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Patrick P L Tam - One of the best experts on this subject based on the ideXlab platform.
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7 somitogenesis segmentation of the Paraxial Mesoderm and the delineation of tissue compartments
Mouse Development#R##N#Patterning Morphogenesis and Organogenesis, 2002Co-Authors: Achim Gossler, Patrick P L TamAbstract:This chapter discusses the segmentation of the Paraxial Mesoderm and the delineation of tissue compartments. Segmentation is a fundamental developmental process that subdivides the body, or parts thereof, into a series of serially repeated subunits and thereby generates a segmental pattern. In the mouse embryo, like other vertebrate embryos, the earliest manifestation of tissue segmentation is the formation of somites during organogenesis. Somites are blocks of Mesodermal cells located on either side of the neural tube and the notochord. Together with the mesenchyme that envelops the cephalic neural tube, they constitute the Paraxial Mesoderm of the embryo. The segmental arrangement of the somites along the anterior–posterior body axis prefigures and underlies the metarnerism of the somite derived vertebral column and epaxial muscles and also determines the segmented arrangement of parts of the peripheral nervous system.
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early events of somitogenesis in higher vertebrates allocation of precursor cells during gastrulation and the organization of a meristic pattern in the Paraxial Mesoderm
Current Topics in Developmental Biology, 1999Co-Authors: Patrick P L Tam, Devorah Goldman, Anne Camus, Gary C. SchoenwolfAbstract:Publisher Summary Through the fate mapping of mouse and avian embryos, the localization of the somite precursors in the germ layers, the primitive streak, and the tail bud has been extensively elucidated. A stage-by-stage study of the localization of the precursor population and the distribution of the clonal descendants of labeled or transplanted cells has enabled the reconstruction of the morphogenetic movement of the prospective somitic cells during gastrulation and early organogenesis. Studies mouse and avians have revealed substantial homology in the allocation of cell lineages, the morphogenesis of the Paraxial Mesoderm, and the specification of the somites, such that some generalization may be warranted for both species. Results of fate mapping experiments have shown that the primitive streak and the tail bud are continuous sources of new cells for somitogenesis. It is currently not known how this self-renewing population is maintained and how the prospective somitic cells are incorporated into the Paraxial Mesoderm. Of significant interest is how the somitogenic potential of this self-renewing population is regulated, especially toward the cessation of somitogenesis.
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cranial Paraxial Mesoderm and neural crest cells of the mouse embryo co distribution in the craniofacial mesenchyme but distinct segregation in branchial arches
Development, 1995Co-Authors: Paul A Trainor, Patrick P L TamAbstract:The spatial distribution of the cranial Paraxial Mesoderm and the neural crest cells during craniofacial morphogenesis of the mouse embryo was studied by micromanipulative cell grafting and cell labelling. Results of this study show that the Paraxial Mesoderm and neural crest cells arising at the same segmental position share common destinations. Mesodermal cells from somitomeres I, III, IV and VI were distributed to the same craniofacial tissues as neural crest cells of the forebrain, the caudal midbrain, and the rostral, middle and caudal hindbrains found respectively next to these Mesodermal segments. This finding suggests that a basic meristic pattern is established globally in the neural plate ectoderm and Paraxial Mesoderm during early mouse development. Cells from these two sources mixed extensively in the peri-ocular, facial, periotic and cervical mesenchyme. However, within the branchial arches a distinct segregation of these two cell populations was discovered. Neural crest cells colonised the periphery of the branchial arches and enveloped the somitomere-derived core tissues on the rostral, lateral and caudal sides of the arch. Such segregation of cell populations in the first three branchial arches is apparent at least until the 10.5-day hindlimb bud stage and could be important for the patterning of the skeletal and myogenic derivatives of the arches.
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cranial Paraxial Mesoderm regionalisation of cell fate and impact on craniofacial development in mouse embryos
Development, 1994Co-Authors: Paul A Trainor, Seongseng Tan, Patrick P L TamAbstract:A combination of micromanipulative cell grafting and fluorescent cell labelling techniques were used to examine the developmental fate of the cranial Paraxial Mesoderm of the 8.5-day early-somite-stage mouse embryo. Mesodermal cells isolated from seven regions of the cranial Mesoderm, identified on the basis of their topographical association with specific brain segments were assessed for their contribution to craniofacial morphogenesis during 48 hours of in vitro development. The results demonstrate extensive cell mixing between adjacent but not alternate groups of Mesodermal cells and a strict cranial-to-caudal distribution of the Paraxial Mesoderm to craniofacial structures. A two-segment periodicity similar to the origins of the branchial motor neurons and the distribution of the rhombencephalic neural crest cells was observed as the Paraxial Mesoderm migrates during formation of the first three branchial arches. The Paraxial Mesoderm colonises the mesenchymal core of the branchial arches, consistent with the location of the muscle plates. A dorsoventral regionalisation of cell fate similar to that of the somitic Mesoderm is also found. This suggests evolution has conserved the fate of the murine cranial Paraxial Mesoderm as a multiprogenitor population which displays a predominantly myogenic fate. Heterotopic transplantation of cells to different regions of the cranial Mesoderm revealed no discernible restriction in cell potency in the craniocaudal axis, reflecting considerable plasticity in the developmental fate of the cranial Mesoderm at least at the time of experimentation. The distribution of the different groups of cranial Mesoderm matches closely with that of the cranial neural crest cells suggesting the two cell populations may share a common segmental origin and similar destination.
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specification and segmentation of the Paraxial Mesoderm
Anatomy and Embryology, 1994Co-Authors: Patrick P L Tam, Paul A TrainorAbstract:Somite formation in the mouse embryo begins with the recruitment of mesenchymal cells into the Paraxial Mesoderm. Cells destined for the Paraxial Mesoderm are recruited from a progenitor population found first in the embryonic ectoderm and later in the primitive streak and the tail bud. Experimental evidence suggests that the allocation of precursor cells to different Mesodermal lineages may be related to the site at which the cells ingress through the primitive streak. An increasing number of genes, such as those encoding growth factor and transcription factors, are now known to be expressed in the primitive streak. It is not known whether the specification of Mesodermal cell fate has any relationship with the activity of genes that are expressed in the restricted cell populations of the primitive streak. Somitomeres, which are spherical clusters of mesenchymal cells in the presomitic Mesoderm, presage the segmentation of somites in the Paraxial Mesoderm. The somitomeric organization denotes a pre-pattern of segmentation that defines the physical boundary and the bilateral symmetry of the Mesodermal segments in the body axis. The establishment of new somitomeres seems to require the interaction of a resident cell population in the presomitic Mesoderm and the incoming primitive streak cells. Cell mixing, which occurs in the somitomeres prior to somite segmentation, poses problems in understanding the developmental role of the somitomere and the real significance of the partitioning of the node-derived and primitive streak-derived cells in the Mesodermal segments. In the presomitic Mesoderm, the expression of some genes that encode transcription factors, growth factors or tyrosine kinase receptor, and the localization of certain cell adhesion molecules are closely associated with distinct morphogenetic events, such as cell clustering in the presomitic Mesoderm and the formation of epithelial somites. There is, however, very little direct relationship between the spatial pattern of gene expression and the somitomeric organization in the presomitic Mesoderm. Results of somite transplantation experiments suggest that both the segmental address and the morphogenetic characteristics of the somite may be determined during somite segmentation. Regional identity of the Paraxial Mesodermal segment is conferred by the expression of a combination of Hox genes in the sclerotome and probably other lineage-specific genes that are subject to imprinting. Superimposed on the global metameric pattern, two orthogonal polarities of cell differentiation are endowed in each Mesodermal segment. The rostro-caudal polarity is established prior to somite segmentation. This polarity is later manifested by the subdivision of the sclerotome and the alliance of the neural crest cells and motor axons with the rostral half-somite.(ABSTRACT TRUNCATED AT 400 WORDS)
Olivier Pourquié - One of the best experts on this subject based on the ideXlab platform.
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Paraxial Mesoderm organoids model development of human somites
bioRxiv, 2021Co-Authors: Christoph Budjan, Olivier Pourquié, Sophia Liu, Adrian Ranga, S Gayen, Sahand HormozAbstract:Abstract During the development of the vertebrate embryo, segmented structures called somites are periodically formed from the presomitic Mesoderm (PSM), and give rise to the vertebral column. While somite formation has been studied in several animal models, it is less clear how well this process is conserved in humans. Recent progress has made it possible to study aspects of human Paraxial Mesoderm development such as the human segmentation clock in vitro using human pluripotent stem cells (hPSCs), however, somite formation has not been observed in these monolayer cultures. Here, we describe the generation of human Paraxial Mesoderm (PM) organoids from hPSCs (termed Somitoids), which recapitulate the molecular, morphological and functional features of Paraxial Mesoderm development, including formation of somite-like structures in vitro. Using a quantitative image-based screen, we identify critical parameters such as initial cell number and signaling modulations that reproducibly yielded somite formation in our organoid system. In addition, using single-cell RNA sequencing and 3D imaging, we show that PM organoids both transcriptionally and morphologically resemble their in vivo counterparts and can be differentiated into somite derivatives. Our organoid system is reproducible and scalable, allowing for the systematic and quantitative analysis of human spinal cord development and disease in vitro.
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Signaling Gradients during Paraxial Mesoderm Development
Cold Spring Harbor perspectives in biology, 2009Co-Authors: Alexander Aulehla, Olivier PourquiéAbstract:The sequential formation of somites along the anterior-posterior axis is under control of multiple signaling gradients involving the Wnt, FGF, and retinoic acid (RA) pathways. These pathways show graded distribution of signaling activity within the Paraxial Mesoderm of vertebrate embryos. Although Wnt and FGF signaling show highest activity in the posterior, unsegmented Paraxial Mesoderm (presomitic Mesoderm [PSM]), RA signaling establishes a countergradient with the highest activity in the somites. The generation of these graded activities relies both on classical source-sink mechanisms (for RA signaling) and on an RNA decay mechanism (for FGF signaling). Numerous studies reveal the tight interconnection among Wnt, FGF, and RA signaling in controlling Paraxial Mesoderm differentiation and in defining the somite-forming unit. In particular, the relationship to a molecular oscillator acting in somite precursors in the PSM-called the segmentation clock-has been recently addressed. These studies indicate that high levels of Wnt and FGF signaling are required for the segmentation clock activity. Furthermore, we discuss how these signaling gradients act in a dose-dependent manner in the progenitors of the Paraxial Mesoderm, partly by regulating cell movements during gastrulation. Finally, links between the process of axial specification of vertebral segments and Hox gene expression are discussed.
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Dual mode of Paraxial Mesoderm formation during chick gastrulation
Proceedings of the National Academy of Sciences of the United States of America, 2007Co-Authors: Tadahiro Iimura, Xuesong Yang, Cornelis J. Weijer, Olivier PourquiéAbstract:The skeletal muscles and axial skeleton of vertebrates derive from the embryonic Paraxial Mesoderm. In amniotes, Paraxial Mesoderm is formed bilaterally to the nerve cord as a result of primitive streak and tail-bud regression during body axis formation. In chick and mouse embryos, Paraxial Mesoderm was proposed to derive from a population of resident cells located in the regressing primitive streak and tail bud. In contrast, in lower vertebrates, Paraxial Mesoderm is formed as a result of the continuation of ingression movements of gastrulation. Here, we reinvestigate Paraxial Mesoderm formation in the chicken embryo and demonstrate that these two modes are concomitantly at work to set up the Paraxial Mesoderm. Although the medial part of somites derives from stem cells resident in the primitive streak/tail bud, the lateral part derives from continuous ingression of epiblastic material. Our fate mapping further shows that the Paraxial Mesoderm territory in the epiblast is regionalized along the anteroposterior axis as in lower vertebrates. These observations suggest that the mechanisms responsible for Paraxial Mesoderm formation are largely conserved across vertebrates.
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vertebrate somitogenesis a novel paradigm for animal segmentation
The International Journal of Developmental Biology, 2003Co-Authors: Olivier PourquiéAbstract:In vertebrates, the primary segmented tissue of the body axis is the Paraxial Mesoderm, which lies bilaterally to the axial organs, neural tube and notochord. The segmental pattern of the Paraxial Mesoderm is established during embryogenesis through the production of the somites which are transient embryonic segments giving rise to the vertebrae, the skeletal muscles and the dorsal dermis. Somitogenesis can be subdivided into three major phases (see Fig. 1). First a growth phase during which new Paraxial Mesoderm cells are produced by a growth zone (epiblast and blastopore margin or primitive streak and later on tail bud) and become organized as two rods of mesenchymal tissue,forming the presomitic Mesoderm. Second a patterning phase occuring in the PSM, during which the segmental pattern is established at the molecular level. Third, the somitic boundaries are formed during the morphological segmentation phase. In all vertebrates, all cells of the Paraxial Mesoderm, during their maturation in the PSM, go successively through these three phases, which are tightly regulated at the spatio-temporal level. The first phase of Paraxial Mesoderm production falls out of the scope of this review, as it essentially pertains to the gastrulation process. Here, I essentially discuss the segmental patterning phase in vertebrates. Recent data suggest that establishment of the segmental pattern relies on a clock and wavefront mechanism which has been conserved in vertebrates. Furthermore, conservation of this system could extend to invertebrates, suggesting that the clock and wavefront is an ancestral mechanism.
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Segmentation of the Paraxial Mesoderm and vertebrate somitogenesis.
Current topics in developmental biology, 2000Co-Authors: Olivier PourquiéAbstract:Somites are the most obviously segmented features of the vertebrate embryo. Although the way segmentation is achieved in the fly is now well described, little was known about the molecular mechanisms underlying vertebrate somitogenesis. Through the recent identification of genes important for vertebrate somitogenesis and the analysis of their function, several theoretical models accounting for somitogenesis such as the clock and wavefront model, which have been proposed over the past 20 years, are now starting to receive experimental support. A molecular clock linked to somitogenesis has been identified which might act as a periodicity generator in the presomitic cells. This temporal periodicity is then translated into a tightly controlled spatial periodicity which is revealed by the expression of several genes. Analysis of mouse mutants in the Notch-Delta pathway suggest that this signaling mechanism might play an important role at this level. The final step of the cascade is to translate these genetically specified segments into morphological units: the somites. Importantly, these studies have helped in dissociating the segmentation and the somitogenesis processes in vertebrates. In addition, although segmentation was classically thought to have arisen independently in protostomes and deuterostomes, recent evidence suggests that part of the segmentation machinery might actually have been conserved. The conservation of segmentation mechanisms reported in the fly such as the pair-rule pattern, however, remain a subject of controversy.
Virginia E. Papaioannou - One of the best experts on this subject based on the ideXlab platform.
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Interaction of Wnt3a, Msgn1 and Tbx6 in neural versus Paraxial Mesoderm lineage commitment and Paraxial Mesoderm differentiation in the mouse embryo.
Developmental biology, 2012Co-Authors: Sonja Nowotschin, Anna Ferrer-vaquer, Daniel Concepcion, Virginia E. Papaioannou, Anna-katerina HadjantonakisAbstract:Paraxial Mesoderm is the tissue which gives rise to the skeletal muscles and vertebral column of the body. A gene regulatory network operating in the formation of Paraxial Mesoderm has been described. This network hinges on three key factors, Wnt3a, Msgn1 and Tbx6, each of which is critical for Paraxial Mesoderm formation, since absence of any one of these factors results in complete absence of posterior somites. In this study we determined and compared the spatial and temporal patterns of expression of Wnt3a, Msgn1 and Tbx6 at a time when Paraxial Mesoderm is being formed. Then, we performed a comparative characterization of mutants in Wnt3a, Msgn1 and Tbx6. To determine the epistatic relationship between these three genes, and begin to decipher the complex interplay between them, we analyzed double mutant embryos and compared their phenotypes to the single mutants. Through the analysis of molecular markers in mutants, our data support the bipotential nature of the progenitor cells for Paraxial Mesoderm and establish regulatory relationships between genes involved in the choice between neural and Mesoderm fates.
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tbx6 dependent sox2 regulation determines neural or Mesodermal fate in axial stem cells
Nature, 2011Co-Authors: Tatsuya Takemoto, Virginia E. Papaioannou, Masanori Uchikawa, Megumi Yoshida, Donald M Bell, Robin Lovellbadge, Hisato KondohAbstract:The classical view of neural plate development held that it arises from the ectoderm, after its separation from the Mesodermal and endodermal lineages. However, recent cell-lineage-tracing experiments indicate that the caudal neural plate and Paraxial Mesoderm are generated from common bipotential axial stem cells originating from the caudal lateral epiblast. Tbx6 null mutant mouse embryos which produce ectopic neural tubes at the expense of Paraxial Mesoderm must provide a clue to the regulatory mechanism underlying this neural versus Mesodermal fate choice. Here we demonstrate that Tbx6-dependent regulation of Sox2 determines the fate of axial stem cells. In wild-type embryos, enhancer N1 of the neural primordial gene Sox2 is activated in the caudal lateral epiblast, and the cells staying in the superficial layer sustain N1 activity and activate Sox2 expression in the neural plate. In contrast, the cells destined to become Mesoderm activate Tbx6 and turn off enhancer N1 before migrating into the Paraxial Mesoderm compartment. In Tbx6 mutant embryos, however, enhancer N1 activity persists in the Paraxial Mesoderm compartment, eliciting ectopic Sox2 activation and transforming the Paraxial Mesoderm into neural tubes. An enhancer-N1-specific deletion mutation introduced into Tbx6 mutant embryos prevented this Sox2 activation in the Mesodermal compartment and subsequent development of ectopic neural tubes, indicating that Tbx6 regulates Sox2 via enhancer N1. Tbx6-dependent repression of Wnt3a in the Paraxial Mesodermal compartment is implicated in this regulatory process. Paraxial Mesoderm-specific misexpression of a Sox2 transgene in wild-type embryos resulted in ectopic neural tube development. Thus, Tbx6 represses Sox2 by inactivating enhancer N1 to inhibit neural development, and this is an essential step for the specification of Paraxial Mesoderm from the axial stem cells.
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tbx6 a mouse t box gene implicated in Paraxial Mesoderm formation at gastrulation
Developmental Biology, 1996Co-Authors: Deborah L Chapman, Irina Agulnik, Sarah Hancock, Lee M Silver, Virginia E. PapaioannouAbstract:Abstract The T-box genes constitute an evolutionarily conserved family of putative transcription factors which are expressed in discrete domains during embryogenesis, suggesting that they may play roles in inductive interactions. Members have been identified by virtue of their homology to the prototypical T-box gene,TorBrachyury,which is required for Mesoderm formation and axial elongation during embryogenesis. We have previously reported the discovery of six new mouse T-box genes,Tbx1–Tbx6,and described the expression patterns ofTbx1–Tbx5(Bollaget al.,1994; Agulniket al.,1996; Chapmanet al.,1996; Gibson-Brownet al.,1996). We have obtained cDNA clones encoding the full-length Tbx6 protein from screens of gastrulation-stage mouse cDNA libraries and determined the spatial and temporal distribution ofTbx6transcripts during embryogenesis. The gene codes for a 1.9-kb transcript with an open reading frame coding for a 540-amino acid protein, with a predicted molecular weight of 59 kDa.Tbx6maps to chromosome 7 and does not appear to be linked to any known mutation. Unlike other members of the mouse T-box gene family which are expressed in a wide variety of tissues derived from all germ layers,Tbx6expression is quite restricted.Tbx6transcripts are first detected in the gastrulation stage embryo in the primitive streak and newly recruited Paraxial Mesoderm. Later in development,Tbx6expression is restricted to presomitic, Paraxial Mesoderm and to the tail bud, which replaces the streak as the source of Mesoderm. Expression in the tail bud persists until 12.5 days postcoitus.Tbx6expression thus overlaps that ofBrachyuryin the primitive streak and tail bud, althoughBrachyuryis expressed earlier in the primitive streak.Brachyuryis also expressed in a second domain, the node and notochord, that is not shared withTbx6.The onset ofTbx6expression is not affected in homozygous nullBrachyurymutant embryos at 7.5 days postcoitus. However,Tbx6expression is extinguished in mutant embryos as soon as theBrachyuryphenotype becomes evident at 8.5 days postcoitus, indicating that the continued expression ofTbx6is directly or indirectly dependent uponBrachyuryexpression.
Anne H Monsoroburq - One of the best experts on this subject based on the ideXlab platform.
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intracellular enhancement of bmp signaling by lim domain protein fhl3 controls spatiotemporal emergence of the neural crest driven by wnt signaling
Social Science Research Network, 2020Co-Authors: Mansour Alkobtawi, Patrick Pla, Anne H MonsoroburqAbstract:How multiple morphogens are coordinated in space and time to position key embryonic tissues remains elusive. During neural crest formation, bone morphogenetic protein (BMP), fibroblast growth factor (FGF) and WNT signaling cooperate by acting either on the Paraxial Mesoderm or directly on the neural border ectoderm, but how each tissue interprets this complex information remains poorly understood. Here we show that Fhl3, a scaffold LIM domain protein of previously unknown developmental function, is essential for neural crest formation by linking BMP and WNT signaling thereby positioning the neural crest-inducing signaling center in the Paraxial Mesoderm. During gastrulation, Fhl3 promotes Smad phosphorylation and Smad-dependent wnt8 activation in the Paraxial Mesoderm, thus enhancing the response of Mesoderm compared to ectoderm although both germ layers are subjected to a common extracellular BMP gradient. This promotes a non-cell autonomous WNT signaling ensuring neural border ectoderm specification by the underlying Mesoderm. During neurulation, in turn, neural crest inducers activate fhl3 in the ectoderm, promoting BMP/Smad-dependent WNT activity required for neural crest specification. We show that Fhl3 binds Smads downstream of BMP signals and promotes Smad binding to wnt8 regulatory elements. Our findings highlight how Fhl3, acting cell-autonomously, ensures a fine spatial and temporal coordination of BMP and WNT signaling at several steps of neural crest development.
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intracellular enhancement of bmp signaling by lim domain protein fhl3 controls spatiotemporal emergence of the neural crest driven by wnt signaling
bioRxiv, 2019Co-Authors: Mansour Alkobtawi, Patrick Pla, Anne H MonsoroburqAbstract:Abstract How multiple morphogen signals are coordinated in space and time to position key embryonic tissues remains elusive. During neural crest formation, bone morphogenetic protein (BMP), fibroblast growth factor (FGF) and WNT signaling cooperate by acting either on the Paraxial Mesoderm or directly on the neural border ectoderm, but how each tissue interprets this complex information remains poorly understood. Here we show that Fhl3, a scaffold LIM domain protein of previously unknown developmental function, is essential for neural crest formation by linking BMP and WNT signaling thereby positioning the neural crest-inducing signaling center in the Paraxial Mesoderm. During gastrulation, Fhl3 promotes Smad phosphorylation and Smad-dependent wnt8 activation specifically in the Paraxial Mesoderm, thus modifying the respective Mesoderm or ectoderm cell response to the extracellular BMP gradient. This ensures neural border ectoderm specification by the underlying Mesoderm via non-cell autonomous WNT signaling. During neurulation, neural crest inducers activate fhl3, promoting BMP/Smad-dependent WNT activity required for neural crest specification. Our findings highlight how Fhl3, acting cell-autonomously, ensures a fine spatial, temporal and germ layer-specific coordination of BMP and WNT signaling at several steps of neural crest development. Highlights: -FHL3 is a novel intracellular enhancer of BMP signaling during early development. -FHL3 ensures cross-talk between BMP and WNT signaling by Smad1-dependent wnt8 activation in the Paraxial Mesoderm. -FHL3 reiterated function in Paraxial Mesoderm and in neural border ectoderm is essential for neural crest development at the border of the neural plate.
Margaret Buckingham - One of the best experts on this subject based on the ideXlab platform.
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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
Development, 1998Co-Authors: Shahragim Tajbakhsh, E Vivarelli, Jackie Papkoff, Robert G. Kelly, Ugo Borello, Margaret Buckingham, Delphine Duprez, Giulio CossuAbstract: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.
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activation of different myogenic pathways myf 5 is induced by the neural tube and myod by the dorsal ectoderm in mouse Paraxial Mesoderm
Development, 1996Co-Authors: Giulio Cossu, Shahragim Tajbakhsh, E Vivarelli, Robert G. Kelly, S Di Donna, Margaret BuckinghamAbstract:Newly formed somites or unsegmented Paraxial Mesoderm (UPM) have been cultured either in isolation or with adjacent structures to investigate the influence of these tissues on myogenic differentiation in mammals. The extent of differentiation was easily and accurately quantified by counting the number of beta-galactosidase-positive cells, since Mesodermal tissues had been isolated from transgenic mice that carry the n-lacZ gene under the transcriptional control of a myosin light chain promoter, restricting expression to striated muscle. The results obtained showed that axial structures are necessary to promote differentiation of Paraxial Mesoderm, in agreement with previous observations. However, it also appeared that the influence of axial structures could be replaced by dorsolateral tissues, adjacent to the Paraxial Mesoderm. To elucidate which of these tissues exerts this positive effect, we cultured the Paraxial Mesoderm with a variety of adjacent structures, either adherent to the Mesoderm or recombined in vitro. The results of these experiments indicated that the dorsal ectoderm exerts a positive influence on myogenesis but only if left in physical proximity to it. In contrast, lateral Mesoderm delays the positive effect of the ectoderm (and has no effect on its own) suggesting that this tissue produces an inhibitory signal. To investigate whether axial structures and dorsal ectoderm induce myogenesis through common or separate pathways, we dissected the medial half of the unsegmented Paraxial Mesoderm and cultured it with the adjacent neural tube. We also cultured the lateral half of the unsegmented Paraxial Mesoderm with adjacent ectoderm. The induction of the myogenic regulatory factors myf-5 and MyoD was monitored by double staining of cultured cells with antibodies against MyoD and beta-galactosidase since the tissues were isolated from mouse embryos that carry n-lacZ targeted to the myf-5 gene, so that myf-5 expressing cells could be easily identified by either histochemical or immunocytochemical staining for beta-galactosidase. After 1 day in culture myogenic cells from the medial half expressed myf-5 but not MyoD, while myogenic cells from the lateral half expressed MyoD but not myf-5. By the next day in vitro, however, most myogenic cells expressed both gene products. These data suggest that the neural tube activates myogenesis in the medial half of Paraxial Mesoderm through a myf-5-dependent pathway, while the dorsal ectoderm activates myogenesis through a MyoD-dependent pathway. The possible developmental significance of these observations is discussed and a model of myogenic determination in mammals is proposed.
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myoblast differentiation during mammalian somitogenesis is dependent upon a community effect
Proceedings of the National Academy of Sciences of the United States of America, 1995Co-Authors: Giulio Cossu, E Vivarelli, Robert G. Kelly, S Di Donna, Margaret BuckinghamAbstract:Abstract The differentiation potential of early mammalian myogenic cells was tested under clonal culture conditions. Cells were isolated from Paraxial Mesoderm and limb buds of transgenic mouse embryos at 9.5 days after conception and grown in culture at clonal density either on collagen-coated dishes or on various feeder cell layers. The transgene used contained a reporter gene encoding beta-galactosidase with a nuclear localization signal under the control of regulatory sequences from the gene for fast myosin light chain 3, so that beta-galactosidase staining indicated the presence of differentiated muscle cells. After 5 days in culture, the number and size of beta-galactosidase-positive (beta-gal+) clones were recorded. Cells isolated from somites I-V (the last five somites to have formed) or from unsegmented Paraxial Mesoderm did not give rise to any beta-gal+ clones. Cells isolated from somites VI-X or from the forelimb bud gave rise to beta-gal+ clones, but only on feeder cells. Cells from somites XI or older gave rise to beta-gal+ clones independently of the substrate. However, when cells isolated from unsegmented Paraxial Mesoderm or somites I-V were cultured with nontransgenic cells from the trunk (including neural tube and notochord), differentiation occurred on condition that the cells were in a three-dimensional aggregate, even though their specific position in the somite had been lost. By culturing explants ranging in size from 1 to < 100 cells in the presence of an inhibitor of cell division, we determined that a minimal number of 30-40 cells is required for Mesodermal cells to differentiate.
