosts were fixed 3-5 days later and sections treated with antibodies that recognize quail endothelial cells and their precursors. Labeled endothelial cells were found intercalated into normal craniofacial blood vessels both nearby and distant from the site of implantation following grafting of limb core mesoderm from any stage. Identical results were obtained following grafting of limb peripheral mesoderm at stages 16-21. However, peripheral mesoderm from donors older than stage 22 did not contain endothelial precursors. Thus at the onset of appendicular development angioblasts are present throughout the mesoderm of the limb bud. During the fourth day of incubation, these cells are lost from peripheral mesoderm, either through emigration or degeneration. PMID:1750708

  10. Malignant teratoma (image)

    MedlinePlus

    A malignant teratoma is a type of cancer consisting of cysts that contain one or more of the three primary embryonic germ layers: ectoderm, mesoderm, and endoderm. Because malignant teratomas have usually spread by the time of diagnosis, ...

  11. Evolution of the head-trunk interface in tetrapod vertebrates.

    PubMed

    Sefton, Elizabeth M; Bhullar, Bhart-Anjan S; Mohaddes, Zahra; Hanken, James

    2016-01-01

    Vertebrate neck musculature spans the transition zone between head and trunk. The extent to which the cucullaris muscle is a cranial muscle allied with the gill levators of anamniotes or is instead a trunk muscle is an ongoing debate. Novel computed tomography datasets reveal broad conservation of the cucullaris in gnathostomes, including coelacanth and caecilian, two sarcopterygians previously thought to lack it. In chicken, lateral plate mesoderm (LPM) adjacent to occipital somites is a recently identified embryonic source of cervical musculature. We fate-map this mesoderm in the axolotl (Ambystoma mexicanum), which retains external gills, and demonstrate its contribution to posterior gill-levator muscles and the cucullaris. Accordingly, LPM adjacent to the occipital somites should be regarded as posterior cranial mesoderm. The axial position of the head-trunk border in axolotl is congruent between LPM and somitic mesoderm, unlike in chicken and possibly other amniotes. PMID:27090084

  12. Antagonizing the Spemann organizer: role of the homeobox gene Xvent-1.

    PubMed Central

    Gawantka, V; Delius, H; Hirschfeld, K; Blumenstock, C; Niehrs, C

    1995-01-01

    We have identified a novel homeobox gene, Xvent-1, that is differentially expressed in the ventral marginal zone of the early Xenopus gastrula. Evidence is presented from mRNA microinjection experiments for a role for this gene in dorsoventral patterning of mesoderm. First, Xvent-1 is induced by BMP-4, a gene known to be a key regulator of ventral mesoderm development. Second, Xvent-1 and the organizer-specific gene goosecoid are able to interact, directly or indirectly, in a cross-regulatory loop suppressing each other's expression, consistent with their mutually exclusive expression in the marginal zone. Third, microinjection of Xvent-1 mRNA ventralizes dorsal mesoderm. The results suggest that Xvent-1 functions in a ventral signaling pathway that maintains the ventral mesodermal state and antagonizes the Spemann organizer. Images PMID:8557046

  13. Comparative morphology and development of extra-ocular muscles in the lamprey and gnathostomes reveal the ancestral state and developmental patterns of the vertebrate head.

    PubMed

    Suzuki, Daichi G; Fukumoto, Yuma; Yoshimura, Miho; Yamazaki, Yuji; Kosaka, Jun; Kuratani, Shigeru; Wada, Hiroshi

    2016-01-01

    The ancestral configuration of the vertebrate head has long been an intriguing topic in comparative morphology and evolutionary biology. One peculiar component of the vertebrate head is the presence of extra-ocular muscles (EOMs), the developmental mechanism and evolution of which remain to be determined. The head mesoderm of elasmobranchs undergoes local epithelialization into three head cavities, precursors of the EOMs. In contrast, in avians, these muscles appear to develop mainly from the mesenchymal head mesoderm. Importantly, in the basal vertebrate lamprey, the head mesoderm does not show overt head cavities or signs of segmental boundaries, and the development of the EOMs is not well described. Furthermore, the disposition of the lamprey EOMs differs from those the rest of vertebrates, in which the morphological pattern of EOMs is strongly conserved. To better understand the evolution and developmental origins of the vertebrate EOMs, we explored the development of the head mesoderm and EOMs of the lamprey in detail. We found that the disposition of lamprey EOM primordia differed from that in gnathostomes, even during the earliest period of development. We also found that three components of the paraxial head mesoderm could be distinguished genetically (premandibular mesoderm: Gsc+/TbxA-; mandibular mesoderm: Gsc-/TbxA-; hyoid mesoderm: Gsc-/TbxA+), indicating that the genetic mechanisms of EOMs are conserved in all vertebrates. We conclude that the tripartite developmental origin of the EOMs is likely to have been possessed by the latest common ancestor of the vertebrates. This ancestor's EOM developmental pattern was also suggested to have resembled more that of the lamprey, and the gnathostome EOMs' disposition is likely to have been established by a secondary modification that took place in the common ancestor of crown gnathostomes. PMID:27081572

  14. Visualizing the lateral somitic frontier in the Prx1Cre transgenic mouse

    PubMed Central

    Durland, J Logan; Sferlazzo, Matteo; Logan, Malcolm; Burke, Ann Campbell

    2008-01-01

    Changes in the organization of the musculoskeletal system have accounted for many evolutionary adaptations in the vertebrate body plan. The musculoskeletal system develops from two mesodermal populations: somitic mesoderm gives rise to the axial skeleton and all of the skeletal muscle of the body, and lateral plate mesoderm gives rise to the appendicular skeleton. The recognition of embryonic domains resulting from the dynamics of morphogenesis has inspired new terminology based on developmental criteria. Two mesodermal domains are defined, primaxial and abaxial. The primaxial domain includes musculoskeletal structures comprising just somitic cells. The abaxial domain contains somitic myoblasts in connective tissue derived from lateral plate mesoderm, as well as lateral plate-derived skeletal structures. The boundary between these two domains is the lateral somitic frontier. Recent studies have described the developmental relationship between these two domains in the chick. In the present study, we describe the labelling pattern in the body of the Prx1/Cre/Z/AP compound transgenic mouse. The enhancer employed in this transgenic leads to reporter expression in the postcranial, somatic lateral plate mesoderm. The boundary between labelled and unlabelled cell populations is described at embryonic day (E)13.5 and E15.5. We argue that the distribution of labelled cells is consistent with the somatic lateral plate lineage, and therefore provides an estimate of the position of the lateral somitic frontier. The role of the frontier in both development and evolution is discussed. PMID:18430087

  15. The repressor function of snail is required for Drosophila gastrulation and is not replaceable by Escargot or Worniu.

    PubMed

    Hemavathy, Kirugaval; Hu, Xiaodi; Ashraf, Shovon I; Small, Stephen J; Ip, Y Tony

    2004-05-15

    Mesoderm formation in the Drosophila embryo depends on the maternal Toll signaling pathway. The Toll pathway establishes the Dorsal nuclear gradient, which regulates many zygotic genes to establish the mesodermal fate and promote the invagination of ventral cells. An important target gene of Dorsal is snail, which is required for proper mesoderm invagination. The Snail protein contains five zinc fingers and is a transcriptional repressor. However, it is not clear whether repressing target genes is a requirement for Snail to control ventral invagination. To examine such requirement, we conducted a series of genetic rescue experiments in snail mutant embryos. Snail, Worniu, and Escargot are closely related zinc-finger proteins and have equal functions during neuroblast development. However, among these three proteins, only Snail can rescue the mesoderm invagination phenotype. Moreover, the ability of various Snail mutant constructs to repress gene expression correlates with their ability to control invagination. This unique property of Snail in mesoderm formation can be attributed mostly to the CtBP co-repressor interaction motifs in the N-terminus, not to the C-terminal DNA-binding zinc fingers. Ectopic expression of Snail outside the ventral domain is not sufficient to induce cell movement even though repression of target genes still occurs. Together, the results show that the repressor function of Snail is essential for gastrulation. The repression of target genes by Snail may permit other factors in the ventral cells to positively promote mesoderm invagination. PMID:15110709

  16. Retinoic Acid Activity in Undifferentiated Neural Progenitors Is Sufficient to Fulfill Its Role in Restricting Fgf8 Expression for Somitogenesis

    PubMed Central

    Cunningham, Thomas J.; Brade, Thomas; Sandell, Lisa L.; Lewandoski, Mark; Trainor, Paul A.; Colas, Alexandre; Mercola, Mark; Duester, Gregg

    2015-01-01

    Bipotent axial stem cells residing in the caudal epiblast during late gastrulation generate neuroectodermal and presomitic mesodermal progeny that coordinate somitogenesis with neural tube formation, but the mechanism that controls these two fates is not fully understood. Retinoic acid (RA) restricts the anterior extent of caudal fibroblast growth factor 8 (Fgf8) expression in both mesoderm and neural plate to control somitogenesis and neurogenesis, however it remains unclear where RA acts to control the spatial expression of caudal Fgf8. Here, we found that mouse Raldh2-/- embryos, lacking RA synthesis and displaying a consistent small somite defect, exhibited abnormal expression of key markers of axial stem cell progeny, with decreased Sox2+ and Sox1+ neuroectodermal progeny and increased Tbx6+ presomitic mesodermal progeny. The Raldh2-/- small somite defect was rescued by treatment with an FGF receptor antagonist. Rdh10 mutants, with a less severe RA synthesis defect, were found to exhibit a small somite defect and anterior expansion of caudal Fgf8 expression only for somites 1–6, with normal somite size and Fgf8 expression thereafter. Rdh10 mutants were found to lack RA activity during the early phase when somites are small, but at the 6-somite stage RA activity was detected in neural plate although not in presomitic mesoderm. Expression of a dominant-negative RA receptor in mesoderm eliminated RA activity in presomitic mesoderm but did not affect somitogenesis. Thus, RA activity in the neural plate is sufficient to prevent anterior expansion of caudal Fgf8 expression associated with a small somite defect. Our studies provide evidence that RA restriction of Fgf8 expression in undifferentiated neural progenitors stimulates neurogenesis while also restricting the anterior extent of the mesodermal Fgf8 mRNA gradient that controls somite size, providing new insight into the mechanism that coordinates somitogenesis with neurogenesis. PMID:26368825

  17. BRACHYURY and CDX2 Mediate BMP-Induced Differentiation of Human and Mouse Pluripotent Stem Cells into Embryonic and Extraembryonic Lineages

    PubMed Central

    Bernardo, Andreia S.; Faial, Tiago; Gardner, Lucy; Niakan, Kathy K.; Ortmann, Daniel; Senner, Claire E.; Callery, Elizabeth M.; Trotter, Matthew W.; Hemberger, Myriam; Smith, James C.; Bardwell, Lee; Moffett, Ashley; Pedersen, Roger A.

    2011-01-01

    Summary BMP is thought to induce hESC differentiation toward multiple lineages including mesoderm and trophoblast. The BMP-induced trophoblast phenotype is a long-standing paradox in stem cell biology. Here we readdressed BMP function in hESCs and mouse epiblast-derived cells. We found that BMP4 cooperates with FGF2 (via ERK) to induce mesoderm and to inhibit endoderm differentiation. These conditions induced cells with high levels of BRACHYURY (BRA) that coexpressed CDX2. BRA was necessary for and preceded CDX2 expression; both genes were essential for expression not only of mesodermal genes but also of trophoblast-associated genes. Maximal expression of the latter was seen in the absence of FGF but these cells coexpressed mesodermal genes and moreover they differed in cell surface and epigenetic properties from placental trophoblast. We conclude that BMP induces human and mouse pluripotent stem cells primarily to form mesoderm, rather than trophoblast, acting through BRA and CDX2. PMID:21816365

  18. MiR-24 is required for hematopoietic differentiation of mouse embryonic stem cells.

    PubMed

    Roy, Lynn; Bikorimana, Emmanuel; Lapid, Danica; Choi, Hyewon; Nguyen, Tan; Dahl, Richard

    2015-01-01

    Overexpression of miRNA, miR-24, in mouse hematopoietic progenitors increases monocytic/ granulocytic differentiation and inhibits B cell development. To determine if endogenous miR-24 is required for hematopoiesis, we antagonized miR-24 in mouse embryonic stem cells (ESCs) and performed in vitro differentiations. Suppression of miR-24 resulted in an inability to produce blood and hematopoietic progenitors (HPCs) from ESCs. The phenotype is not a general defect in mesoderm production since we observe production of nascent mesoderm as well as mesoderm derived cardiac muscle and endothelial cells. Results from blast colony forming cell (BL-CFC) assays demonstrate that miR-24 is not required for generation of the hemangioblast, the mesoderm progenitor that gives rise to blood and endothelial cells. However, expression of the transcription factors Runx1 and Scl is greatly reduced, suggesting an impaired ability of the hemangioblast to differentiate. Lastly, we observed that known miR-24 target, Trib3, is upregulated in the miR-24 antagonized embryoid bodies (EBs). Overexpression of Trib3 alone in ESCs was able to decrease HPC production, though not as great as seen with miR-24 knockdown. These results demonstrate an essential role for miR-24 in the hematopoietic differentiation of ESCs. Although many miRNAs have been implicated in regulation of hematopoiesis, this is the first miRNA observed to be required for the specification of mammalian blood progenitors from early mesoderm. PMID:25634354

  19. A Biomechanical Analysis of Ventral Furrow Formation in the Drosophila Melanogaster Embryo

    PubMed Central

    Conte, Vito; Ulrich, Florian; Baum, Buzz; Muñoz, Jose; Veldhuis, Jim; Brodland, Wayne; Miodownik, Mark

    2012-01-01

    The article provides a biomechanical analysis of ventral furrow formation in the Drosophila melanogaster embryo. Ventral furrow formation is the first large-scale morphogenetic movement in the fly embryo. It involves deformation of a uniform cellular monolayer formed following cellularisation, and has therefore long been used as a simple system in which to explore the role of mechanics in force generation. Here we use a quantitative framework to carry out a systematic perturbation analysis to determine the role of each of the active forces observed. The analysis confirms that ventral furrow invagination arises from a combination of apical constriction and apical–basal shortening forces in the mesoderm, together with a combination of ectodermal forces. We show that the mesodermal forces are crucial for invagination: the loss of apical constriction leads to a loss of the furrow, while the mesodermal radial shortening forces are the primary cause of the internalisation of the future mesoderm as the furrow rises. Ectodermal forces play a minor but significant role in furrow formation: without ectodermal forces the furrow is slower to form, does not close properly and has an aberrant morphology. Nevertheless, despite changes in the active mesodermal and ectodermal forces lead to changes in the timing and extent of furrow, invagination is eventually achieved in most cases, implying that the system is robust to perturbation and therefore over-determined. PMID:22511944

  20. BMP-SMAD signaling: From pluripotent stem cells to cardiovascular commitment.

    PubMed

    Orlova, Valeria V; Chuva de Sousa Lopes, Susana; Valdimarsdottir, Gudrun

    2016-02-01

    Human pluripotent stem cells (hPSCs) can form all somatic cells of the body. They thus offer opportunities for understanding (i) the basic steps of early human development, (ii) the pathophysiology in human degenerative diseases and (iii) approaches to regenerative medicine and drug development. Methods for improving their differentiation to defined mesodermal derivatives in particular will benefit their use in all of these areas but most particularly applications that require cardiac and vascular tissue. However, the molecular mechanisms that regulate mesodermal development in humans are still poorly understood. Gene ablation studies in mice have shown that the signaling pathways activated by the transforming growth factor beta (TGFβ) superfamily, including the bone morphogenetic proteins (BMP), play crucial roles in mesoderm differentiation and patterning the early embryo. Understanding their interplay and interaction with other signaling pathways, how they activate and inhibit transcription factors and epigenetic regulators during self-renewal, maintenance and exit from pluripotency and differentiation could provide vital information for a range of applications. This includes disease modeling when the hPSCs are derived from patients or drug screens for diseases of mesodermal organs. Here, we review the role of the BMP-SMAD signaling pathway in pluripotent stem cells and during mesoderm differentiation with focus on the cells that make up the cardiovascular system. PMID:26651597

  1. What are Head Cavities? - A History of Studies on Vertebrate Head Segmentation.

    PubMed

    Kuratani, Shigeru; Adachi, Noritaka

    2016-06-01

    Motivated by the discovery of segmental epithelial coeloms, or "head cavities," in elasmobranch embryos toward the end of the 19th century, the debate over the presence of mesodermal segments in the vertebrate head became a central problem in comparative embryology. The classical segmental view assumed only one type of metamerism in the vertebrate head, in which each metamere was thought to contain one head somite and one pharyngeal arch, innervated by a set of cranial nerves serially homologous to dorsal and ventral roots of spinal nerves. The non-segmental view, on the other hand, rejected the somite-like properties of head cavities. A series of small mesodermal cysts in early Torpedo embryos, which were thought to represent true somite homologs, provided a third possible view on the nature of the vertebrate head. Recent molecular developmental data have shed new light on the vertebrate head problem, explaining that head mesoderm evolved, not by the modification of rostral somites of an amphioxus-like ancestor, but through the polarization of unspecified paraxial mesoderm into head mesoderm anteriorly and trunk somites posteriorly. PMID:27268975

  2. Quantitative phosphoproteome analysis of embryonic stem cell differentiation toward blood

    PubMed Central

    Piazzi, Manuela; Williamson, Andrew; Lee, Chia-Fang; Pearson, Stella; Lacaud, Georges; Kouskoff, Valerie; McCubrey, James A.; Cocco, Lucio; Whetton, Anthony D.

    2015-01-01

    Murine embryonic stem (ES) cells can differentiate in vitro into three germ layers (endodermic, mesodermic, ectodermic). Studies on the differentiation of these cells to specific early differentiation stages has been aided by an ES cell line carrying the Green Fluorescent Protein (GFP) targeted to the Brachyury (Bry) locus which marks mesoderm commitment. Furthermore, expression of the Vascular Endothelial Growth Factor receptor 2 (Flk1) along with Bry defines hemangioblast commitment. Isobaric-tag for relative and absolute quantification (iTRAQTM) and phosphopeptide enrichment coupled to liquid chromatography separation and mass spectrometry allow the study of phosphorylation changes occurring at different stages of ES cell development using Bry and Flk1 expression respectively. We identified and relatively quantified 37 phosphoentities which are modulated during mesoderm-induced ES cells differentiation, comparing epiblast-like, early mesoderm and hemangioblast-enriched cells. Among the proteins differentially phosphorylated toward mesoderm differentiation were: the epigenetic regulator Dnmt3b, the protein kinase GSK3b, the chromatin remodeling factor Smarcc1, the transcription factor Utf1; as well as protein specifically related to stem cell differentiation, as Eomes, Hmga2, Ints1 and Rif1. As most key factors regulating early hematopoietic development have also been implicated in various types of leukemia, understanding the post-translational modifications driving their regulation during normal development could result in a better comprehension of their roles during abnormal hematopoiesis in leukemia. PMID:25890499

  3. Regulation of Xenopus gastrulation by ErbB signaling

    PubMed Central

    Nie, Shuyi; Chang, Chenbei

    2016-01-01

    During Xenopus gastrulation, mesendodermal cells are internalized and display different movements. Head mesoderm migrates along the blastocoel roof, while trunk mesoderm undergoes convergent extension (C&E). Different signals are implicated in these processes. Our previous studies reveal that signals through ErbB receptor tyrosine kinases modulate Xenopus gastrulation, but the mechanisms employed are not understood. Here we report that ErbB signals control both C&E and head mesoderm migration. Inhibition of ErbB pathway blocks elongation of dorsal marginal zone explants and activin-treated animal caps without removing mesodermal gene expression. Bipolar cell shape and cell mixing in the dorsal region are impaired. Inhibition of ErbB signaling also interferes with migration of prechordal mesoderm on fibronectin. Cell–cell and cell–matrix interaction and cell spreading are reduced when ErbB signaling is blocked. Using antisense morpholino oligonucleotides, we show that ErbB4 is involved in Xenopus gastrulation morphogenesis, and it partially regulates cell movements through modulation of cell adhesion and membrane protrusions. Our results reveal for the first time that vertebrate ErbB signaling modulates gastrulation movements, thus providing a novel pathway, in addition to non-canonical Wnt and FGF signals, that controls gastrulation. We further demonstrate that regulation of cell adhesive properties and cell morphology may underlie the functions of ErbBs in gastrulation. PMID:17134691

  4. Gastrulation EMT Is Independent of P-Cadherin Downregulation

    PubMed Central

    Moly, Pricila K.; Cooley, James R.; Zeltzer, Sebastian L.; Yatskievych, Tatiana A.

    2016-01-01

    Epithelial-mesenchymal transition (EMT) is an evolutionarily conserved process during which cells lose epithelial characteristics and gain a migratory phenotype. Although downregulation of epithelial cadherins by Snail and other transcriptional repressors is generally considered a prerequisite for EMT, recent studies have challenged this view. Here we investigate the relationship between E-cadherin and P-cadherin expression and localization, Snail function and EMT during gastrulation in chicken embryos. Expression analyses show that while E-cadherin transcripts are detected in the epiblast but not in the primitive streak or mesoderm, P-cadherin mRNA and protein are present in the epiblast, primitive and mesoderm. Antibodies that specifically recognize E-cadherin are not presently available. During EMT, P-cadherin relocalizes from the lateral surfaces of epithelial epiblast cells to a circumferential distribution in emerging mesodermal cells. Cells electroporated with an E-cadherin expression construct undergo EMT and migrate into the mesoderm. An examination of Snail function showed that reduction of Slug (SNAI2) protein levels using a morpholino fails to inhibit EMT, and expression of human or chicken Snail in epiblast cells fails to induce EMT. In contrast, cells expressing the Rho inhibitor peptide C3 rapidly exit the epiblast without activating Slug or the mesoderm marker N-cadherin. Together, these experiments show that epiblast cells undergo EMT while retaining P-cadherin, and raise questions about the mechanisms of EMT regulation during avian gastrulation. PMID:27097030

  5. Prune belly syndrome associated with cloacal anomaly, patent urachal remnant, and omphalocele in a female infant.

    PubMed

    Giuliani, Stefano; Vendryes, Christopher; Malhotra, Ajay; Shaul, Donald B; Anselmo, Dean M

    2010-11-01

    Prune belly syndrome (PBS), megacystis-microcolon-intestinal hypoperistalsis (MMIH), and omphalocele-exstrophy of the bladder-imperforate anus-spine abnormalities complex (OEIS) are rare congenital malformations of the newborn that lead to incomplete formation of the gastrointestinal and genitourinary tract systems. To date, incomplete mesodermal development is identified as the cause for all these complex genetic syndromes even if the etiology is still unknown. We present an original case sharing characteristics common to PBS, MMIH, and OEIS complex, without a clear inclination toward any particular one. This case hints toward a common pathway in the creation of the 3 syndromes. We hypothesize that they are a spectrum of malformations based on the time frame when the mesoderm fails to create a normal interaction between infraumbilical mesoderm, urorectal septum, lumbosacral somites in the formation of the abdominal wall and the genitourinary and lower gastrointestinal tracts. PMID:21034928

  6. Interfering with Wnt signalling alters the periodicity of the segmentation clock

    PubMed Central

    Gibb, Sarah; Zagorska, Anna; Melton, Kristin; Tenin, Gennady; Vacca, Irene; Trainor, Paul; Maroto, Miguel; Dale, J. Kim

    2009-01-01

    Somites are embryonic precursors of the ribs, vertebrae and certain dermis tissue. Somite formation is a periodic process regulated by a molecular clock which drives cyclic expression of a number of clock genes in the presomitic mesoderm. To date the mechanism regulating the period of clock gene oscillations is unknown. Here we show that chick homologues of the Wnt pathway genes that oscillate in mouse do not cycle across the chick presomitic mesoderm. Strikingly we find that modifying Wnt signalling changes the period of Notch driven oscillations in both mouse and chick but these oscillations continue. We propose that the Wnt pathway is a conserved mechanism that is involved in regulating the period of cyclic gene oscillations in the presomitic mesoderm. PMID:19272372

  7. Early embryonic expression patterns of the mouse Flamingo and Prickle orthologues.

    PubMed

    Crompton, Lucy A; Du Roure, Camille; Rodriguez, Tristan A

    2007-11-01

    The Drosophila melanogaster proteins Flamingo and Prickle act in the planar cell polarity (PCP) pathway, which is required for acquisition of epithelial polarity in the wing, eye, and epidermis. In mammals, PCP signaling has been shown to regulate cell movements and polarity in a variety of tissues. Here, we show that the murine Flamingo orthologues Celsr1-3 and the Prickle orthologues Prickle1, Prickle2, and Testin have dynamic patterns of expression during pregastrulation and gastrulation stages. Celsr1 is expressed in the anterior visceral endoderm and nascent mesoderm, Celsr2 and Celsr3 mark the prospective neuroectoderm, Prickle1 is expressed in the primitive streak and mesoderm, Prickle2 in the node, and Testin in the anterior visceral endoderm, the extraembryonic ectoderm, primitive streak, and mesoderm. Analysis of a gene-trap mutation in Testin indicates that this gene is not required for embryogenesis; therefore, other Prickle homologues may compensate for its function during development. PMID:17937400

  8. Developmental angiogenesis: quail embryonic vasculature.

    PubMed

    Poole, T J; Coffin, J D

    1988-03-01

    We have examined the segregation and early morphogenesis of the embryonic vasculature by using a monoclonal antibody for immunofluorescence and by scanning electron microscopy. This antibody labels the presumptive endothelial cells (PECs) as they segregate from mesoderm. Similar embryos prepared for SEM revealed finer details of how these segregated cells interact to form the rudiments of the major blood vessels. Here we concentrate on the development of the dorsal aortae and the posterior cardinal veins. The dorsal aortae form from single PECs which segregate from the lateral mesoderm and aggregate into a loose cord ventral to the somites. These cells become more closely associated and a lumen forms. The posterior cardinal veins form from a loose plexus of cells segregated from the lateral mesoderm on its dorsal surface. These cells become intimately associated with the Wolffian ducts. PMID:3285464

  9. Retinoic acid regulates size, pattern and alignment of tissues at the head-trunk transition.

    PubMed

    Lee, Keun; Skromne, Isaac

    2014-11-01

    At the head-trunk transition, hindbrain and spinal cord alignment to occipital and vertebral bones is crucial for coherent neural and skeletal system organization. Changes in neural or mesodermal tissue configuration arising from defects in the specification, patterning or relative axial placement of territories can severely compromise their integration and function. Here, we show that coordination of neural and mesodermal tissue at the zebrafish head-trunk transition crucially depends on two novel activities of the signaling factor retinoic acid (RA): one specifying the size and the other specifying the axial position relative to mesodermal structures of the hindbrain territory. These activities are each independent but coordinated with the well-established function of RA in hindbrain patterning. Using neural and mesodermal landmarks we demonstrate that the functions of RA in aligning neural and mesodermal tissues temporally precede the specification of hindbrain and spinal cord territories and the activation of hox transcription. Using cell transplantation assays we show that RA activity in the neuroepithelium regulates hindbrain patterning directly and territory size specification indirectly. This indirect function is partially dependent on Wnts but independent of FGFs. Importantly, RA specifies and patterns the hindbrain territory by antagonizing the activity of the spinal cord specification gene cdx4; loss of Cdx4 rescues the defects associated with the loss of RA, including the reduction in hindbrain size and the loss of posterior rhombomeres. We propose that at the head-trunk transition, RA coordinates specification, patterning and alignment of neural and mesodermal tissues that are essential for the organization and function of the neural and skeletal systems. PMID:25371368

  10. The outflow tract of the heart is recruited from a novel heart-forming field.

    PubMed

    Mjaatvedt, C H; Nakaoka, T; Moreno-Rodriguez, R; Norris, R A; Kern, M J; Eisenberg, C A; Turner, D; Markwald, R R

    2001-10-01

    As classically described, the precardiac mesoderm of the paired heart-forming fields migrate and fuse anteriomedially in the ventral midline to form the first segment of the straight heart tube. This segment ultimately forms the right trabeculated ventricle. Additional segments are added to the caudal end of the first in a sequential fashion from the posteriolateral heart-forming field mesoderm. In this study we report that the final major heart segment, which forms the cardiac outflow tract, does not follow this pattern of embryonic development. The cardiac outlet, consisting of the conus and truncus, does not derive from the paired heart-forming fields, but originates separately from a previously unrecognized source of mesoderm located anterior to the initial primitive heart tube segment. Fate-mapping results show that cells labeled in the mesoderm surrounding the aortic sac and anterior to the primitive right ventricle are incorporated into both the conus and the truncus. Conversely, if cells are labeled in the existing right ventricle no incorporation into the cardiac outlet is observed. Tissue explants microdissected from this anterior mesoderm region are capable of forming beating cardiac muscle in vitro when cocultured with explants of the primitive right ventricle. These findings establish the presence of another heart-forming field. This anterior heart-forming field (AHF) consists of mesoderm surrounding the aortic sac immediately anterior to the existing heart tube. This new concept of the heart outlet's embryonic origin provides a new basis for explaining a variety of gene-expression patterns and cardiac defects described in both transgenic animals and human congenital heart disease. PMID:11783996

  11. Isolation of Sna, a mouse gene homologous to the Drosophila genes snail and escargot: its expression pattern suggests multiple roles during postimplantation development.

    PubMed

    Smith, D E; Franco del Amo, F; Gridley, T

    1992-12-01

    The Drosophila gene snail encodes a zinc-finger protein that is required zygotically for mesoderm formation. Snail acts as a transcriptional repressor during the period of mesoderm formation by preventing expression of mesectodermal and ectodermal genes in the mesoderm anlage. A Xenopus homolog (xsnail) of snail has been cloned and it too is expressed early in the mesodermal germ layer. We have isolated cDNA clones of a mouse gene (termed Sna) closely related to snail and xsnail and another Drosophila gene termed escargot that also encodes a zinc-finger protein. Sna encodes a 264 amino acid protein that contains four zinc fingers. Developmental RNA blot analysis showed that Sna transcripts are expressed throughout postimplantation development. Analysis of the spatial and temporal localization of Sna transcripts by in situ hybridization to both whole-mount and sectioned embryos revealed that, in the gastrulating embryo, Sna is expressed throughout the primitive streak and in the entire mesodermal germ layer. By 9.5 days post coitum (dpc) Sna is expressed at high levels in cephalic neural crest and limb bud mesenchyme. In fact, by 10.5 dpc Sna expression is observed in most mesenchymal cells, whether of neural crest or mesodermal origin. Later in gestation, high levels of Sna expression are observed in condensing cartilage and in the mesenchymal component of several tissues (lung, kidney, teeth and vibrissae) that undergo epithelial-mesenchymal inductive interactions during development. These results suggest multiple roles for the Sna gene in gastrulation and organogenesis during murine development. PMID:1295727

  12. Nkx3.2 AND HEDGEHOG PATHWAY INTERSECT TO PATTERN AXIAL SKELETON

    Technology Transfer Automated Retrieval System (TEKTRAN)

    BACKGROUND: NK homeobox family members are tissue-specific transcription factors important for regulation of developmental genes. Nkx3.2 is one of the earliest markers of sclerotome and gut mesoderm. Targeted disruption of Nkx3.2 expression in mice produced severe developmental defects, including ma...

  13. Position-dependent plasticity of distinct progenitor types in the primitive streak

    PubMed Central

    Wymeersch, Filip J; Huang, Yali; Blin, Guillaume; Cambray, Noemí; Wilkie, Ron; Wong, Frederick CK; Wilson, Valerie

    2016-01-01

    The rostrocaudal (head-to-tail) axis is supplied by populations of progenitors at the caudal end of the embryo. Despite recent advances characterising one of these populations, the neuromesodermal progenitors, their nature and relationship to other populations remains unclear. Here we show that neuromesodermal progenitors are a single Sox2lowTlow entity whose choice of neural or mesodermal fate is dictated by their position in the progenitor region. The choice of mesoderm fate is Wnt/β-catenin dependent. Wnt/β-catenin signalling is also required for a previously unrecognised phase of progenitor expansion during mid-trunk formation. Lateral/ventral mesoderm progenitors represent a distinct committed state that is unable to differentiate to neural fates, even upon overexpression of the neural transcription factor Sox2. They do not require Wnt/β-catenin signalling for mesoderm differentiation. This information aids the correct interpretation of in vivo genetic studies and the development of in vitro protocols for generating physiologically-relevant cell populations of clinical interest. DOI: http://dx.doi.org/10.7554/eLife.10042.001 PMID:26780186

  14. Somites in zebrafish doubly mutant for knypek and trilobite form without internal mesenchymal cells or compaction.

    PubMed

    Henry, C A; Hall, L A; Burr Hille, M; Solnica-Krezel, L; Cooper, M S

    2000-09-01

    In vertebrates, paraxial mesoderm is partitioned into repeating units called somites. It is thought that the mechanical forces arising from compaction of the presumptive internal cells of prospective somites cause them to detach from the unsegmented presomitic mesoderm [1-3]. To determine how prospective somites physically segregate from each other, we used time-lapse microscopy to analyze the mechanics underlying early somitogenesis in wild-type zebrafish and in the mutants trilobite(m209) (tri), knypek(m119) (kny), and kny;tri, which are defective in convergent extension during gastrulation. Formation of somite boundaries in all of these embryos involved segregation, local alignment, and cell-shape changes of presumptive epitheloid border cells along nascent intersomitic boundaries. Although kny;tri somites formed without convergence of the presomitic mesoderm and were composed of only two cells in their anteroposterior (AP) dimension, they still exhibited AP intrasegmental polarity. Furthermore, morphogenesis of somite boundaries in these embryos proceeded in a manner similar to that in wild-type embryos. Thus, intersomitic boundary formation in zebrafish involves short-range movements of presumptive border cells that do not require mechanical forces generated by internal cells or compaction of the presomitic mesoderm. PMID:10996075

  15. Red light, green light: Signals that control endothelial cell proliferation during embryonic vascular development

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The proper regulation of endothelial cell proliferation is critical for vascular development in the embryo. VEGF-A and bFGF, which are important in the induction of mesodermal progenitors to form a capillary plexus, are also key mitogenic signals. Disruption in VEGF-A or bFGF decreases endothelial c...

  16. Distinct modes of floor plate induction in the chick embryo.

    PubMed

    Patten, Iain; Kulesa, Paul; Shen, Michael M; Fraser, Scott; Placzek, Marysia

    2003-10-01

    To begin to reconcile models of floor plate formation in the vertebrate neural tube, we have performed experiments aimed at understanding the development of the early floor plate in the chick embryo. Using real-time analyses of cell behaviour, we provide evidence that the principal contributor to the early neural midline, the future anterior floor plate, exists as a separate population of floor plate precursor cells in the epiblast of the gastrula stage embryo, and does not share a lineage with axial mesoderm. Analysis of the tissue interactions associated with differentiation of these cells to a floor plate fate reveals a role for the nascent prechordal mesoderm, indicating that more than one inductive event is associated with floor plate formation along the length of the neuraxis. We show that Nr1, a chick nodal homologue, is expressed in the nascent prechordal mesoderm and we provide evidence that Nodal signalling can cooperate with Shh to induce the epiblast precursors to a floor-plate fate. These results indicate that a shared lineage with axial mesoderm cells is not a pre-requisite for floor plate differentiation and suggest parallels between the development of the floor plate in amniote and anamniote embryos. PMID:12917296

  17. Polarization of PI3K Activity Initiated by Ooplasmic Segregation Guides Nuclear Migration in the Mesendoderm.

    PubMed

    Takatori, Naohito; Oonuma, Kouhei; Nishida, Hiroki; Saiga, Hidetoshi

    2015-11-01

    Asymmetric localization of RNA is a widely observed mechanism of cell polarization. Using embryos of the ascidian, Halocynthia roretzi, we previously showed that mesoderm and endoderm fates are separated by localization of mRNA encoding a transcription factor, Not, to the future mesoderm-side cytoplasm of the mesendoderm cell through asymmetric positioning of the nucleus. Here, we investigated the mechanism that defines the direction of the nuclear migration. We show that localization of PtdIns(3,4,5)P3 to the future mesoderm region determines the direction of nuclear migration. Localization of PtdIns(3,4,5)P3 was dependent on the localization of PI3Kα to the future mesoderm region. PI3Kα was first localized at the 1-cell stage by the ooplasmic movement. Activity of localized PI3Kα at the 4-cell stage was required for the localization of PI3Kα up to the nuclear migration. Our results provide the scaffold for understanding the chain of causality leading to the separation of germ layer fates. PMID:26555053

  18. A novel gain-of-function mutation of the proneural IRX1 and IRX2 genes disrupts axis elongation in the Araucana rumpless chicken.

    PubMed

    Freese, Nowlan H; Lam, Brianna A; Staton, Meg; Scott, Allison; Chapman, Susan C

    2014-01-01

    Axis elongation of the vertebrate embryo involves the generation of cell lineages from posterior progenitor populations. We investigated the molecular mechanism governing axis elongation in vertebrates using the Araucana rumpless chicken. Araucana embryos exhibit a defect in axis elongation, failing to form the terminal somites and concomitant free caudal vertebrae, pygostyle, and associated tissues of the tail. Through whole genome sequencing of six Araucana we have identified a critical 130 kb region, containing two candidate causative SNPs. Both SNPs are proximal to the IRX1 and IRX2 genes, which are required for neural specification. We show that IRX1 and IRX2 are both misexpressed within the bipotential chordoneural hinge progenitor population of Araucana embryos. Expression analysis of BRA and TBX6, required for specification of mesoderm, shows that both are downregulated, whereas SOX2, required for neural patterning, is expressed in ectopic epithelial tissue. Finally, we show downregulation of genes required for the protection and maintenance of the tailbud progenitor population from the effects of retinoic acid. Our results support a model where the disruption in balance of mesoderm and neural fate results in early depletion of the progenitor population as excess neural tissue forms at the expense of mesoderm, leading to too few mesoderm cells to form the terminal somites. Together this cascade of events leads to axis truncation. PMID:25372603

  19. A Novel Gain-Of-Function Mutation of the Proneural IRX1 and IRX2 Genes Disrupts Axis Elongation in the Araucana Rumpless Chicken

    PubMed Central

    Freese, Nowlan H.; Lam, Brianna A.; Staton, Meg; Scott, Allison; Chapman, Susan C.

    2014-01-01

    Axis elongation of the vertebrate embryo involves the generation of cell lineages from posterior progenitor populations. We investigated the molecular mechanism governing axis elongation in vertebrates using the Araucana rumpless chicken. Araucana embryos exhibit a defect in axis elongation, failing to form the terminal somites and concomitant free caudal vertebrae, pygostyle, and associated tissues of the tail. Through whole genome sequencing of six Araucana we have identified a critical 130 kb region, containing two candidate causative SNPs. Both SNPs are proximal to the IRX1 and IRX2 genes, which are required for neural specification. We show that IRX1 and IRX2 are both misexpressed within the bipotential chordoneural hinge progenitor population of Araucana embryos. Expression analysis of BRA and TBX6, required for specification of mesoderm, shows that both are downregulated, whereas SOX2, required for neural patterning, is expressed in ectopic epithelial tissue. Finally, we show downregulation of genes required for the protection and maintenance of the tailbud progenitor population from the effects of retinoic acid. Our results support a model where the disruption in balance of mesoderm and neural fate results in early depletion of the progenitor population as excess neural tissue forms at the expense of mesoderm, leading to too few mesoderm cells to form the terminal somites. Together this cascade of events leads to axis truncation. PMID:25372603

  20. EphrinB/EphB Signaling Controls Embryonic Germ Layer Separation by Contact-Induced Cell Detachment

    PubMed Central

    Rohani, Nazanin; Canty, Laura; Luu, Olivia

    2011-01-01

    Background The primordial organization of the metazoan body is achieved during gastrulation by the establishment of the germ layers. Adhesion differences between ectoderm, mesoderm, and endoderm cells could in principle be sufficient to maintain germ layer integrity and prevent intermixing. However, in organisms as diverse as fly, fish, or amphibian, the ectoderm-mesoderm boundary not only keeps these germ layers separated, but the ectoderm also serves as substratum for mesoderm migration, and the boundary must be compatible with repeated cell attachment and detachment. Principal Findings We show that localized detachment resulting from contact-induced signals at the boundary is at the core of ectoderm-mesoderm segregation. Cells alternate between adhesion and detachment, and detachment requires ephrinB/EphB signaling. Multiple ephrinB ligands and EphB receptors are expressed on each side of the boundary, and tissue separation depends on forward signaling across the boundary in both directions, involving partially redundant ligands and receptors and activation of Rac and RhoA. Conclusion This mechanism differs from a simple differential adhesion process of germ layer formation. Instead, it involves localized responses to signals exchanged at the tissue boundary and an attachment/detachment cycle which allows for cell migration across a cellular substratum. PMID:21390298

  1. Expression pattern of annelid Zic in embryonic development of the oligochaete Tubifex tubifex.

    PubMed

    Takahashi, Hirokazu; Shimizu, Takashi; Aruga, Jun

    2008-10-01

    Embryonic expression of a Zic homologue (Ttu-Zic) was examined in the oligochaete annelid Tubifex tubifex. The body plan of T. tubifex is characterized by obvious segmentation in the ectoderm and mesoderm. Ttu-Zic expression is detected in the mesodermal germ band and a subset of micromere descendants. Ttu-Zic is transiently expressed in primary m-blast cells (i.e., founder cells of mesodermal segments) as early as the time of their birth from M teloblasts. During its development, each mesodermal segment experiences two additional phases of Ttu-Zic expression. Ttu-Zic expression in micromere descendants is seen on the anterior surfaces of embryos undergoing teloblastogenesis; subsequently, these cells proliferate to form bilateral clusters, which then become internalized. Finally, clusters of Ttu-Zic-expressing cells are found in the center of the prostomium, corresponding to the cerebral ganglion. The Ttu-Zic expression profile in the early embryogenesis of T. tubifex may be homologous to those of evolutionarily distant animals. PMID:18810489

  2. Embryological Development: Evolutionary History, Genetic Bias, and Cellular Environment Control the Flow of Developmental Events, Part II.

    ERIC Educational Resources Information Center

    Caplan, Arnold I.

    1981-01-01

    Emphasizes ectodermal-mesodermal interaction but focuses on the genesis of specialized structures like feathers (ectodermal) and muscles, cartilage, and bone. The sum of these interactions and other factors which govern normal development may be important in regulating the regeneration of particular structures in postembryonic individuals.…

  3. Evolution of the head-trunk interface in tetrapod vertebrates

    PubMed Central

    Sefton, Elizabeth M; Bhullar, Bhart-Anjan S; Mohaddes, Zahra; Hanken, James

    2016-01-01

    Vertebrate neck musculature spans the transition zone between head and trunk. The extent to which the cucullaris muscle is a cranial muscle allied with the gill levators of anamniotes or is instead a trunk muscle is an ongoing debate. Novel computed tomography datasets reveal broad conservation of the cucullaris in gnathostomes, including coelacanth and caecilian, two sarcopterygians previously thought to lack it. In chicken, lateral plate mesoderm (LPM) adjacent to occipital somites is a recently identified embryonic source of cervical musculature. We fate-map this mesoderm in the axolotl (Ambystoma mexicanum), which retains external gills, and demonstrate its contribution to posterior gill-levator muscles and the cucullaris. Accordingly, LPM adjacent to the occipital somites should be regarded as posterior cranial mesoderm. The axial position of the head-trunk border in axolotl is congruent between LPM and somitic mesoderm, unlike in chicken and possibly other amniotes. DOI: http://dx.doi.org/10.7554/eLife.09972.001 PMID:27090084

  4. Position-dependent plasticity of distinct progenitor types in the primitive streak.

    PubMed

    Wymeersch, Filip J; Huang, Yali; Blin, Guillaume; Cambray, Noemí; Wilkie, Ron; Wong, Frederick Ck; Wilson, Valerie

    2016-01-01

    The rostrocaudal (head-to-tail) axis is supplied by populations of progenitors at the caudal end of the embryo. Despite recent advances characterising one of these populations, the neuromesodermal progenitors, their nature and relationship to other populations remains unclear. Here we show that neuromesodermal progenitors are a single Sox2(low)T(low) entity whose choice of neural or mesodermal fate is dictated by their position in the progenitor region. The choice of mesoderm fate is Wnt/β-catenin dependent. Wnt/β-catenin signalling is also required for a previously unrecognised phase of progenitor expansion during mid-trunk formation. Lateral/ventral mesoderm progenitors represent a distinct committed state that is unable to differentiate to neural fates, even upon overexpression of the neural transcription factor Sox2. They do not require Wnt/β-catenin signalling for mesoderm differentiation. This information aids the correct interpretation of in vivo genetic studies and the development of in vitro protocols for generating physiologically-relevant cell populations of clinical interest. PMID:26780186

  5. PAPC mediates self/non–self-distinction during Snail1-dependent tissue separation

    PubMed Central

    Luu, Olivia; Damm, Erich W.; Parent, Serge E.; Barua, Debanjan; Smith, Tamara H.L.; Wen, Jason W.H.; Lepage, Stephanie E.; Nagel, Martina; Ibrahim-Gawel, Hady; Huang, Yunyun

    2015-01-01

    Cleft-like boundaries represent a type of cell sorting boundary characterized by the presence of a physical gap between tissues. We studied the cleft-like ectoderm–mesoderm boundary in Xenopus laevis and zebrafish gastrulae. We identified the transcription factor Snail1 as being essential for tissue separation, showed that its expression in the mesoderm depends on noncanonical Wnt signaling, and demonstrated that it enables paraxial protocadherin (PAPC) to promote tissue separation through two novel functions. First, PAPC attenuates planar cell polarity signaling at the ectoderm–mesoderm boundary to lower cell adhesion and facilitate cleft formation. Second, PAPC controls formation of a distinct type of adhesive contact between mesoderm and ectoderm cells that shows properties of a cleft-like boundary at the single-cell level. It consists of short stretches of adherens junction–like contacts inserted between intermediate-sized contacts and large intercellular gaps. These roles of PAPC constitute a self/non–self-recognition mechanism that determines the site of boundary formation at the interface between PAPC-expressing and -nonexpressing cells. PMID:25778923

  6. The development of the action potential mechanism of amphibian neurons isolated in culture.

    PubMed

    Spitzer, N C; Lamborghini, J E

    1976-05-01

    Nerve and muscle cells differentiated morphologically, in cultures of dissociated cells prepared from amphibian neural plate and underlying mesoderm (Xenopus laevis, Nieuwkoop and Faber stage 15). Cultures were grown in a defined medium containing sterile Steinberg's salt solution and 0.1% bovine serum albumin, and maintained for periods up to 5 days. PMID:1064036

  7. The development of the action potential mechanism of amphibian neurons isolated in culture.

    PubMed Central

    Spitzer, N C; Lamborghini, J E

    1976-01-01

    Nerve and muscle cells differentiated morphologically, in cultures of dissociated cells prepared from amphibian neural plate and underlying mesoderm (Xenopus laevis, Nieuwkoop and Faber stage 15). Cultures were grown in a defined medium containing sterile Steinberg's salt solution and 0.1% bovine serum albumin, and maintained for periods up to 5 days. Images PMID:1064036

  8. The thin red line: angiogenesis in normal and malignant hematopoiesis.

    PubMed

    Bertolini, F; Mancuso, P; Gobbi, A; Pruneri, G

    2000-09-01

    This review describes the current knowledge about cell subsets involved in vasculogenesis (i.e., differentiation of endothelial cells from mesodermal precursors) and angiogenesis (i.e., blood vessel generation from pre-existing vessels), together with recent findings about angiogenesis and antiangiogenic therapies in hematopoietic malignancies such as leukemia, lymphoma, myeloma, and myelodysplastic syndromes. PMID:11008011

  9. Cell lineage in mammalian craniofacial mesenchyme.

    PubMed

    Yoshida, Toshiyuki; Vivatbutsiri, Philaiporn; Morriss-Kay, Gillian; Saga, Yumiko; Iseki, Sachiko

    2008-01-01

    We have analysed the contributions of neural crest and mesoderm to mammalian craniofacial mesenchyme and its derivatives by cell lineage tracing experiments in mouse embryos, using the permanent genetic markers Wnt1-cre for neural crest and Mesp1-cre for mesoderm, combined with the Rosa26 reporter. At the end of neural crest cell migration (E9.5) the two patterns are reciprocal, with a mutual boundary just posterior to the eye. Mesodermal cells expressing endothelial markers (angioblasts) are found not to respect this boundary; they are associated with the migrating neural crest from the 5-somite stage, and by E9.5 they form a pre-endothelial meshwork throughout the cranial mesenchyme. Mesodermal cells of the myogenic lineage also migrate with neural crest cells, as the branchial arches form. By E17.5 the neural crest-mesoderm boundary in the subectodermal mesenchyme becomes out of register with that of the underlying skeletogenic layer, which is between the frontal and parietal bones. At E13.5 the primordia of these bones lie basolateral to the brain, extending towards the vertex of the skull during the following 4-5 days. We used DiI labelling of the bone primordia in ex-utero E13.5 embryos to distinguish between two possibilities for the origin of the frontal and parietal bones: (1) recruitment from adjacent connective tissue or (2) proliferation of the original primordia. The results clearly demonstrated that the bone primordia extend vertically by intrinsic growth, without detectable recruitment of adjacent mesenchymal cells. PMID:18617001

  10. The emergence of Pax7-expressing muscle stem cells during vertebrate head muscle development

    PubMed Central

    Nogueira, Julia Meireles; Hawrot, Katarzyna; Sharpe, Colin; Noble, Anna; Wood, William M.; Jorge, Erika C.; Goldhamer, David J.; Kardon, Gabrielle; Dietrich, Susanne

    2015-01-01

    Pax7 expressing muscle stem cells accompany all skeletal muscles in the body and in healthy individuals, efficiently repair muscle after injury. Currently, the in vitro manipulation and culture of these cells is still in its infancy, yet muscle stem cells may be the most promising route toward the therapy of muscle diseases such as muscular dystrophies. It is often overlooked that muscular dystrophies affect head and body skeletal muscle differently. Moreover, these muscles develop differently. Specifically, head muscle and its stem cells develop from the non-somitic head mesoderm which also has cardiac competence. To which extent head muscle stem cells retain properties of the early head mesoderm and might even be able to switch between a skeletal muscle and cardiac fate is not known. This is due to the fact that the timing and mechanisms underlying head muscle stem cell development are still obscure. Consequently, it is not clear at which time point one should compare the properties of head mesodermal cells and head muscle stem cells. To shed light on this, we traced the emergence of head muscle stem cells in the key vertebrate models for myogenesis, chicken, mouse, frog and zebrafish, using Pax7 as key marker. Our study reveals a common theme of head muscle stem cell development that is quite different from the trunk. Unlike trunk muscle stem cells, head muscle stem cells do not have a previous history of Pax7 expression, instead Pax7 expression emerges de-novo. The cells develop late, and well after the head mesoderm has committed to myogenesis. We propose that this unique mechanism of muscle stem cell development is a legacy of the evolutionary history of the chordate head mesoderm. PMID:26042028

  11. Small molecule-directed specification of sclerotome-like chondroprogenitors and induction of a somitic chondrogenesis program from embryonic stem cells.

    PubMed

    Zhao, Jiangang; Li, Songhui; Trilok, Suprita; Tanaka, Makoto; Jokubaitis-Jameson, Vanta; Wang, Bei; Niwa, Hitoshi; Nakayama, Naoki

    2014-10-01

    Pluripotent embryonic stem cells (ESCs) generate rostral paraxial mesoderm-like progeny in 5-6 days of differentiation induced by Wnt3a and Noggin (Nog). We report that canonical Wnt signaling introduced either by forced expression of activated β-catenin, or the small-molecule inhibitor of Gsk3, CHIR99021, satisfied the need for Wnt3a signaling, and that the small-molecule inhibitor of BMP type I receptors, LDN193189, was able to replace Nog. Mesodermal progeny generated using such small molecules were chondrogenic in vitro, and expressed trunk paraxial mesoderm markers such as Tcf15 and Meox1, and somite markers such as Uncx, but failed to express sclerotome markers such as Pax1. Induction of the osteochondrogenically committed sclerotome from somite requires sonic hedgehog and Nog. Consistently, Pax1 and Bapx1 expression was induced when the isolated paraxial mesodermal progeny were treated with SAG1 (a hedgehog receptor agonist) and LDN193189, then Sox9 expression was induced, leading to cartilaginous nodules and particles in the presence of BMP, indicative of chondrogenesis via sclerotome specification. By contrast, treatment with TGFβ also supported chondrogenesis and stimulated Sox9 expression, but failed to induce the expression of Pax1 and Bapx1. On ectopic transplantation to immunocompromised mice, the cartilage particles developed under either condition became similarly mineralized and formed pieces of bone with marrow. Thus, the use of small molecules led to the effective generation from ESCs of paraxial mesodermal progeny, and to their further differentiation in vitro through sclerotome specification into growth plate-like chondrocytes, a mechanism resembling in vivo somitic chondrogenesis that is not recapitulated with TGFβ. PMID:25294938

  12. Revisiting the involvement of signaling gradients in somitogenesis.

    PubMed

    Mallo, Moisés

    2016-04-01

    During embryonic development, formation of individual vertebrae requires that the paraxial mesoderm becomes divided into regular segmental units known as somites. Somites are sequentially formed at the anterior end of the presomitic mesoderm (PSM) resulting from functional interactions between the oscillatory activity of signals promoting segmentation and a moving wavefront of tissue competence to those signals, eventually generating a constant flow of new somites at regular intervals. According to the current model for somitogenesis, the wavefront results from the combined activity of two opposing functional gradients in the PSM involving the Fgf, Wnt and retinoic acid (RA) signaling pathways. Here, I use published data to evaluate the wavefront model. A critical analysis of those studies seems to support a role for Wnt signaling, but raise doubts regarding the extent to which Fgf and RA signaling contribute to this process. PMID:26662366

  13. Metastable primordial germ cell-like state induced from mouse embryonic stem cells by Akt activation

    SciTech Connect

    Yamano, Noriko; Kimura, Tohru; Watanabe-Kushima, Shoko; Shinohara, Takashi; Nakano, Toru

    2010-02-12

    Specification to primordial germ cells (PGCs) is mediated by mesoderm-induction signals during gastrulation. We found that Akt activation during in vitro mesodermal differentiation of embryonic stem cells (ESCs) generated self-renewing spheres with differentiation states between those of ESCs and PGCs. Essential regulators for PGC specification and their downstream germ cell-specific genes were expressed in the spheres, indicating that the sphere cells had commenced differentiation to the germ lineage. However, the spheres did not proceed to spermatogenesis after transplantation into testes. Sphere cell transfer to the original feeder-free ESC cultures resulted in chaotic differentiation. In contrast, when the spheres were cultured on mouse embryonic fibroblasts or in the presence of ERK-cascade and GSK3 inhibitors, reversion to the ESC-like state was observed. These results indicate that Akt signaling promotes a novel metastable and pluripotent state that is intermediate to those of ESCs and PGCs.

  14. Teratomas: a multimodality review.

    PubMed

    Peterson, Christine M; Buckley, Celine; Holley, Susan; Menias, Christine O

    2012-01-01

    Germ cell tumors (GCTs) may occur in both children and adults and include a broad array of histologic subtypes, such as teratoma, seminoma (known as dysgerminoma in the ovary and germinoma in the pineal gland), choriocarcinoma, yolk sac tumor, embryonal cell carcinoma, and mixed GCT. In adults, GCTs occur most commonly in the gonads. In children, sacrococcygeal tumors predominate. Teratomas are a common form of GCT. They are defined histologically as containing tissues derived from all 3 germ cell layers: ectoderm, mesoderm (most teratomas contain fat, an imaging hallmark, which is a mesodermal derivative), and endoderm. Teratomas are also classified as mature or immature, depending on the degree of differentiation of its components, and in adults, immature tumors are more likely to exhibit malignant behavior. PMID:23009771

  15. Dose-dependent Nodal/Smad signals pattern the early mouse embryo.

    PubMed

    Robertson, Elizabeth J

    2014-08-01

    Nodal signals in the early post-implantation stage embryo are essential to establish initial proximal-distal (P-D) polarity and generate the final anterior-posterior (A-P) body axis. Nodal signaling in the epiblast results in the phosphorylation of Smad2 in the overlying visceral endoderm necessary to induce the AVE, in part via Smad2-dependent activation of the T-box gene Eomesodermin. Slightly later following mesoderm induction a continuum of dose-dependent Nodal signaling during the process of gastrulation underlies specification of mesodermal and definitive endoderm progenitors. Dynamic Nodal expression during the critical 72 h time window immediately following implantation, accomplished by a series of feed-back and feed-forward mechanisms serves to provide key positional cues required for establishment of the body plan and controls cell fate decisions in the early mammalian embryo. PMID:24704361

  16. Planar induction of anteroposterior pattern in the developing central nervous system of Xenopus laevis

    NASA Technical Reports Server (NTRS)

    Doniach, T.; Phillips, C. R.; Gerhart, J. C.

    1992-01-01

    It has long been thought that anteroposterior (A-P) pattern in the vertebrate central nervous system is induced in the embryo's dorsal ectoderm exclusively by signals passing vertically from underlying, patterned dorsal mesoderm. Explants from early gastrulae of the frog Xenopus laevis were prepared in which vertical contact between dorsal ectoderm and mesoderm was prevented but planar contact was maintained. In these, four position-specific neural markers (engrailed-2, Krox-20, XlHbox 1, and XlHbox 6) were expressed in the ectoderm in the same A-P order as in the embryo. Thus, planar signals alone, following a path available in the normal embryo, can induce A-P neural pattern.

  17. Generation of the germ layers along the animal-vegetal axis in Xenopus laevis.

    PubMed

    Yasuo, H; Lemaire, P

    2001-01-01

    After completion of gastrulation, typical vertebrate embryos consist of three cell sheets, called germ layers. The outer layer, the ectoderm, which produces the cells of the epidermis and the nervous system; the inner layer, the endoderm, producing the lining of the digestive tube and its associated organs (pancreas, liver, lungs etc.) and the middle layer, the mesoderm, which gives rise to several organs (heart, kidney, gonads), connective tissues (bone, muscles, tendons, blood vessels), and blood cells. The formation of the germ layers is one of the earliest embryonic events to subdivide multicellular embryos into a few compartments. In Xenopus laevis, the spatial domains of three germ layers are largely separated along the animal-vegetal axis even before gastrulation; ectoderm in the animal pole region; mesoderm in the equatorial region and endoderm in the vegetal pole region. In this review, we summarise the recent advances in our understanding of the formation of the germ layers in Xenopus laevis. PMID:11291851

  18. SOX17 links gut endoderm morphogenesis and germ layer segregation.

    PubMed

    Viotti, Manuel; Nowotschin, Sonja; Hadjantonakis, Anna-Katerina

    2014-12-01

    Gastrulation leads to three germ layers--ectoderm, mesoderm and endoderm--that are separated by two basement membranes. In the mouse embryo, the emergent gut endoderm results from the widespread intercalation of cells of two distinct origins: pluripotent epiblast-derived definitive endoderm (DE) and extra-embryonic visceral endoderm (VE). Here we image the trajectory of prospective DE cells before intercalating into the VE epithelium. We show that the transcription factor SOX17, which is activated in prospective DE cells before intercalation, is necessary for gut endoderm morphogenesis and the assembly of the basement membrane that separates gut endoderm from mesoderm. Our results mechanistically link gut endoderm morphogenesis and germ layer segregation, two central and conserved features of gastrulation. PMID:25419850

  19. An Fgf-Shh signaling hierarchy regulates early specification of the zebrafish skull

    PubMed Central

    McCarthy, Neil; Sidik, Alfire; Bertrand, Julien Y.; Eberhart, Johann K.

    2016-01-01

    The neurocranium generates most of the craniofacial skeleton and consists of prechordal and postchordal regions. Although development of the prechordal is well studied, little is known of the postchordal region. Here we characterize a signaling hierarchy necessary for postchordal neurocranial development involving Fibroblast growth factor (Fgf) signaling for early specification of mesodermally-derived progenitor cells. The expression of hyaluron synthetase 2 (has2) in the cephalic mesoderm requires Fgf signaling and Has2 function, in turn, is required for postchordal neurocranial development. While Hedgehog (Hh)-deficient embryos also lack a postchordal neurocranium, this appears primarily due to a later defect in chondrocyte differentiation. Inhibitor studies demonstrate that postchordal neurocranial development requires early Fgf and later Hh signaling. Collectively, our results provide a mechanistic understanding of early postchordal neurocranial development and demonstrate a hierarchy of signaling between Fgf and Hh in the development of this structure. PMID:27060628

  20. The Drosophila melanogaster developmental gene g1 encodes a variant zinc-finger-motif protein.

    PubMed

    Bouchard, M L; Côté, S

    1993-03-30

    In Drosophila melanogaster, the mechanisms involved in the pattern formation of complex internal organs are still largely unknown. However, the identity of the molecular determinants that control the development of these specific tissues is emerging from the combined use of genetic and molecular approaches. We have cloned a gene that is expressed in the mesoderm, one of the fundamental embryonic germ layers which gives rise to internal structures, such as the musculature. Here, we describe the molecular characterization of this gene, designated as g1. The nucleotide (nt) sequence of its cDNA shows an open reading frame of 852 nt, which encodes a 32-kDa protein with two putative zinc fingers, and a serine/glutamine/proline-rich region. These features indicate a functional role for g1, which remains to be elucidated, in regulating gene expression during mesoderm formation. PMID:8462875

  1. An Fgf-Shh signaling hierarchy regulates early specification of the zebrafish skull.

    PubMed

    McCarthy, Neil; Sidik, Alfire; Bertrand, Julien Y; Eberhart, Johann K

    2016-07-15

    The neurocranium generates most of the craniofacial skeleton and consists of prechordal and postchordal regions. Although development of the prechordal is well studied, little is known of the postchordal region. Here we characterize a signaling hierarchy necessary for postchordal neurocranial development involving Fibroblast growth factor (Fgf) signaling for early specification of mesodermally-derived progenitor cells. The expression of hyaluron synthetase 2 (has2) in the cephalic mesoderm requires Fgf signaling and Has2 function, in turn, is required for postchordal neurocranial development. While Hedgehog (Hh)-deficient embryos also lack a postchordal neurocranium, this appears primarily due to a later defect in chondrocyte differentiation. Inhibitor studies demonstrate that postchordal neurocranial development requires early Fgf and later Hh signaling. Collectively, our results provide a mechanistic understanding of early postchordal neurocranial development and demonstrate a hierarchy of signaling between Fgf and Hh in the development of this structure. PMID:27060628

  2. Pleiotropic patterning response to activation of Shh signaling in the limb Apical Ectodermal Ridge

    PubMed Central

    Wang, Chi-Kuang Leo; Tsugane, Mizuyo H.; Scranton, Victoria; Kosher, Robert A.; Pierro, Louis J.; Upholt, William B.; Dealy, Caroline N.

    2012-01-01

    Sonic hedgehog (Shh) signaling in the limb plays a central role in coordination of limb patterning and outgrowth. Shh expression in the limb is limited to the cells of the Zone of Polarizing Activity (ZPA), located in posterior limb bud mesoderm. Shh is not expressed by limb ectoderm or AER, but recent studies suggest a role for AER-Shh signaling in limb patterning. Here, we have examined the effects of activation of Shh signaling in the AER. We find that targeted expression of Shh in the AER activates constitutive Shh signaling throughout the AER and subjacent limb mesoderm, and causes a range of limb patterning defects with progressive severity from mild polydactyly, to polysyndactyly with proximal defects, to severe oligodactyly with phocomelia and partial limb ventralization. Our studies emphasize the importance of control of the timing, level and location of Shh pathway signaling for limb AP, PD and DV patterning. PMID:21465622

  3. A patient with Mullerian abnormalities, renal dysplasia, cervical spine fusion, cataracts and intellectual disability: MURCS-plus?

    PubMed

    Tan, Tiong Yang; Whitelaw, Charlotte; Savarirayan, Ravi

    2007-10-01

    We report a 15-year-old girl with features of the MURCS (Mullerian abnormalities, renal agenesis/ectopy and cervicothoracic somite dysplasia) association and birth defects not typically associated with MURCS. In addition to seizures and intellectual disability, she has cortical brain heterotopia, bilateral subclinical cataracts, submucous cleft palate and patent ductus arteriosus. We propose that this patient represents a more severe form of MURCS, or 'MURCS-plus', which may represent a defect of or insult to mesodermal morphogenesis. PMID:17786121

  4. Formation of the extracellular matrix during the epimorphic anterior regeneration of Owenia fusiformis: autoradiographical and in situ hybridization studies.

    PubMed

    Dupin, F; Coulon, J; Le Parco, Y; Fontes, M; Thouveny, Y

    1991-06-01

    During post-traumatic regeneration of the polychaete annelid Owenia fusiformis, the extracellular matrix (ECM) formation was studied by light and electron microscopy and by histoautoradiography after incorporation of tritiated proline as marker for collagenic proteins. Three days after amputation, a new basement membrane was reformed in the blastema between the ectoderm and the mesoderm. At the same time, the cytoskeleton and the anchoring structures (hemidesmosomes) were differentiated in the basal part of the ectodermal cells. Four days after amputation, collagen fibers appeared in the extracellular matrix newly reformed between the ectodermal and mesodermal layers. The existence of a proximo-distal gradient in the organization of the new extracellular matrix and the accumulation of molecules labeled by 3H-proline was shown. This accumulation started at the level of the injured segment of the stump. Differences in labeling intensity were seen in the regenerate. Within specific organogenetic zones, i.e. the epidermal gland analagen, the branchial buds and the stomodeal invagination, the labeling between the ectodermal and mesodermal layers was less intense than in other parts of the regenerate. In the mesodermal connective septa (dissepiments), located between consecutive segments, the labeling and the accumulation of extracellular material occurred later than the formation of the ectodermal basement membrane. In situ hybridization of a DNA molecular probe corresponding partially to the coding region of the collagen-like gene Ocg8, showed a spatio-temporal expression of this gene. Northern blot analysis showed a single transcript of 6.6 kb. Four days after amputation the accumulation of this transcript was exclusively localized at the level of the ectodermal layer during differentiation of the regenerate. The ectoderm was thus shown to play a dynamic role during the first stages of traumatic regeneration, although it did not seem to be directly involved in the

  5. Derivation of the human embryonic stem cell line RCe008-A (RC-4).

    PubMed

    De Sousa, P A; Tye, B; Bruce, K; Dand, P; Gardner, J; Downie, J M; Bateman, M; Courtney, A

    2016-05-01

    The human embryonic stem cell line RCe008-A (RC-4) was derived from a blastocyst voluntarily donated as unsuitable and surplus to fertility requirements following ethics committee approved informed consent under licence from the UK Human Fertilisation and Embryology Authority. The cell line shows normal pluripotency marker expression and differentiation to ectoderm and mesoderm in vitro. It has a mixed 46XX/45X female karyotype and microsatellite PCR identity and blood group typing data is available. PMID:27346193

  6. Male urethral sarcoma: a case report and literature review.

    PubMed

    Nogueira, Magno Almeida; Santos, Guilherme Campelo Lopes Dos; Lopes, Roberto Iglesias; Campos, Octavio Henrique Arcos; Dall'Oglio, Marcos Francisco; Sant'Anna, Alexandre Crippa

    2016-03-01

    Urethral tumors are rare and aggressive. They usually affect men (2:1) and occur more commonly in white (85% of cases). Soft tissue sarcomas are a heterogeneous group of tumors that arise from embryonic mesoderm. It represents 1% of all cases of urinary tract malignancies and rarely primary affect the ureter. We report a case of male urethral sarcoma. To date, only two similar cases have been published in literature. PMID:26398363

  7. ABCC5 is required for cAMP-mediated hindgut invagination in sea urchin embryos.

    PubMed

    Shipp, Lauren E; Hill, Rose Z; Moy, Gary W; Gökırmak, Tufan; Hamdoun, Amro

    2015-10-15

    ATP-binding cassette (ABC) transporters are evolutionarily conserved proteins that pump diverse substrates across membranes. Many are known to efflux signaling molecules and are extensively expressed during development. However, the role of transporters in moving extracellular signals that regulate embryogenesis is largely unexplored. Here, we show that a mesodermal ABCC (MRP) transporter is necessary for endodermal gut morphogenesis in sea urchin embryos. This transporter, Sp-ABCC5a (C5a), is expressed in pigment cells and their precursors, which are a subset of the non-skeletogenic mesoderm (NSM) cells. C5a expression depends on Delta/Notch signaling from skeletogenic mesoderm and is downstream of Gcm in the aboral NSM gene regulatory network. Long-term imaging of development reveals that C5a knockdown embryos gastrulate, but ∼90% develop a prolapse of the hindgut by the late prism stage (∼8 h after C5a protein expression normally peaks). Since C5a orthologs efflux cyclic nucleotides, and cAMP-dependent protein kinase (Sp-CAPK/PKA) is expressed in pigment cells, we examined whether C5a could be involved in gastrulation through cAMP transport. Consistent with this hypothesis, membrane-permeable pCPT-cAMP rescues the prolapse phenotype in C5a knockdown embryos, and causes archenteron hyper-invagination in control embryos. In addition, the cAMP-producing enzyme soluble adenylyl cyclase (sAC) is expressed in pigment cells, and its inhibition impairs gastrulation. Together, our data support a model in which C5a transports sAC-derived cAMP from pigment cells to control late invagination of the hindgut. Little is known about the ancestral functions of ABCC5/MRP5 transporters, and this study reveals a novel role for these proteins in mesoderm-endoderm signaling during embryogenesis. PMID:26395488

  8. Ancestral state reconstruction by comparative analysis of a GRN kernel operating in echinoderms.

    PubMed

    Erkenbrack, Eric M; Ako-Asare, Kayla; Miller, Emily; Tekelenburg, Saira; Thompson, Jeffrey R; Romano, Laura

    2016-01-01

    Diverse sampling of organisms across the five major classes in the phylum Echinodermata is beginning to reveal much about the structure and function of gene regulatory networks (GRNs) in development and evolution. Sea urchins are the most studied clade within this phylum, and recent work suggests there has been dramatic rewiring at the top of the skeletogenic GRN along the lineage leading to extant members of the euechinoid sea urchins. Such rewiring likely accounts for some of the observed developmental differences between the two major subclasses of sea urchins-cidaroids and euechinoids. To address effects of topmost rewiring on downstream GRN events, we cloned four downstream regulatory genes within the skeletogenic GRN and surveyed their spatiotemporal expression patterns in the cidaroid Eucidaris tribuloides. We performed phylogenetic analyses with homologs from other non-vertebrate deuterostomes and characterized their spatiotemporal expression by quantitative polymerase chain reaction (qPCR) and whole-mount in situ hybridization (WMISH). Our data suggest the erg-hex-tgif subcircuit, a putative GRN kernel, exhibits a mesoderm-specific expression pattern early in Eucidaris development that is directly downstream of the initial mesodermal GRN circuitry. Comparative analysis of the expression of this subcircuit in four echinoderm taxa allowed robust ancestral state reconstruction, supporting hypotheses that its ancestral function was to stabilize the mesodermal regulatory state and that it has been co-opted and deployed as a unit in mesodermal subdomains in distantly diverged echinoderms. Importantly, our study supports the notion that GRN kernels exhibit structural and functional modularity, locking down and stabilizing clade-specific, embryonic regulatory states. PMID:26781941

  9. BMP-mediated functional cooperation between Dlx5;Dlx6 and Msx1;Msx2 during mammalian limb development.

    PubMed

    Vieux-Rochas, Maxence; Bouhali, Kamal; Mantero, Stefano; Garaffo, Giulia; Provero, Paolo; Astigiano, Simonetta; Barbieri, Ottavia; Caratozzolo, Mariano F; Tullo, Apollonia; Guerrini, Luisa; Lallemand, Yvan; Robert, Benoît; Levi, Giovanni; Merlo, Giorgio R

    2013-01-01

    The Dlx and Msx homeodomain transcription factors play important roles in the control of limb development. The combined disruption of Msx1 and Msx2, as well as that of Dlx5 and Dlx6, lead to limb patterning defects with anomalies in digit number and shape. Msx1;Msx2 double mutants are characterized by the loss of derivatives of the anterior limb mesoderm which is not observed in either of the simple mutants. Dlx5;Dlx6 double mutants exhibit hindlimb ectrodactyly. While the morphogenetic action of Msx genes seems to involve the BMP molecules, the mode of action of Dlx genes still remains elusive. Here, examining the limb phenotypes of combined Dlx and Msx mutants we reveal a new Dlx-Msx regulatory loop directly involving BMPs. In Msx1;Dlx5;Dlx6 triple mutant mice (TKO), beside the expected ectrodactyly, we also observe the hallmark morphological anomalies of Msx1;Msx2 double mutants suggesting an epistatic role of Dlx5 and Dlx6 over Msx2. In Msx2;Dlx5;Dlx6 TKO mice we only observe an aggravation of the ectrodactyly defect without changes in the number of the individual components of the limb. Using a combination of qPCR, ChIP and bioinformatic analyses, we identify two Dlx/Msx regulatory pathways: 1) in the anterior limb mesoderm a non-cell autonomous Msx-Dlx regulatory loop involves BMP molecules through the AER and 2) in AER cells and, at later stages, in the limb mesoderm the regulation of Msx2 by Dlx5 and Dlx6 occurs also cell autonomously. These data bring new elements to decipher the complex AER-mesoderm dialogue that takes place during limb development and provide clues to understanding the etiology of congenital limb malformations. PMID:23382810

  10. Stem cells in asexual reproduction of Enchytraeus japonensis (Oligochaeta, Annelid): proliferation and migration of neoblasts.

    PubMed

    Sugio, Mutsumi; Yoshida-Noro, Chikako; Ozawa, Kaname; Tochinai, Shin

    2012-05-01

    Enchytraeus japonensis is a small oligochaete that reproduces mainly asexually by fragmentation (autotomy) and regeneration. As sexual reproduction can also be induced, it is a good animal model for the study of both somatic and germline stem cells. To clarify the features of stem cells in regeneration, we investigated the proliferation and lineage of stem cells in E. japonensis. Neoblasts, which have the morphological characteristics of undifferentiated cells, were found to firmly adhere to the posterior surface of septa in each trunk segment. Also, smaller neoblast-like cells, which are designated as N-cells in this study, were located dorsal to the neoblasts on the septa. By conducting 5-bromo-2'-deoxyuridine (BrdU)-labeling-experiments, we have shown that neoblasts are slow-cycling (or quiescent) in intact growing worms, but proliferate rapidly in response to fragmentation. N-cells proliferate more actively than do neoblasts in intact worms. The results of pulse-chase experiments indicated that neoblast and N-cell lineage mesodermal cells that incorporated BrdU early in regeneration migrated toward the autotomized site to form the mesodermal region of the blastema, while the epidermal and intestinal cells also contributed to the blastema locally near the autotomized site. We have also shown that neoblasts have stem cell characteristics by expressing Ej-vlg2 and by the activity of telomerase during regeneration. Telomerase activity was high in the early stage of regeneration and correlated with the proliferation activity in the neoblast lineage of mesodermal stem cells. Taken together, our results indicate that neoblasts are mesodermal stem cells involved in the regeneration of E. japonensis. PMID:22417296

  11. The spatial and temporal expression of Ch-en, the engrailed gene in the polychaete Chaetopterus, does not support a role in body axis segmentation

    NASA Technical Reports Server (NTRS)

    Seaver, E. C.; Paulson, D. A.; Irvine, S. Q.; Martindale, M. Q.

    2001-01-01

    We are interested in understanding whether the annelids and arthropods shared a common segmented ancestor and have approached this question by characterizing the expression pattern of the segment polarity gene engrailed (en) in a basal annelid, the polychaete Chaetopterus. We have isolated an en gene, Ch-en, from a Chaetopterus cDNA library. Genomic Southern blotting suggests that this is the only en class gene in this animal. The predicted protein sequence of the 1.2-kb cDNA clone contains all five domains characteristic of en proteins in other taxa, including the en class homeobox. Whole-mount in situ hybridization reveals that Ch-en is expressed throughout larval life in a complex spatial and temporal pattern. The Ch-en transcript is initially detected in a small number of neurons associated with the apical organ and in the posterior portion of the prototrochophore. At later stages, Ch-en is expressed in distinct patterns in the three segmented body regions (A, B, and C) of Chaetopterus. In all segments, Ch-en is expressed in a small set of segmentally iterated cells in the CNS. In the A region, Ch-en is also expressed in a small group of mesodermal cells at the base of the chaetal sacs. In the B region, Ch-en is initially expressed broadly in the mesoderm that then resolves into one band/segment coincident with morphological segmentation. The mesodermal expression in the B region is located in the anterior region of each segment, as defined by the position of ganglia in the ventral nerve cord, and is involved in the morphogenesis of segment-specific feeding structures late in larval life. We observe banded mesodermal and ectodermal staining in an anterior-posterior sequence in the C region. We do not observe a segment polarity pattern of expression of Ch-en in the ectoderm, as is observed in arthropods. Copyright 2001 Academic Press.

  12. Penile agenesis: a fatal variation of an uncommon lesion.

    PubMed

    Gilbert, J; Clark, R D; Koyle, M A

    1990-02-01

    Two male neonates (46XY karyotype) were born with a triad of penile agenesis associated with imperforate anus and complete absence of the median raphe. Both patients died of renal dysplasia and secondary pulmonary hypoplasia shortly after birth. This triad appears to be secondary to a lack of caudal mesoderm migration during month 1 of gestation, leading to severe developmental defects in the caudal axis. In all reported cases to date this triad of findings has been incompatible with extrauterine life. PMID:2299726

  13. Male urethral sarcoma: a case report and literature review

    PubMed Central

    Nogueira, Magno Almeida; dos Santos, Guilherme Campelo Lopes; Lopes, Roberto Iglesias; Campos, Octavio Henrique Arcos; Dall’Oglio, Marcos Francisco; Sant’Anna, Alexandre Crippa

    2016-01-01

    ABSTRACT Urethral tumors are rare and aggressive. They usually affect men (2:1) and occur more commonly in white (85% of cases). Soft tissue sarcomas are a heterogeneous group of tumors that arise from embryonic mesoderm. It represents 1% of all cases of urinary tract malignancies and rarely primary affect the ureter. We report a case of male urethral sarcoma. To date, only two similar cases have been published in literature. PMID:26398363

  14. Injury, nerve, and wound epidermis related electrophoretic and fluorographic protein patterns in forelimbs of adult newts

    SciTech Connect

    Garling, D.J.; Tassava, R.A.

    1984-08-01

    Polyacrylamide slab gel electrophoresis and (/sup 35/S)methionine fluorography were used to examine proteins in regenerating newt limbs, amputated denervated limbs, unamputated denervated limbs, and separated blastema mesodermal core and wound epidermis. A total of 27 protein electrophoretic bands were obtained from amputated limbs and 24 bands from unamputated limbs. Amputation resulted in the appearance of 4 new bands and the loss of 1 band as compared to unamputated limbs. These 5 banding differences were apparent on stained gels 3 days postamputation and were maintained through 10 weeks postamputation (complete regenerate stage). Only one band in unamputated limbs was always detectable on fluorographs, whereas virtually all of the stainable bands of amputated limbs were visible on fluorographs. Amputation clearly stimulated a marked, generalized increase in the synthesis of limb proteins. The 5 amputation induced changes were equally evident in stained gels of both innervated and denervated limbs. Amputated denervated limbs possessed a full set of fluorographic bands (including the 5 differences) through 18 days postamputation. However, denervation without amputation was not sufficient to alter the stainable banding pattern. Wound epidermis and mesodermal core both displayed the 5 banding differences and had identical banding patterns with the exception of one epidermal specific band. This band was also present in whole limb skin but was absent in unamputated mesodermal limb tissue. This was the only band of unamputated limbs that was consistently detectable by fluorography. It is concluded that amputation induces nerve independent changes in protein synthesis that are common to both mesodermal core and wound epidermis. These changes may represent preparation for cellular proliferation.

  15. FRL-1, a member of the EGF-CFC family, is essential for neural differentiation in Xenopus early development.

    PubMed

    Yabe, Shin-Ichiro; Tanegashima, Kousuke; Haramoto, Yoshikazu; Takahashi, Shuji; Fujii, Tomoyuki; Kozuma, Siro; Taketani, Yuji; Asashima, Makoto

    2003-05-01

    Recent studies indicate an essential role for the EGF-CFC family in vertebrate development, particularly in the regulation of nodal signaling. Biochemical evidence suggests that EGF-CFC genes can also activate certain cellular responses independently of nodal signaling. Here, we show that FRL-1, a Xenopus EGF-CFC gene, suppresses BMP signaling to regulate an early step in neural induction. Overexpression of FRL-1 in animal caps induced the early neural markers zic3, soxD and Xngnr-1, but not the pan-mesodermal marker Xbra or the dorsal mesodermal marker chordin. Furthermore, overexpression of FRL-1 suppressed the expression of the BMP-responsive genes, Xvent-1 and Xmsx-1, which are expressed in animal caps and induced by overexpressed BMP-4. Conversely, loss of function analysis using morpholino-antisense oligonucleotides against FRL-1 (FRL-1MO) showed that FRL-1 is required for neural development. FRL-1MO-injected embryos lacked neural structures but contained mesodermal tissue. It was suggested previously that expression of early neural genes that mark the start of neuralization is activated in the presumptive neuroectoderm of gastrulae. FRL-1MO also inhibited the expression of these genes in dorsal ectoderm, but did not affect the expression of chordin, which acts as a neural inducer from dorsal mesoderm. FRL-1MO also inhibited the expression of neural markers that were induced by chordin in animal caps, suggesting that FRL-1 enables the response to neural inducing signals in ectoderm. Furthermore, we showed that the activation of mitogen-activated protein kinase by FRL-1 is required for neural induction and BMP inhibition. Together, these results suggest that FRL-1 is essential in the establishment of the neural induction response. PMID:12668622

  16. Not just inductive: a crucial mechanical role for the endoderm during heart tube assembly

    PubMed Central

    Varner, Victor D.; Taber, Larry A.

    2012-01-01

    The heart is the first functioning organ to form during development. During gastrulation, the cardiac progenitors reside in the lateral plate mesoderm but maintain close contact with the underlying endoderm. In amniotes, these bilateral heart fields are initially organized as a pair of flat epithelia that move towards the embryonic midline and fuse above the anterior intestinal portal (AIP) to form the heart tube. This medial motion is typically attributed to active mesodermal migration over the underlying endoderm. In this model, the role of the endoderm is twofold: to serve as a mechanically passive substrate for the crawling mesoderm and to secrete various growth factors necessary for cardiac specification and differentiation. Here, using computational modeling and experiments on chick embryos, we present evidence supporting an active mechanical role for the endoderm during heart tube assembly. Label-tracking experiments suggest that active endodermal shortening around the AIP accounts for most of the heart field motion towards the midline. Results indicate that this shortening is driven by cytoskeletal contraction, as exposure to the myosin-II inhibitor blebbistatin arrested any shortening and also decreased both tissue stiffness (measured by microindentation) and mechanical tension (measured by cutting experiments). In addition, blebbistatin treatment often resulted in cardia bifida and abnormal foregut morphogenesis. Moreover, finite element simulations of our cutting experiments suggest that the endoderm (not the mesoderm) is the primary contractile tissue layer during this process. Taken together, these results indicate that contraction of the endoderm actively pulls the heart fields towards the embryonic midline, where they fuse to form the heart tube. PMID:22492358

  17. The spatial and temporal dynamics of Sax1 (CHox3) homeobox gene expression in the chick's spinal cord.

    PubMed

    Spann, P; Ginsburg, M; Rangini, Z; Fainsod, A; Eyal-Giladi, H; Gruenbaum, Y

    1994-07-01

    Sax1 (previously CHox3) is a chicken homeobox gene belonging to the same homeobox gene family as the Drosophila NK1 and the honeybee HHO genes. Sax1 transcripts are present from stage 2 H&H until at least 5 days of embryonic development. However, specific localization of Sax1 transcripts could not be detected by in situ hybridization prior to stage 8-, when Sax1 transcripts are specifically localized in the neural plate, posterior to the hindbrain. From stages 8- to 15 H&H, Sax1 continues to be expressed only in the spinal part of the neural plate. The anterior border of Sax1 expression was found to be always in the transverse plane separating the youngest somite from the yet unsegmented mesodermal plate and to regress with similar dynamics to that of the segregation of the somites from the mesodermal plate. The posterior border of Sax1 expression coincides with the posterior end of the neural plate. In order to study a possible regulation of Sax1 expression by its neighboring tissues, several embryonic manipulation experiments were performed. These manipulations included: removal of somites, mesodermal plate or notochord and transplantation of a young ectopic notochord in the vicinity of the neural plate or transplantation of neural plate sections into the extraembryonic area. The results of these experiments revealed that the induction of the neural plate by the mesoderm has already occurred in full primitive streak embryos, after which Sax1 is autonomously regulated within the spinal part of the neural plate. PMID:7924989

  18. GATA2 regulates Wnt signaling to promote primitive red blood cell fate.

    PubMed

    Mimoto, Mizuho S; Kwon, Sunjong; Green, Yangsook Song; Goldman, Devorah; Christian, Jan L

    2015-11-01

    Primitive erythropoiesis is regulated in a non cell-autonomous fashion across evolution from frogs to mammals. In Xenopus laevis, signals from the overlying ectoderm are required to induce the mesoderm to adopt an erythroid fate. Previous studies in our lab identified the transcription factor GATA2 as a key regulator of this ectodermal signal. To identify GATA2 target genes in the ectoderm required for red blood cell formation in the mesoderm, we used microarray analysis to compare gene expression in ectoderm from GATA2 depleted and wild type embryos. Our analysis identified components of the non-canonical and canonical Wnt pathways as being reciprocally up- and down-regulated downstream of GATA2 in both mesoderm and ectoderm. We show that up-regulation of canonical Wnt signaling during gastrulation blocks commitment to a hematopoietic fate while down-regulation of non-canonical Wnt signaling impairs erythroid differentiation. Our results are consistent with a model in which GATA2 contributes to inhibition of canonical Wnt signaling, thereby permitting progenitors to exit the cell cycle and commit to a hematopoietic fate. Subsequently, activation of non-canonical Wnt signaling plays a later role in enabling these progenitors to differentiate as mature red blood cells. PMID:26365900

  19. Analysis of coelom development in the sea urchin Holopneustes purpurescens yielding a deuterostome body plan.

    PubMed

    Morris, Valerie B

    2016-01-01

    An analysis of early coelom development in the echinoid Holopneustes purpurescens yields a deuterostome body plan that explains the disparity between the pentameral plan of echinoderms and the bilateral plans of chordates and hemichordates, the three major phyla of the monophyletic deuterostomes. The analysis shows an early separation into a medial hydrocoele and lateral coelomic mesoderm with an enteric channel between them before the hydrocoele forms the pentameral plan of five primary podia. The deuterostome body plan thus has a single axial or medial coelom and a pair of lateral coeloms, all surrounding an enteric channel, the gut channel. Applied to the phyla, the medial coelom is the hydrocoele in echinoderms, the notochord in chordates and the proboscis coelom in hemichordates: the lateral coeloms are the coelomic mesoderm in echinoderms, the paraxial mesoderm in chordates and the lateral coeloms in hemichordates. The plan fits frog and chick development and the echinoderm fossil record, and predicts genes involved in coelomogenesis as the source of deuterostome macroevolution. PMID:26892238

  20. Spatiotemporal transcriptomics reveals the evolutionary history of the endoderm germ layer.

    PubMed

    Hashimshony, Tamar; Feder, Martin; Levin, Michal; Hall, Brian K; Yanai, Itai

    2015-03-12

    The concept of germ layers has been one of the foremost organizing principles in developmental biology, classification, systematics and evolution for 150 years (refs 1 - 3). Of the three germ layers, the mesoderm is found in bilaterian animals but is absent in species in the phyla Cnidaria and Ctenophora, which has been taken as evidence that the mesoderm was the final germ layer to evolve. The origin of the ectoderm and endoderm germ layers, however, remains unclear, with models supporting the antecedence of each as well as a simultaneous origin. Here we determine the temporal and spatial components of gene expression spanning embryonic development for all Caenorhabditis elegans genes and use it to determine the evolutionary ages of the germ layers. The gene expression program of the mesoderm is induced after those of the ectoderm and endoderm, thus making it the last germ layer both to evolve and to develop. Strikingly, the C. elegans endoderm and ectoderm expression programs do not co-induce; rather the endoderm activates earlier, and this is also observed in the expression of endoderm orthologues during the embryology of the frog Xenopus tropicalis, the sea anemone Nematostella vectensis and the sponge Amphimedon queenslandica. Querying the phylogenetic ages of specifically expressed genes reveals that the endoderm comprises older genes. Taken together, we propose that the endoderm program dates back to the origin of multicellularity, whereas the ectoderm originated as a secondary germ layer freed from ancestral feeding functions. PMID:25487147

  1. The segmented urbilateria: a testable scenario.

    PubMed

    Balavoine, Guillaume; Adoutte, André

    2003-02-01

    The idea that the last common ancestor of bilaterian animals (Urbilateria) was segmented has been raised recently on evidence coming from comparative molecular embryology. Leaving aside the complex debate on the value of genetic evidence, the morphological and developmental evidence in favor of a segmented Urbilateria are discussed in the light of the emerging molecular phylogeny of metazoans. Applying a cladistic character optimization procedure to the question of segmentation is vastly complicated by the problem of defining without ambiguity what segmentation is and to what taxa this definition applies. An ancestral segmentation might have undergone many complex derivations in each different phylum, thus rendering the cladistics approaches problematic. Taking the most general definitions of coelom and segmentation however, some remarkably similar patterns are found across the bilaterian tree in the way segments are formed by the posterior addition of mesodermal segments or somites. Postulating that these striking similarities in mesodermal patterns are ancestral, a scenario for the diversification of bilaterians from a metameric ancestor is presented. Several types of evolutionary mechanisms (specialization, tagmosis, progenesis) operating on a segmented ancestral body plan would explain the rapid emergence of body plans during the Cambrian. We finally propose to test this hypothesis by comparing genes involved in mesodermal segmentation. PMID:21680418

  2. Development, triploblastism, physics of wetting and the Cambrian explosion.

    PubMed

    Fleury, Vincent

    2013-09-01

    The Cambrian explosion is characterized by the sudden outburst of organized animal plans, which occurred circa 530 M years ago. Around that time, many forms of animal life appeared, including several which have since disappeared. There is no general consensus about "why" this happened, and why it had any form of suddenness. However, all organized animal plans share a common feature: they are triploblastic, i.e., composed of 3 layers of tissue, endoderm, ectoderm and mesoderm. I show here that, within simple hypotheses, the formation of the mesoderm has intrinsically a physical exponential dynamics, leading rapidly to triploblastism, and eventually, to animal formation. A novel physico-mathematical framework including epithelium-mesenchyme transition, visco-elastic constitutive equations, and conservation laws, is presented which allows one to describe gastrulation as a self-wetting phenomenon of a soft solid onto itself. This phenomenon couples differentiation and migration during gastrulation, and leads in a closed form to an exponential scaling law for the formation of the mesoderm. Therefore, the Cambrian explosion might have started, actually, by a true viscoelastic "explosion": the exponential run-away of mesenchymal cells. PMID:23959076

  3. Distinct regulation of the anterior and posterior myeloperoxidase expression by Etv2 and Gata1 during primitive Granulopoiesis in zebrafish.

    PubMed

    Glenn, Nicole O; Schumacher, Jennifer A; Kim, Hyon J; Zhao, Emma J; Skerniskyte, Jurate; Sumanas, Saulius

    2014-09-01

    Neutrophilic granulocytes are the most abundant type of myeloid cells and form an essential part of the innate immune system. In vertebrates the first neutrophils are thought to originate during primitive hematopoiesis, which precedes hematopoietic stem cell formation. In zebrafish embryos, it has been suggested that primitive neutrophils may originate in two distinct sites, the anterior (ALPM) and posterior lateral plate mesoderm (PLPM). An ETS-family transcription factor Etsrp/Etv2/ER71 has been implicated in vasculogenesis and hematopoiesis in multiple vertebrates. However, its role during neutrophil development is not well understood. Here we demonstrate using zebrafish embryos that Etv2 has a specific cell-autonomous function during primitive neutropoiesis in the anterior lateral plate mesoderm (ALPM) but has little effect on erythropoiesis or the posterior lateral plate mesoderm (PLPM) expression of neutrophil marker myeloperoxidase mpo/mpx. Our results argue that ALPM-derived neutrophils originate from etv2-expressing cells which downregulate etv2 during neutropoiesis. We further show that Scl functions downstream of Etv2 in anterior neutropoiesis. Additionally, we demonstrate that mpx expression within the PLPM overlaps with gata1 expression, potentially marking the cells with a dual myelo-erythroid potential. Intriguingly, initiation of mpx expression in the PLPM is dependent on gata1 but not etv2 function. Our results demonstrate that mpx expression is controlled differently in the ALPM and PLPM regions and describe novel roles for etv2 and gata1 during primitive neutropoiesis. PMID:24956419

  4. Evolutionary origins of the vertebrate heart: Specification of the cardiac lineage in Ciona intestinalis

    PubMed Central

    Davidson, Brad; Levine, Michael

    2003-01-01

    Here we exploit the extensive cell lineage information and streamlined genome of the ascidian, Ciona intestinalis, to investigate heart development in a basal chordate. Several cardiac genes were analyzed, including the sole Ciona ortholog of the Drosophila tinman gene, and tissue-specific enhancers were isolated for some of the genes. Conserved sequence motifs within these enhancers facilitated the isolation of a heart enhancer for the Ciona Hand-like gene. Altogether, these studies provide a regulatory framework for the differentiation of the cardiac mesoderm, beginning at the 110-cell stage, and extending through the fusion of cardiac progenitors during tail elongation. The cardiac lineage shares a common origin with the germ line, and zygotic transcription is first detected in the heart progenitors only after its separation from the germ line at the 64-cell stage. We propose that germ-line determinants influence the specification of the cardiac mesoderm, both by inhibiting inductive signals required for the development of noncardiac mesoderm lineages, and by providing a localized source of Wnt-5 and other signals required for heart development. We discuss the possibility that the germ line also influences the specification of the vertebrate heart. PMID:14500781

  5. Myoblasts and myoblast-conditioned medium attract the earliest spinal neurites from frog embryos.

    PubMed Central

    McCaig, C D

    1986-01-01

    A study was made of the capacity of newly segmented somites, unsegmented mesoderm and medium conditioned by each of these tissues to attract the growth of the earliest spinal neurites from the neural tube of Xenopus laevis in tissue culture. When presented with segmented somitic myoblasts or sheets of skin, spinal neurites grew selectively towards the somitic myoblasts. Neurites were not attracted specifically to somitic myoblasts from their own rostrocaudal level. A variable proportion of myoblasts from unsegmented caudal mesoderm differentiated and elongated in co-culture with neural tube and skin. These myoblasts also attracted neural outgrowths, but only if present in sufficient numbers. An agar slab containing medium conditioned by the presence of segmented myoblasts for 1 day attracted neurite outgrowths. A source of medium conditioned by the presence of undifferentiated, unsegmented myotomal mesoderm alone did not attract neurite outgrowths. Nerve growth factor (NGF) at a range of concentrations in the agar source (500-10,000 ng/ml) did not attract the earliest neurite outgrowths. It is concluded that the earliest skeletal myoblasts from Xenopus laevis embryos may attract neural outgrowths by releasing a soluble factor. Myoblasts may have to develop to the stage of somite segmentation before secretion of such an agent begins. The release of a myoblast-derived factor so early in development may assist directed nerve growth in vivo. Images Plate 1 Plate 2 PMID:3795063

  6. Analysis of coelom development in the sea urchin Holopneustes purpurescens yielding a deuterostome body plan

    PubMed Central

    Morris, Valerie B.

    2016-01-01

    ABSTRACT An analysis of early coelom development in the echinoid Holopneustes purpurescens yields a deuterostome body plan that explains the disparity between the pentameral plan of echinoderms and the bilateral plans of chordates and hemichordates, the three major phyla of the monophyletic deuterostomes. The analysis shows an early separation into a medial hydrocoele and lateral coelomic mesoderm with an enteric channel between them before the hydrocoele forms the pentameral plan of five primary podia. The deuterostome body plan thus has a single axial or medial coelom and a pair of lateral coeloms, all surrounding an enteric channel, the gut channel. Applied to the phyla, the medial coelom is the hydrocoele in echinoderms, the notochord in chordates and the proboscis coelom in hemichordates: the lateral coeloms are the coelomic mesoderm in echinoderms, the paraxial mesoderm in chordates and the lateral coeloms in hemichordates. The plan fits frog and chick development and the echinoderm fossil record, and predicts genes involved in coelomogenesis as the source of deuterostome macroevolution. PMID:26892238

  7. The transcription factor Vox represses endoderm development by interacting with Casanova and Pou2.

    PubMed

    Zhao, Jue; Lambert, Guillaume; Meijer, Annemarie H; Rosa, Frederic M

    2013-03-01

    Endoderm and mesoderm are both formed upon activation of Nodal signaling but how endoderm differentiates from mesoderm is still poorly explored. The sox-related gene casanova (sox32) acts downstream of the Nodal signal, is essential for endoderm development and requires the co-factor Pou2 (Pou5f1, Oct3, Oct4) in this process. Conversely, BMP signals have been shown to inhibit endoderm development by an as yet unexplained mechanism. In a search for Casanova regulators in zebrafish, we identified two of its binding partners as the transcription factors Pou2 and Vox, a member of the Vent group of proteins also involved in the patterning of the gastrula. In overexpression studies we show that vox and/or Vent group genes inhibit the capacity of Casanova to induce endoderm, even in the presence of its co-factor Pou2, and that Vox acts as a repressor in this process. We further show that vox, but not other members of the Vent group, is essential for defining the proper endodermal domain size at gastrulation. In this process, vox acts downstream of BMPs. Cell fate analysis further shows that Vox plays a key role downstream of BMP signals in regulating the capacity of Nodal to induce endoderm versus mesoderm by modulating the activity of the Casanova/Pou2 regulatory system. PMID:23364327

  8. Eya 1 acts as a critical regulator for specifying the metanephric mesenchyme.

    PubMed

    Sajithlal, Gangadharan; Zou, Dan; Silvius, Derek; Xu, Pin-Xian

    2005-08-15

    Although it is well established that the Gdnf-Ret signal transduction pathway initiates metanephric induction, no single regulator has yet been identified to specify the metanephric mesenchyme or blastema within the intermediate mesoderm, the earliest step of metanephric kidney development and the molecular mechanisms controlling Gdnf expression are essentially unknown. Previous studies have shown that a loss of Eya 1 function leads to renal agenesis that is a likely result of failure of metanephric induction. The studies presented here demonstrate that Eya 1 specifies the metanephric blastema within the intermediate mesoderm at the caudal end of the nephrogenic cord. In contrast to its specific roles in metanephric development, Eya 1 appears dispensable for the formation of nephric duct and mesonephric tubules. Using a combination of null and hypomorphic Eya 1 mutants, we now demonstrated that approximately 20% of normal Eya 1 protein level is sufficient for establishing the metanephric blastema and inducing the ureteric bud formation but not for its normal branching. Using Eya 1, Gdnf, Six 1 and Pax 2 mutant mice, we show that Eya 1 probably functions at the top of the genetic hierarchy controlling kidney organogenesis and it acts in combination with Six 1 and Pax 2 to regulate Gdnf expression during UB outgrowth and branching. These findings uncover an essential function for Eya 1 as a critical determination factor in acquiring metanephric fate within the intermediate mesoderm and as a key regulator of Gdnf expression during ureteric induction and branching morphogenesis. PMID:16018995

  9. Pax3 and Tbx5 specify whether PDGFRα+ cells assume skeletal or cardiac muscle fate in differentiating ES cells

    PubMed Central

    Magli, Alessandro; Schnettler, Erin; Swanson, Scott A; Borges, Luciene; Hoffman, Kirsta; Stewart, Ron; Thomson, James A; Keirstead, Susan A.; Perlingeiro, Rita C. R.

    2014-01-01

    Embryonic stem (ES) cells represent an ideal model to study how lineage decisions are established during embryonic development. Using a doxycycline-inducible mouse ES cell line, we have previously shown that expression of the transcriptional activator Pax3 in early mesodermal cells leads to the robust generation of paraxial mesoderm progenitors that ultimately differentiate into skeletal muscle precursors. Here we show that the ability of this transcription factor to induce the skeletal myogenic cell fate occurs at the expenses of the cardiac lineage. Our results show that the PDGFRα+FLK1− sub-fraction represents the main population affected by Pax3, through down-regulation of several transcripts encoding for proteins involved in cardiac development. We demonstrate that although Nkx2-5, Tbx5 and Gata4 negatively affect Pax3 skeletal myogenic activity, the cardiac potential of embryoid body (EB)-derived cultures is restored solely by forced expression of Tbx5. Taking advantage of this model, we employed an unbiased genome wide approach to identify genes whose expression is rescued by Tbx5, and which could represent important regulators of cardiac development. These findings elucidate mechanisms regulating the commitment of mesodermal cells in the early embryo and identify the Tbx5 cardiac transcriptome. PMID:24677751

  10. Robustness and Accuracy in Sea Urchin Developmental Gene Regulatory Networks

    PubMed Central

    Ben-Tabou de-Leon, Smadar

    2016-01-01

    Developmental gene regulatory networks robustly control the timely activation of regulatory and differentiation genes. The structure of these networks underlies their capacity to buffer intrinsic and extrinsic noise and maintain embryonic morphology. Here I illustrate how the use of specific architectures by the sea urchin developmental regulatory networks enables the robust control of cell fate decisions. The Wnt-βcatenin signaling pathway patterns the primary embryonic axis while the BMP signaling pathway patterns the secondary embryonic axis in the sea urchin embryo and across bilateria. Interestingly, in the sea urchin in both cases, the signaling pathway that defines the axis controls directly the expression of a set of downstream regulatory genes. I propose that this direct activation of a set of regulatory genes enables a uniform regulatory response and a clear cut cell fate decision in the endoderm and in the dorsal ectoderm. The specification of the mesodermal pigment cell lineage is activated by Delta signaling that initiates a triple positive feedback loop that locks down the pigment specification state. I propose that the use of compound positive feedback circuitry provides the endodermal cells enough time to turn off mesodermal genes and ensures correct mesoderm vs. endoderm fate decision. Thus, I argue that understanding the control properties of repeatedly used regulatory architectures illuminates their role in embryogenesis and provides possible explanations to their resistance to evolutionary change. PMID:26913048

  11. The type I activin receptor ActRIB is required for egg cylinder organization and gastrulation in the mouse

    PubMed Central

    Gu, Zhenyu; Nomura, Masatoshi; Simpson, Brenda B.; Lei, Hong; Feijen, Alie; van den Eijnden-van Raaij, Janny; Donahoe, Patricia K.; Li, En

    1998-01-01

    ActRIB is a type I transmembrane serine/threonine kinase receptor that has been shown to form heteromeric complexes with the type II activin receptors to mediate activin signal. To investigate the function of ActRIB in mammalian development, we generated ActRIB-deficient ES cell lines and mice by gene targeting. Analysis of the ActRIB−/− embryos showed that the epiblast and the extraembryonic ectoderm were disorganized, resulting in disruption and developmental arrest of the egg cylinder before gastrulation. To assess the function of ActRIB in mesoderm formation and gastrulation, chimera analysis was conducted. We found that ActRIB−/− ES cells injected into wild-type blastocysts were able to contribute to the mesoderm in chimeric embryos, suggesting that ActRIB is not required for mesoderm formation. Primitive streak formation, however, was impaired in chimeras when ActRIB−/− cells contributed highly to the epiblast. Further, chimeras generated by injection of wild-type ES cells into ActRIB−/− blastocysts formed relatively normal extraembryonic tissues, but the embryo proper developed poorly probably resulting from severe gastrulation defect. These results provide genetic evidence that ActRIB functions in both epiblast and extraembryonic cells to mediate signals that are required for egg cylinder organization and gastrulation. PMID:9512518

  12. Human pluripotent embryonal carcinoma NTERA2 cl.D1 cells maintain their typical morphology in an angiomyogenic medium

    PubMed Central

    Simões, Pedro D; Ramos, Teresa

    2007-01-01

    Background Pluripotent embryonal carcinomas are good potential models, to study, "in vitro," the mechanisms that control differentiation during embryogenesis. The NTERA2cl.D1 (NT2/D1) cell line is a well known system of ectodermal differentiation. Retinoic acid (RA) induces a dorsal pattern of differentiation (essentially neurons) and bone morphogenetic protein (BMP) or hexamethylenebisacetamide (HMBA) induces a more ventral (epidermal) pattern of differentiation. However, whether these human cells could give rise to mesoderm derivatives as their counterpart in mouse remained elusive. We analyzed the morphological characteristics and transcriptional activation of genes pertinent in cardiac muscle and endothelium differentiation, during the growth of NT2/D1 cells in an inductive angiomyogenic medium with or without Bone Morphogenetic Protein 2 (BMP2). Results Our experiments showed that NT2/D1 maintains their typical actin organization in angiomyogenic medium. Although the beta myosin heavy chain gene was never detected, all the other 15 genes analyzed maintained their expression throughout the time course of the experiment. Among them were early and late cardiac, endothelial, neuronal and teratocarcinoma genes. Conclusion Our results suggest that despite the NT2/D1 cells natural tendency to differentiate into neuroectodermal lineages, they can activate genes of mesodermal lineages. Therefore, we believe that these pluripotent cells might still be a good model to study biological development of mesodermal derivatives, provided the right culture conditions are met. PMID:17442106

  13. ENDODERMAL ORIGIN OF BLADDER TRIGONE INFERRED FROM MESENCHYMAL-EPITHELIAL INTERACTIONS

    PubMed Central

    Tanaka, Stacy T.; Ishii, Kenichiro; DeMarco, Romano T.; Pope, John C.; Brock, John W.; Hayward, Simon W.

    2009-01-01

    Purpose In the classic view of bladder development, the trigone originates from the mesoderm-derived Wolffian ducts while the remainder of the bladder originates from the endoderm-derived urogenital sinus. Recent molecular developmental studies have questioned the veracity of this received wisdom, suggesting an endodermal origin for the trigone. To shed further light on this issue we observed mesenchymalepithelial interactions between trigone epithelium and fetal urogenital sinus mesenchyme (UGM) to infer the germ layer of origin of the trigone. Materials and methods Mouse trigone epithelium was recombined with fetal rat UGM in tissue recombinant grafts that were placed beneath the renal capsule of athymic mouse hosts. Grafts were harvested at 4 weeks. Control grafts with bladder dome and ureteral epithelium were also examined. Tissues were evaluated with hematoxylin / eosin and Hoechst dye 33258 (to confirm cell species origin). Immunohistochemistry to androgen receptor, broad spectrum uroplakin, dorsal lateral prostate secretions and seminal vesicle secretions were used to differentiate prostatic and seminal vesicle differentiation. Results Grafts of mouse trigone epithelium with fetal rat UGM yielded epithelial tissue which stained for dorsal lateral prostate secretions but not for seminal vesicle secretions. Control grafts of bladder dome epithelium yielded the expected endodermal prostate differentiation. Control grafts of ureteral epithelium yielded the expected mesodermal seminal vesicle differentiation. Conclusions The consistent finding of prostatic epithelium in tissue recombinants of trigone epithelium and fetal UGM reinforces the hypothesis that the trigone is derived from the endoderm and not the mesoderm as commonly accepted. PMID:19914648

  14. Spatiotemporal transcriptomics reveals the evolutionary history of the endoderm germ layer

    PubMed Central

    Hashimshony, Tamar; Feder, Martin; Levin, Michal; Hall, Brian K.; Yanai, Itai

    2014-01-01

    The germ layer concept has been one of the foremost organizing principles in developmental biology, classification, systematics and evolution for 150 years1-3. Of the three germ layers, the mesoderm is found in bilaterian animals but is absent in species in the phyla Cnidaria and Ctenophora, which has been taken as evidence that the mesoderm was the final germ layer to evolve1,4,5. The origin of the ectoderm and endoderm germ layers, however, remains unclear with models supporting the antecedence of each as well as a simultaneous origin4,6-9. Here, we determine the temporal and spatial components of gene expression spanning embryonic development for all Caenorhabditis elegans genes and use it to determine the evolutionary ages of the germ layers. The gene expression program of the mesoderm is induced after those of the ectoderm and endoderm, thus making it the last germ layer to both evolve and develop. Strikingly, the C. elegans endoderm and ectoderm expression programs do not co-induce; rather the endoderm activates earlier, and this is observed also in the expression of endoderm orthologs during the embryology of Xenopus tropicalis, Nematostella vectensis, and the sponge Amphimedon queenslandica. Querying for the phylogenetic ages of specifically expressed genes revealed that the endoderm is comprised of older genes. Taken together, we propose that the endoderm program dates back to the origin of multicellularity, while the ectoderm originated as a secondary germ layer freed from ancestral feeding functions. PMID:25487147

  15. Endoderm differentiation and inductive effect of activin-treated ectoderm in Xenopus.

    PubMed

    Ninomiya, H; Takahashi, S; Tanegashima, K; Yokota, C; Asashima, M

    1999-08-01

    When presumptive ectoderm is treated with high concentrations of activin A, it mainly differentiates into axial mesoderm (notochord, muscle) in Xenopus and into yolk-rich endodermal cells in newt (Cynops pyrrhogaster). Xenopus ectoderm consists of multiple layers, different from the single layer of Cynops ectoderm. This multilayer structure of Xenopus ectoderm may prevent complete treatment of activin A and subsequent whole differentiation into endoderm. In the present study, therefore, Xenopus ectoderm was separated into an outer layer and an inner layer, which were individually treated with a high concentration of activin A (100 ng/mL). Then the differentiation and inductive activity of these ectodermal cells were examined in explantation and transplantation experiments. In isolation culture, ectoderm treated with activin A formed endoderm. Ectodermal and mesodermal tissues were seldom found in these explants. The activin-treated ectoderm induced axial mesoderm and neural tissues, and differentiated into endoderm when it was sandwiched between two sheets of ectoderm or was transplanted into the ventral marginal zone of other blastulae. These findings suggest that Xenopus ectoderm treated with a high concentration of activin A forms endoderm and mimics the properties of the organizer as in Cynops. PMID:10466926

  16. Evolution and development of the vertebrate neck

    PubMed Central

    Ericsson, Rolf; Knight, Robert; Johanson, Zerina

    2013-01-01

    Muscles of the vertebrate neck include the cucullaris and hypobranchials. Although a functional neck first evolved in the lobe-finned fishes (Sarcopterygii) with the separation of the pectoral/shoulder girdle from the skull, the neck muscles themselves have a much earlier origin among the vertebrates. For example, lampreys possess hypobranchial muscles, and may also possess the cucullaris. Recent research in chick has established that these two muscles groups have different origins, the hypobranchial muscles having a somitic origin but the cucullaris muscle deriving from anterior lateral plate mesoderm associated with somites 1–3. Additionally, the cucullaris utilizes genetic pathways more similar to the head than the trunk musculature. Although the latter results are from experiments in the chick, cucullaris homologues occur in a variety of more basal vertebrates such as the sharks and zebrafish. Data are urgently needed from these taxa to determine whether the cucullaris in these groups also derives from lateral plate mesoderm or from the anterior somites, and whether the former or the latter represent the basal vertebrate condition. Other lateral plate mesoderm derivatives include the appendicular skeleton (fins, limbs and supporting girdles). If the cucullaris is a definitive lateral plate-derived structure it may have evolved in conjunction with the shoulder/limb skeleton in vertebrates and thereby provided a greater degree of flexibility to the heads of predatory vertebrates. PMID:22697305

  17. The Hydra FGFR, Kringelchen, partially replaces the Drosophila Heartless FGFR.

    PubMed

    Rudolf, Anja; Hübinger, Christine; Hüsken, Katrin; Vogt, Angelika; Rebscher, Nicole; Onel, Susanne-Filiz; Renkawitz-Pohl, Renate; Hassel, Monika

    2013-05-01

    Fibroblast growth factor receptors (FGFR) are highly conserved receptor tyrosine kinases, and evolved early in metazoan evolution. In order to investigate their functional conservation, we asked whether the Kringelchen FGFR in the freshwater polyp Hydra vulgaris, is able to functionally replace FGFR in fly embryos. In Drosophila, two endogenous FGFR, Breathless (Btl) and Heartless (Htl), ensure formation of the tracheal system and mesodermal cell migration as well as formation of the heart. Using UAS-kringelchen-5xmyc transgenic flies and targeted expression, we show that Kringelchen is integrated correctly into the cell membrane of mesodermal and tracheal cells in Drosophila. Nevertheless, Kringelchen expression driven in tracheal cells failed to rescue the btl (LG19) mutant. The Hydra FGFR was able to substitute for Heartless in the htl (AB42) null mutant; however, this occurred only during early mesodermal cell migration. Our data provide evidence for functional conservation of this early-diverged FGFR across these distantly related phyla, but also selectivity for the Htl FGFR in the Drosophila system. PMID:23111653

  18. Twist1 Controls a Cell-Specification Switch Governing Cell Fate Decisions within the Cardiac Neural Crest

    PubMed Central

    Vincentz, Joshua W.; Firulli, Beth A.; Lin, Andrea; Spicer, Douglas B.; Howard, Marthe J.; Firulli, Anthony B.

    2013-01-01

    Neural crest cells are multipotent progenitor cells that can generate both ectodermal cell types, such as neurons, and mesodermal cell types, such as smooth muscle. The mechanisms controlling this cell fate choice are not known. The basic Helix-loop-Helix (bHLH) transcription factor Twist1 is expressed throughout the migratory and post-migratory cardiac neural crest. Twist1 ablation or mutation of the Twist-box causes differentiation of ectopic neuronal cells, which molecularly resemble sympathetic ganglia, in the cardiac outflow tract. Twist1 interacts with the pro-neural factor Sox10 via its Twist-box domain and binds to the Phox2b promoter to repress transcriptional activity. Mesodermal cardiac neural crest trans-differentiation into ectodermal sympathetic ganglia-like neurons is dependent upon Phox2b function. Ectopic Twist1 expression in neural crest precursors disrupts sympathetic neurogenesis. These data demonstrate that Twist1 functions in post-migratory neural crest cells to repress pro-neural factors and thereby regulate cell fate determination between ectodermal and mesodermal lineages. PMID:23555309

  19. Tailbud-derived Bmp4 drives proliferation and inhibits maturation of zebrafish chordamesoderm.

    PubMed

    Esterberg, Robert; Delalande, Jean-Marie; Fritz, Andreas

    2008-12-01

    In zebrafish, BMP signaling establishes cell identity along the dorsoventral (DV) axis during gastrulation. Owing to the early requirements of BMP activity in DV patterning, it has been difficult to assign later roles in cell fate specification to specific BMP ligands. In this study, we have taken advantage of two follistatin-like genes (fstl1 and fstl2), as well as a transgenic zebrafish line carrying an inducible truncated form of the BMP-type 1 receptor to study the role of Bmp4 outside of the context of DV specification. Characterization of fstl1/2 suggests that they exert a redundant role as BMP antagonists during late gastrulation, regulating BMP activity in axial mesoderm. Maintenance of appropriate levels of BMP signaling is crucial for the proper development of chordamesoderm, a subset of axial mesoderm that gives rise to the notochord, but not prechordal mesoderm, which gives rise to the prechordal plate. Bmp4 activity in particular is required during a crucial window beginning at late gastrulation and lasting through early somitogenesis to promote chordamesoderm proliferation. In the absence of Bmp4, the notochord precursor pool is depleted, and the notochord differentiates prematurely. Our results illustrate a role for Bmp4 in the proliferation and timely differentiation of axial tissue after DV axis specification. PMID:18948415

  20. Stepwise renal lineage differentiation of mouse embryonic stem cells tracing in vivo development

    SciTech Connect

    Nishikawa, Masaki; Yanagawa, Naomi; Kojima, Nobuhiko; Yuri, Shunsuke; Hauser, Peter V.; Jo, Oak D.; Yanagawa, Norimoto

    2012-01-13

    Highlights: Black-Right-Pointing-Pointer We induced renal lineages from mESCs by following the in vivo developmental cues. Black-Right-Pointing-Pointer We induced nephrogenic intermediate mesoderm by stepwise addition of factors. Black-Right-Pointing-Pointer We induced two types of renal progenitor cells by reciprocal conditioned media. Black-Right-Pointing-Pointer We propose the potential role of CD24 for the enrichment of renal lineage cells. -- Abstract: The in vitro derivation of renal lineage progenitor cells is essential for renal cell therapy and regeneration. Despite extensive studies in the past, a protocol for renal lineage induction from embryonic stem cells remains unestablished. In this study, we aimed to induce renal lineages from mouse embryonic stem cells (mESC) by following in vivo developmental stages, i.e., the induction of mesoderm (Stage I), intermediate mesoderm (Stage II) and renal lineages (Stage III). For stage I induction, in accordance with known signaling pathways involved in mesoderm development in vivo, i.e., Nodal, bone morphogenic proteins (BMPs) and Wnt, we found that the sequential addition of three factors, i.e., Activin-A (A), a surrogate for Nodal signaling, during days 0-2, A plus BMP-4 (4) during days 2-4, and A4 plus lithium (L), a surrogate for Wnt signaling, during days 4-6, was most effective to induce the mesodermal marker, Brachyury. For stage II induction, the addition of retinoic acid (R) in the continuous presence of A4L during days 6-8 was most effective to induce nephrogenic intermediate mesodermal markers, such as Pax2 and Lim1. Under this condition, more than 30% of cells were stained positive for Pax2, and there was a concomitant decrease in the expression of non-mesodermal markers. For stage III induction, in resemblance to the reciprocal induction between ureteric bud (UB) and metanephric mesenchyme (MM) during kidney development, we found that the exposure to conditioned media derived from UB and MM cells was

  1. Dorsal-Ventral Patterning and Neural Induction in Xenopus Embryos

    PubMed Central

    De Robertis, Edward M.; Kuroda, Hiroki

    2008-01-01

    We review the current status of research in dorsal-ventral (D-V) patterning in vertebrates. Emphasis is placed on recent work on Xenopus, which provides a paradigm for vertebrate development based on a rich heritage of experimental embryology. D-V patterning starts much earlier than previously thought, under the influence of a dorsal nuclear β-Catenin signal. At mid-blastula two signaling centers are present on the dorsal side: The prospective neuroectoderm expresses bone morphogenetic protein (BMP) antagonists, and the future dorsal endoderm secretes Nodal-related mesoderm-inducing factors. When dorsal mesoderm is formed at gastrula, a cocktail of growth factor antagonists is secreted by the Spemann organizer and further patterns the embryo. A ventral gastrula signaling center opposes the actions of the dorsal organizer, and another set of secreted antagonists is produced ventrally under the control of BMP4. The early dorsal β-Catenin signal inhibits BMP expression at the transcriptional level and promotes expression of secreted BMP antagonists in the prospective central nervous system (CNS). In the absence of mesoderm, expression of Chordin and Noggin in ectoderm is required for anterior CNS formation. FGF (fibroblast growth factor) and IGF (insulin-like growth factor) signals are also potent neural inducers. Neural induction by anti-BMPs such as Chordin requires mitogen-activated protein kinase (MAPK) activation mediated by FGF and IGF. These multiple signals can be integrated at the level of Smad1. Phosphorylation by BMP receptor stimulates Smad1 transcriptional activity, whereas phosphorylation by MAPK has the opposite effect. Neural tissue is formed only at very low levels of activity of BMP-transducing Smads, which require the combination of both low BMP levels and high MAPK signals. Many of the molecular players that regulate D-V patterning via regulation of BMP signaling have been conserved between Drosophila and the vertebrates. PMID:15473842

  2. The Notch Ligand Delta-Like 4 Regulates Multiple Stages of Early Hemato-Vascular Development

    PubMed Central

    Neves, Hélia; Gomes, Andreia C.; Saavedra, Pedro; Carvalho, Catarina C.; Duarte, António; Cidadão, António; Parreira, Leonor

    2012-01-01

    Background In mouse embryos, homozygous or heterozygous deletions of the gene encoding the Notch ligand Dll4 result in early embryonic death due to major defects in endothelial remodeling in the yolk sac and embryo. Considering the close developmental relationship between endothelial and hematopoietic cell lineages, which share a common mesoderm-derived precursor, the hemangioblast, and many key regulatory molecules, we investigated whether Dll4 is also involved in the regulation of early embryonic hematopoiesis. Methodology/Principal Findings Using Embryoid Bodies (EBs) derived from embryonic stem cells harboring hetero- or homozygous Dll4 deletions, we observed that EBs from both genotypes exhibit an abnormal endothelial remodeling in the vascular sprouts that arise late during EB differentiation, indicating that this in vitro system recapitulates the angiogenic phenotype of Dll4 mutant embryos. However, analysis of EB development at early time points revealed that the absence of Dll4 delays the emergence of mesoderm and severely reduces the number of blast-colony forming cells (BL-CFCs), the in vitro counterpart of the hemangioblast, and of endothelial cells. Analysis of colony forming units (CFU) in EBs and yolk sacs from Dll4+/− and Dll4−/− embryos, showed that primitive erythropoiesis is specifically affected by Dll4 insufficiency. In Dll4 mutant EBs, smooth muscle cells (SMCs) were seemingly unaffected and cardiomyocyte differentiation was increased, indicating that SMC specification is Dll4-independent while a normal dose of this Notch ligand is essential for the quantitative regulation of cardiomyogenesis. Conclusions/Significance This study highlights a previously unnoticed role for Dll4 in the quantitative regulation of early hemato-vascular precursors, further indicating that it is also involved on the timely emergence of mesoderm in early embryogenesis. PMID:22514637

  3. Pattern formation during gastrulation in the sea urchin embryo.

    PubMed

    McClay, D R; Armstrong, N A; Hardin, J

    1992-01-01

    The sea urchin embryo follows a relatively simple cell behavioral sequence in its gastrulation movements. To form the mesoderm, primary mesenchyme cells ingress from the vegetal plate and then migrate along the basal lamina lining the blastocoel. The presumptive secondary mesenchyme and endoderm then invaginate from the vegetal pole of the embryo. The archenteron elongates and extends across the blastocoel until the tip of the archenteron touches and attaches to the opposite side of the blastocoel. Secondary mesenchyme cells, originally at the tip of the archenteron, differentiate to form a variety of structures including coelomic pouches, esophageal muscles, pigment cells and other cell types. After migration of the secondary mesenchyme cells from their original position at the tip of the archenteron, the endoderm fuses with an invagination of the ventral ectoderm (the stomodaem), to form the mouth and complete the process of gastrulation. A larval skeleton is made by primary mesenchyme cells during the time of archenteron and mouth formation. A number of experiments have established that these morphogenetic movements involve a number of cell autonomous behaviors plus a series of cell interactions that provide spatial, temporal and scalar information to cells of the mesoderm and endoderm. The cell autonomous behaviors can be demonstrated by the ability of micromeres or endoderm to perform their morphogenetic functions if either is isolated and grown in culture. The requirement for cell interactions has been demonstrated by manipulative experiments where it has been shown that axial information, temporal information, spatial information and scalar information is obtained by mesoderm and endoderm from other embryonic cells.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:1299366

  4. Human adipose tissue possesses a unique population of pluripotent stem cells with nontumorigenic and low telomerase activities: potential implications in regenerative medicine.

    PubMed

    Ogura, Fumitaka; Wakao, Shohei; Kuroda, Yasumasa; Tsuchiyama, Kenichiro; Bagheri, Mozhdeh; Heneidi, Saleh; Chazenbalk, Gregorio; Aiba, Setsuya; Dezawa, Mari

    2014-04-01

    In this study, we demonstrate that a small population of pluripotent stem cells, termed adipose multilineage-differentiating stress-enduring (adipose-Muse) cells, exist in adult human adipose tissue and adipose-derived mesenchymal stem cells (adipose-MSCs). They can be identified as cells positive for both MSC markers (CD105 and CD90) and human pluripotent stem cell marker SSEA-3. They intrinsically retain lineage plasticity and the ability to self-renew. They spontaneously generate cells representative of all three germ layers from a single cell and successfully differentiate into targeted cells by cytokine induction. Cells other than adipose-Muse cells exist in adipose-MSCs, however, do not exhibit these properties and are unable to cross the boundaries from mesodermal to ectodermal or endodermal lineages even under cytokine inductions. Importantly, adipose-Muse cells demonstrate low telomerase activity and transplants do not promote teratogenesis in vivo. When compared with bone marrow (BM)- and dermal-Muse cells, adipose-Muse cells have the tendency to exhibit higher expression in mesodermal lineage markers, while BM- and dermal-Muse cells were generally higher in those of ectodermal and endodermal lineages. Adipose-Muse cells distinguish themselves as both easily obtainable and versatile in their capacity for differentiation, while low telomerase activity and lack of teratoma formation make these cells a practical cell source for potential stem cell therapies. Further, they will promote the effectiveness of currently performed adipose-MSC transplantation, particularly for ectodermal and endodermal tissues where transplanted cells need to differentiate across the lineage from mesodermal to ectodermal or endodermal in order to replenish lost cells for tissue repair. PMID:24256547

  5. Downregulation of H19 improves the differentiation potential of mouse parthenogenetic embryonic stem cells.

    PubMed

    Ragina, Neli P; Schlosser, Karianne; Knott, Jason G; Senagore, Patricia K; Swiatek, Pamela J; Chang, Eun Ah; Fakhouri, Walid D; Schutte, Brian C; Kiupel, Matti; Cibelli, Jose B

    2012-05-01

    Parthenogenetic embryonic stem cells (P-ESCs) offer an alternative source of pluripotent cells, which hold great promise for autologous transplantation and regenerative medicine. P-ESCs have been successfully derived from blastocysts of several mammalian species. However, compared with biparental embryonic stem cells (B-ESCs), P-ESCs are limited in their ability to fully differentiate into all 3 germ layers. For example, it has been observed that there is a differentiation bias toward ectoderm derivatives at the expense of endoderm and mesoderm derivatives-muscle in particular-in chimeric embryos, teratomas, and embryoid bodies. In the present study we found that H19 expression was highly upregulated in P-ESCs with more than 6-fold overexpression compared with B-ESCs. Thus, we hypothesized that manipulation of the H19 gene in P-ESCs would alleviate their limitations and allow them to function like B-ESCs. To test this hypothesis we employed a small hairpin RNA approach to reduce the amount of H19 transcripts in mouse P-ESCs. We found that downregulation of H19 led to an increase of mesoderm-derived muscle and endoderm in P-ESCs teratomas similar to that observed in B-ESCs teratomas. This phenomenon coincided with upregulation of mesoderm-specific genes such as Myf5, Myf6, and MyoD. Moreover, H19 downregulated P-ESCs differentiated into a higher percentage of beating cardiomyocytes compared with control P-ESCs. Collectively, these results suggest that P-ESCs are amenable to molecular modifications that bring them functionally closer to true ESCs. PMID:21793658

  6. Downregulation of H19 Improves the Differentiation Potential of Mouse Parthenogenetic Embryonic Stem Cells

    PubMed Central

    Ragina, Neli P.; Schlosser, Karianne; Knott, Jason G.; Senagore, Patricia K.; Swiatek, Pamela J.; Chang, Eun Ah; Fakhouri, Walid D.; Schutte, Brian C.; Kiupel, Matti

    2012-01-01

    Parthenogenetic embryonic stem cells (P-ESCs) offer an alternative source of pluripotent cells, which hold great promise for autologous transplantation and regenerative medicine. P-ESCs have been successfully derived from blastocysts of several mammalian species. However, compared with biparental embryonic stem cells (B-ESCs), P-ESCs are limited in their ability to fully differentiate into all 3 germ layers. For example, it has been observed that there is a differentiation bias toward ectoderm derivatives at the expense of endoderm and mesoderm derivatives—muscle in particular—in chimeric embryos, teratomas, and embryoid bodies. In the present study we found that H19 expression was highly upregulated in P-ESCs with more than 6-fold overexpression compared with B-ESCs. Thus, we hypothesized that manipulation of the H19 gene in P-ESCs would alleviate their limitations and allow them to function like B-ESCs. To test this hypothesis we employed a small hairpin RNA approach to reduce the amount of H19 transcripts in mouse P-ESCs. We found that downregulation of H19 led to an increase of mesoderm-derived muscle and endoderm in P-ESCs teratomas similar to that observed in B-ESCs teratomas. This phenomenon coincided with upregulation of mesoderm-specific genes such as Myf5, Myf6, and MyoD. Moreover, H19 downregulated P-ESCs differentiated into a higher percentage of beating cardiomyocytes compared with control P-ESCs. Collectively, these results suggest that P-ESCs are amenable to molecular modifications that bring them functionally closer to true ESCs. PMID:21793658

  7. Clarifying tetrapod embryogenesis by a dorso-ventral analysis of the tissue flows during early stages of chicken development.

    PubMed

    Fleury, Vincent

    2012-09-01

    The formation of an animal body remains largely a mystery. It is still not clear whether anything like an organization plan or an "archetype" as coined by Darwin himself, actually exists, or whether animals are organized by a succession of stop-and-go genetic, non-linear, instructions with no global pattern. Nevertheless, it was recognized long ago that the early stages of amniote development consist of large scale rotatory movements over a discoidal blastula (Wetzel, 1924). Such rotatory movements reshuffle a mass inside a finite volume, and thus may have to bear physical conservation laws which contribute to establish the plan of animals in a global fashion. In this article I use dual dorso-ventral imaging of the chicken blastula, to show experimentally that the global movement of early vertebrate embryogenesis is organized with a very simple topology, around and away of a series of hyperbolic points in the vector flow of movement. At the first hyperbolic point, a layer of tissue (the mesoderm) ingresses and moves as a viscous sheet radially. It is found that the sheet flows away with a scaling law for the radius R(t)∼exp(t/τ). Also, the movement of this mesoderm changes the flow on the other layer (the ectoderm) by the principle of action and reaction. By mesoderm wetting the ectoderm, the first hyperbolic point migrates from the anal region, to the umbilical region. The final location of the hyperbolic point defines eventually the central part of the body (the umbilical region). Thus, the formation of the vertebrate body is fixed, as a global movement, by the dynamics of singular points in the visco-elastic flow, governed by mechanical forces within the tissue. PMID:22564883

  8. The hedgehog Pathway Gene shifted Functions together with the hmgcr-Dependent Isoprenoid Biosynthetic Pathway to Orchestrate Germ Cell Migration

    PubMed Central

    Deshpande, Girish; Zhou, Keren; Wan, Joy Y.; Friedrich, Jana; Jourjine, Nicholas; Smith, Daniel; Schedl, Paul

    2013-01-01

    The Drosophila embryonic gonad is assembled from two distinct cell types, the Primordial Germ Cells (PGCs) and the Somatic Gonadal Precursor cells (SGPs). The PGCs form at the posterior of blastoderm stage embryos and are subsequently carried inside the embryo during gastrulation. To reach the SGPs, the PGCs must traverse the midgut wall and then migrate through the mesoderm. A combination of local repulsive cues and attractive signals emanating from the SGPs guide migration. We have investigated the role of the hedgehog (hh) pathway gene shifted (shf) in directing PGC migration. shf encodes a secreted protein that facilitates the long distance transmission of Hh through the proteoglycan matrix after it is released from basolateral membranes of Hh expressing cells in the wing imaginal disc. shf is expressed in the gonadal mesoderm, and loss- and gain-of-function experiments demonstrate that it is required for PGC migration. Previous studies have established that the hmgcr-dependent isoprenoid biosynthetic pathway plays a pivotal role in generating the PGC attractant both by the SGPs and by other tissues when hmgcr is ectopically expressed. We show that production of this PGC attractant depends upon shf as well as a second hh pathway gene gγ1. Further linking the PGC attractant to Hh, we present evidence indicating that ectopic expression of hmgcr in the nervous system promotes the release/transmission of the Hh ligand from these cells into and through the underlying mesodermal cell layer, where Hh can contact migrating PGCs. Finally, potentiation of Hh by hmgcr appears to depend upon cholesterol modification. PMID:24068944

  9. Jagged1 functions downstream of Twist1 in the specification of the coronal suture and the formation of a boundary between osteogenic and non-osteogenic cells

    PubMed Central

    Yen, Hai-Yun; Ting, Man-Chun; Maxson, Robert E.

    2011-01-01

    The Notch pathway is crucial for a wide variety of developmental processes including the formation of tissue boundaries. That it may function in calvarial suture development and figure in the pathophysiology of craniosynostosis was suggested by the demonstration that heterozygous loss of function of JAGGED1 in humans can cause Alagille syndrome, which has craniosynostosis as a feature. We used conditional gene targeting to examine the role of Jagged1 in the development of the skull vault. We demonstrate that Jagged1 is expressed in a layer of mesoderm-derived sutural cells that lie along the osteogenic-non-osteogenic boundary. We show that inactivation of Jagged1 in the mesodermal compartment of the coronal suture, but not in the neural crest compartment, results in craniosynostosis. Mesodermal inactivation of Jagged1 also results in changes in the identity of sutural cells prior to overt osteogenic differentiation, as well as defects in the boundary between osteogenic and non-osteogenic compartments at the coronal suture. These changes, surprisingly, are associated with increased expression of Notch2 and the Notch effector, Hes1, in the sutural mesenchyme. They are also associated with an increase in nuclear β-catenin. In Twist1 mutants, Jagged1 expression in the suture is reduced substantially, suggesting an epistatic relationship between Twist1 and Jagged1. Consistent with such a relationship, Twist1-Jagged1 double heterozygotes exhibit a substantial increase in the severity of craniosynostosis over individual heterozygotes. Our results thus suggest that Jagged1 is an effector of Twist1 in coronal suture development. PMID:20727876

  10. Evolutionarily conserved morphogenetic movements at the vertebrate head–trunk interface coordinate the transport and assembly of hypopharyngeal structures

    PubMed Central

    Lours-Calet, Corinne; Alvares, Lucia E.; El-Hanfy, Amira S.; Gandesha, Saniel; Walters, Esther H.; Sobreira, Débora Rodrigues; Wotton, Karl R.; Jorge, Erika C.; Lawson, Jennifer A.; Kelsey Lewis, A.; Tada, Masazumi; Sharpe, Colin; Kardon, Gabrielle; Dietrich, Susanne

    2014-01-01

    The vertebrate head–trunk interface (occipital region) has been heavily remodelled during evolution, and its development is still poorly understood. In extant jawed vertebrates, this region provides muscle precursors for the throat and tongue (hypopharyngeal/hypobranchial/hypoglossal muscle precursors, HMP) that take a stereotype path rostrally along the pharynx and are thought to reach their target sites via active migration. Yet, this projection pattern emerged in jawless vertebrates before the evolution of migratory muscle precursors. This suggests that a so far elusive, more basic transport mechanism must have existed and may still be traceable today. Here we show for the first time that all occipital tissues participate in well-conserved cell movements. These cell movements are spearheaded by the occipital lateral mesoderm and ectoderm that split into two streams. The rostrally directed stream projects along the floor of the pharynx and reaches as far rostrally as the floor of the mandibular arch and outflow tract of the heart. Notably, this stream leads and engulfs the later emerging HMP, neural crest cells and hypoglossal nerve. When we (i) attempted to redirect hypobranchial/hypoglossal muscle precursors towards various attractants, (ii) placed non-migratory muscle precursors into the occipital environment or (iii) molecularly or (iv) genetically rendered muscle precursors non-migratory, they still followed the trajectory set by the occipital lateral mesoderm and ectoderm. Thus, we have discovered evolutionarily conserved morphogenetic movements, driven by the occipital lateral mesoderm and ectoderm, that ensure cell transport and organ assembly at the head–trunk interface. PMID:24662046