Andrea Munsterberg | University of East Anglia (original) (raw)
Papers by Andrea Munsterberg
Mechanisms of Development, Jul 1, 2017
Development, 2020
Understanding how complex organ systems are assembled from simple embryonic tissues is a major ch... more Understanding how complex organ systems are assembled from simple embryonic tissues is a major challenge. Across the animal kingdom a great diversity of visual organs are initiated by a 'master control gene' called Pax6, which is both necessary and sufficient for eye development. Yet precisely how Pax6 achieves this deeply homologous function is poorly understood. Using the chick as a model organism, we show that vertebrate Pax6 interacts with a pair of morphogen-coding genes, Tgfb2 and Fst, to form a putative Turing network, which we have computationally modelled. Computer simulations suggest that this gene network is sufficient to spontaneously polarise the developing retina, establishing the first organisational axis of the eye and prefiguring its further development. Our findings reveal how retinal self-organisation may be initiated independently of the highly ordered tissue interactions that help to assemble the eye in vivo. These results help to explain how stem cell aggregates spontaneously self-organise into functional eye-cups in vitro. We anticipate these findings will help to underpin retinal organoid technology, which holds much promise as a platform for disease modelling, drug development and regenerative therapies.
Methods in molecular biology, Sep 17, 2020
Avian (chick) embryos are an established and accessible model organism making them ideal for stud... more Avian (chick) embryos are an established and accessible model organism making them ideal for studying developmental processes. Chick embryos can be harvested from the egg and cultured allowing real-time observations and imaging. Here, we describe ex vivo culture and preparation of somite tissue followed by time-lapse multi-photon microscopy, image capture and processing. We applied this approach to perform live imaging of somites, the paired segments in vertebrate embryos that form in a regular sequence on either side of the neural tube, posteriorly from presomitic mesoderm (psm). Somites give rise to cell lineages of the musculoskeletal system in the trunk such as skeletal muscle, cartilage and tendon, as well as endothelial cells. Until recently it was not possible to observe the cellular dynamics underlying morphological transitions in live tissue, including in somites which undergo epithelial-to-mesenchymal transitions (EMT) during their differentiation. In addition to the experimental setup, we describe the analytical tools used for image processing.
bioRxiv (Cold Spring Harbor Laboratory), Jan 24, 2023
The mammalian body plan is shaped by rhythmic segmentation of mesoderm into somites, which are tr... more The mammalian body plan is shaped by rhythmic segmentation of mesoderm into somites, which are transient embryonic structures consisting of hundreds of cells that form down each side of the neural tube. We have systematically analysed the genome-wide transcriptional and chromatin dynamics occurring within nascent somites, from early inception of somitogenesis to the latest stages of body plan establishment. We created matched gene expression and open chromatin maps for the three leading pairs of somites at six time points during embryonic development. Here we show that the rate of somite differentiation accelerates as development progresses. We identified a conserved maturation programme followed by all somites after segmentation, but somites from more developed embryos concomitantly switch on differentiation programmes from derivative cell lineages soon after segmentation. Integrated analysis of the somitic transcriptional and chromatin activities revealed opposing regulatory modules controlling the onset of differentiation. We identified transcription factors expressed during early development that inhibit the activity of proteins required for commitment and differentiation of skeletal cell populations. Our results provide a powerful, high-resolution view of the molecular genetics underlying somitic development in mammals. .
Skeletal muscle stem cells (MuSC) are crucial for tissue homeostasis and repair after injury. Fol... more Skeletal muscle stem cells (MuSC) are crucial for tissue homeostasis and repair after injury. Following activation, they proliferate to generate differentiating myoblasts. A proportion of cells selfrenew, re-enter the MuSC niche under the basal lamina outside the myofiber and become quiescent. Quiescent MuSC have a primary cilium, which is disassembled upon cell cycle entry.Ex vivoexperiments suggest cilia are important for MuSC self-renewal, however, their role in muscle regenerationin vivoremains poorly understood. Talpid3 (TA3) is essential for primary cilia formation and Hedgehog (Hh) signalling. Here we use tamoxifen-inducible conditional deletion of TA3in MuSC (iSC-KO) and show that regeneration is impaired in response to cytotoxic injury. Repeat injury exacerbates the regeneration phenotype in TA3iSC-KOmice, indicating depletion of MuSCs. Single cell transcriptomics of MuSC progeny isolated from myofibers identifies components of several signalling pathways, which are deregul...
Understanding how complex organ systems are assembled from simple embryonic tissues is a major ch... more Understanding how complex organ systems are assembled from simple embryonic tissues is a major challenge. Across the animal kingdom a great diversity of visual organs are initiated by a ‘master control gene’ calledPax6, which is both necessary and sufficient for eye development1–6. Yet precisely howPax6achieves this deeply homologous function is poorly understood. Here we show that vertebratePax6interacts with a pair of morphogen-coding genes,Tgfb2andFst, to form a putative Turing network7, which we have computationally modelled. Computer simulations suggest that this gene network is sufficient to spontaneously polarise the developing retina, establishing the eye’s first organisational axis and prefiguring its further development. Our findings reveal how retinal self-organisation may be initiated independent of the highly ordered tissue interactions that help to assemble the eyein vivo. These results help to explain how stem cell aggregates spontaneously self-organise into functiona...
Mechanisms of Development, Jul 1, 2017
Communications Biology
Skeletal muscle stem cells (MuSC) are crucial for tissue homoeostasis and repair after injury. Fo... more Skeletal muscle stem cells (MuSC) are crucial for tissue homoeostasis and repair after injury. Following activation, they proliferate to generate differentiating myoblasts. A proportion of cells self-renew, re-enter the MuSC niche under the basal lamina outside the myofiber and become quiescent. Quiescent MuSC have a primary cilium, which is disassembled upon cell cycle entry. Ex vivo experiments suggest cilia are important for MuSC self-renewal, however, their requirement for muscle regeneration in vivo remains poorly understood. Talpid3 (TA3) is essential for primary cilia formation and Hedgehog (Hh) signalling. Here we use tamoxifen-inducible conditional deletion of TA3 in MuSC (iSC-KO) and show that regeneration is impaired in response to cytotoxic injury. Depletion of MuSC after regeneration suggests impaired self-renewal, also consistent with an exacerbated phenotype in TA3iSC-KO mice after repeat injury. Single cell transcriptomics of MuSC progeny isolated from myofibers iden...
Developmental Cell, Jul 1, 2023
Mechanisms of Development, Jul 1, 2017
Scientific Reports, Aug 23, 2018
Somites are paired embryonic segments that form in a regular sequence from unsegmented mesoderm d... more Somites are paired embryonic segments that form in a regular sequence from unsegmented mesoderm during vertebrate development. Although transient structures they are of fundamental importance as they generate cell lineages of the musculoskeletal system in the trunk such as cartilage, tendon, bone, endothelial cells and skeletal muscle. Surprisingly, very little is known about cellular dynamics underlying the morphological transitions during somite differentiation. Here, we address this by examining cellular rearrangements and morphogenesis in differentiating somites using live multiphoton imaging of transgenic chick embryos, where all cells express a membrane-bound GFP. We specifically focussed on the dynamic cellular changes in two principle regions within the somite, the medial and lateral domains, to investigate extensive morphological transformations. Furthermore, by using quantitative analysis and cell tracking, we capture for the first time a directed movement of dermomyotomal progenitor cells towards the rostro-medial domain of the dermomyotome, where skeletal muscle formation initiates. Embryonic morphogenesis involves dramatic tissue deformation and growth, which often occurs rapidly over short timescales. It is implicit that tissue deformations are driven by local cellular activities, including cell proliferation, changes in morphology and/or size, and cell rearrangements. However, it has been challenging to image, capture and quantify these processes in live tissues. Somites are transient, epithelial, near spherical structures that form during vertebrate development from the presomitic mesoderm (PSM) in a regular sequence and with a rostro-caudal progression 1. Somites can be staged based on morphological landmarks and age of development, using roman numerals 2. Newly formed somites consist of a ball of epithelial cells surrounding a central cavity, the somitocoel, which is filled with mesenchymal cells (stages I-III). As they differentiate, these paired body segments dissociate ventrally (from stage IV) and epithelial-to-mesenchymal transition (EMT) leads to formation of the sclerotome, the source of the axial skeleton. The dorsal somite remains epithelial and produces the dermomyotome and myotome, the source of all trunk and limb skeletal muscles 2,3. Signalling and genetic control of cell lineage specification is well characterised 4-6. For example, expression of the first myogenic marker, the transcription factor Myf5, is first detectable in the medial wall of epithelial somites 7. However, surprisingly very little is known about how individual cell dynamics and cellular rearrangements drive morphogenesis within the somite during its differentiation, for example during the emergence of the myotome. An improved and greater understanding of these processes may also benefit the derivation of musculoskeletal lineages from pluripotent stem cells 8. Along the rostro-caudal axis, each individual somite is flanked by neighbouring somites; other adjacent tissues on the medial, lateral, dorsal and ventral sides are the neural tube (future spinal cord), the intermediate and lateral plate mesoderm, the surface ectoderm and the endoderm respectively. Signalling molecules derived from many of these tissues govern the specification of somite cells towards particular fates 9-20. In addition, these flanking tissues
Development, Oct 1, 2000
The developmental signals that govern cell specification and differentiation in vertebrate somite... more The developmental signals that govern cell specification and differentiation in vertebrate somites are well understood. However, little is known about the downstream signalling pathways involved. We have shown previously that a combination of Shh protein and Wnt1 or Wnt3aexpressing fibroblasts is sufficient to activate skeletal muscle-specific gene expression in somite explants. Here, we have examined the molecular mechanisms by which the Wnt-mediated signal acts on myogenic precursor cells. We show that chick frizzled 1 (Fz1), β-catenin and Lef1 are expressed during somitogenesis. Lef1 and β-catenin transcripts become restricted to the developing myotome. Furthermore, β-catenin is expressed prior to the time at which MyoD transcripts can be detected. Expression of β-catenin mRNA is regulated by positive and negative signals derived from neural tube, notochord and lateral plate mesoderm. These signals include Bmp4, Shh and Wnt1/Wnt3a itself. In somite explants, Fz1, β-catenin and Lef1 are expressed prior to activation of myogenesis in response to Shh and Wnt signals. Thus, our data show that a combination of Shh and Wnt1 upregulates expression of Wnt pathway components in developing somites prior to myogenesis. Thus, Wnt1 could act through β-catenin on cells in the myotome.
The spalt proteins are encoded by a family of evolutionarily conserved genes found in species as ... more The spalt proteins are encoded by a family of evolutionarily conserved genes found in species as diverse as Drosophila, C. elegans and vertebrates. In humans, mutations in some of these genes are associated with several congenital disorders which underscores the importance of spalt gene function in embryonic development. Recent studies have begun to cast light on the functions of this family of proteins with increasing understanding of the developmental processes regulated and the molecular mechanisms used. Here we review what is currently known about the role of spalt genes in vertebrate development and human disease.
Seminars in Cell & Developmental Biology, Jul 1, 2022
A critical stage in the development of all vertebrate embryos is the generation of the body plan ... more A critical stage in the development of all vertebrate embryos is the generation of the body plan and its subsequent patterning and regionalisation along the main anterior-posterior axis. This includes the formation of the vertebral axial skeleton. Its organisation begins during early embryonic development with the periodic formation of paired blocks of mesoderm tissue called somites. Here, we review axial patterning of somites, with a focus on studies using amniote model systems-avian and mouse. We summarise the molecular and cellular mechanisms that generate paraxial mesoderm and review how the different anatomical regions of the vertebral column acquire their specific identity and thus shape the body plan. We also discuss the generation of organoids and embryo-like structures from embryonic stem cells, which provide insights regarding axis formation and promise to be useful for disease modelling.
Frontiers in Cell and Developmental Biology, 2017
*<p>Developmental stage indicated as Hamburger Hamilton (HH) stages <a href="http:/... more *<p>Developmental stage indicated as Hamburger Hamilton (HH) stages <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0051948#pone.0051948-Hamburger1" target="_blank">[30]</a>.</p>∧<p>Developmental stage indicated as Primitive Streak (PS) stages <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0051948#pone.0051948-LopezSanchez2" target="_blank">[31]</a>.</p><p>Table indicates the length of the primitive streak during development from stage HH3 (PS2) to HH3<sup>+</sup> (PS4) and the limits of region A and region B along the anterior-middle primitive streak. An eye piece reticule with 100 subdivisions (0 to100) was used to delineate region A and region B. The length of the primitive streak is shown in the number of subdivisions and in microns (µm). The limits of region A and region B for each developmental stage are shown as subdivisions. The primitive streak of all embryos used in the study was measured to determine developmental stage and a total of 348 embryos were labelled and analysed to determine the limits of each region. Labelling experiments were performed by either injecting vital fluorescent dyes at one to three subdivisions within each region or by grafting GFP fluorescent primitive streak tissue from most of region B of a donor embryo into the equivalent region B of a host wild type embryo.</p
MicroRNAs (miRNAs), short non-coding RNAs, which act post-transcriptionally to regulate gene expr... more MicroRNAs (miRNAs), short non-coding RNAs, which act post-transcriptionally to regulate gene expression, are of widespread significance during development and disease, including muscle disease. Advances in sequencing technology and bioinformatics led to the identification of a large number of miRNAs in vertebrates and other species, however, for many of these miRNAs specific roles have not yet been determined. LNA in situ hybridisation has revealed expression patterns of somite-enriched miRNAs, here we focus on characterising the functions of miR-128. We show that antagomir-mediated knock-down (KD) of miR-128 in developing chick somites has a negative impact on skeletal myogenesis. Computational analysis identified the transcription factor EYA4 as a candidate target consistent with the observation that miR-128 and EYA4 display similar expression profiles. Luciferase assays confirmed that miR-128 interacts with the EYA4 3’UTR. Furthermore, in vivo experiments suggest that EYA4 is reg...
Developmental Biology, 2018
Background: Studying microRNA networks during heart development is essential to obtain a better u... more Background: Studying microRNA networks during heart development is essential to obtain a better understanding of developmental defects and diseases associated with the heart and to identify novel opportunities for therapeutics. Here we highlight the advantages of chicken embryos as a vertebrate model for studying intermediate processes of heart development. Avians develop a four-chambered heart closely resembling human anatomy and they develop ex utero, which makes them easily accessible. Furthermore, embryos are available all year with a steady supply. Results: In this report we established a novel method for the knockdown of microRNA function by microinjecting AntagomiRs into the chicken heart in ovo. Our approach enables the targeted delivery of antagomirs into a locally restricted area and is not impacted by circulation. After further embryo development the successful miRNA knockdown was confirmed. Loss of function phenotypes can be evaluated rapidly, compared to more time-consuming genetic ablation experiments. The local application avoids potential systemic effects of microRNA knockdown, therefore allowing the assessment of impacts on heart development only. The method can be adjusted for different stages of chicken embryos (HH13-HH18) as well as for knockdown or targeted overexpression of coding genes. Conclusion: In conclusion our method allows targeted and locally restricted delivery of Antagomirs to the heart leading to successful knockdown of microRNA function. This method enables rapid phenotypic assessment, for example by gene expression analysis of multiple cardiac genes.
Mechanisms of Development, Jul 1, 2017
Development, 2020
Understanding how complex organ systems are assembled from simple embryonic tissues is a major ch... more Understanding how complex organ systems are assembled from simple embryonic tissues is a major challenge. Across the animal kingdom a great diversity of visual organs are initiated by a 'master control gene' called Pax6, which is both necessary and sufficient for eye development. Yet precisely how Pax6 achieves this deeply homologous function is poorly understood. Using the chick as a model organism, we show that vertebrate Pax6 interacts with a pair of morphogen-coding genes, Tgfb2 and Fst, to form a putative Turing network, which we have computationally modelled. Computer simulations suggest that this gene network is sufficient to spontaneously polarise the developing retina, establishing the first organisational axis of the eye and prefiguring its further development. Our findings reveal how retinal self-organisation may be initiated independently of the highly ordered tissue interactions that help to assemble the eye in vivo. These results help to explain how stem cell aggregates spontaneously self-organise into functional eye-cups in vitro. We anticipate these findings will help to underpin retinal organoid technology, which holds much promise as a platform for disease modelling, drug development and regenerative therapies.
Methods in molecular biology, Sep 17, 2020
Avian (chick) embryos are an established and accessible model organism making them ideal for stud... more Avian (chick) embryos are an established and accessible model organism making them ideal for studying developmental processes. Chick embryos can be harvested from the egg and cultured allowing real-time observations and imaging. Here, we describe ex vivo culture and preparation of somite tissue followed by time-lapse multi-photon microscopy, image capture and processing. We applied this approach to perform live imaging of somites, the paired segments in vertebrate embryos that form in a regular sequence on either side of the neural tube, posteriorly from presomitic mesoderm (psm). Somites give rise to cell lineages of the musculoskeletal system in the trunk such as skeletal muscle, cartilage and tendon, as well as endothelial cells. Until recently it was not possible to observe the cellular dynamics underlying morphological transitions in live tissue, including in somites which undergo epithelial-to-mesenchymal transitions (EMT) during their differentiation. In addition to the experimental setup, we describe the analytical tools used for image processing.
bioRxiv (Cold Spring Harbor Laboratory), Jan 24, 2023
The mammalian body plan is shaped by rhythmic segmentation of mesoderm into somites, which are tr... more The mammalian body plan is shaped by rhythmic segmentation of mesoderm into somites, which are transient embryonic structures consisting of hundreds of cells that form down each side of the neural tube. We have systematically analysed the genome-wide transcriptional and chromatin dynamics occurring within nascent somites, from early inception of somitogenesis to the latest stages of body plan establishment. We created matched gene expression and open chromatin maps for the three leading pairs of somites at six time points during embryonic development. Here we show that the rate of somite differentiation accelerates as development progresses. We identified a conserved maturation programme followed by all somites after segmentation, but somites from more developed embryos concomitantly switch on differentiation programmes from derivative cell lineages soon after segmentation. Integrated analysis of the somitic transcriptional and chromatin activities revealed opposing regulatory modules controlling the onset of differentiation. We identified transcription factors expressed during early development that inhibit the activity of proteins required for commitment and differentiation of skeletal cell populations. Our results provide a powerful, high-resolution view of the molecular genetics underlying somitic development in mammals. .
Skeletal muscle stem cells (MuSC) are crucial for tissue homeostasis and repair after injury. Fol... more Skeletal muscle stem cells (MuSC) are crucial for tissue homeostasis and repair after injury. Following activation, they proliferate to generate differentiating myoblasts. A proportion of cells selfrenew, re-enter the MuSC niche under the basal lamina outside the myofiber and become quiescent. Quiescent MuSC have a primary cilium, which is disassembled upon cell cycle entry.Ex vivoexperiments suggest cilia are important for MuSC self-renewal, however, their role in muscle regenerationin vivoremains poorly understood. Talpid3 (TA3) is essential for primary cilia formation and Hedgehog (Hh) signalling. Here we use tamoxifen-inducible conditional deletion of TA3in MuSC (iSC-KO) and show that regeneration is impaired in response to cytotoxic injury. Repeat injury exacerbates the regeneration phenotype in TA3iSC-KOmice, indicating depletion of MuSCs. Single cell transcriptomics of MuSC progeny isolated from myofibers identifies components of several signalling pathways, which are deregul...
Understanding how complex organ systems are assembled from simple embryonic tissues is a major ch... more Understanding how complex organ systems are assembled from simple embryonic tissues is a major challenge. Across the animal kingdom a great diversity of visual organs are initiated by a ‘master control gene’ calledPax6, which is both necessary and sufficient for eye development1–6. Yet precisely howPax6achieves this deeply homologous function is poorly understood. Here we show that vertebratePax6interacts with a pair of morphogen-coding genes,Tgfb2andFst, to form a putative Turing network7, which we have computationally modelled. Computer simulations suggest that this gene network is sufficient to spontaneously polarise the developing retina, establishing the eye’s first organisational axis and prefiguring its further development. Our findings reveal how retinal self-organisation may be initiated independent of the highly ordered tissue interactions that help to assemble the eyein vivo. These results help to explain how stem cell aggregates spontaneously self-organise into functiona...
Mechanisms of Development, Jul 1, 2017
Communications Biology
Skeletal muscle stem cells (MuSC) are crucial for tissue homoeostasis and repair after injury. Fo... more Skeletal muscle stem cells (MuSC) are crucial for tissue homoeostasis and repair after injury. Following activation, they proliferate to generate differentiating myoblasts. A proportion of cells self-renew, re-enter the MuSC niche under the basal lamina outside the myofiber and become quiescent. Quiescent MuSC have a primary cilium, which is disassembled upon cell cycle entry. Ex vivo experiments suggest cilia are important for MuSC self-renewal, however, their requirement for muscle regeneration in vivo remains poorly understood. Talpid3 (TA3) is essential for primary cilia formation and Hedgehog (Hh) signalling. Here we use tamoxifen-inducible conditional deletion of TA3 in MuSC (iSC-KO) and show that regeneration is impaired in response to cytotoxic injury. Depletion of MuSC after regeneration suggests impaired self-renewal, also consistent with an exacerbated phenotype in TA3iSC-KO mice after repeat injury. Single cell transcriptomics of MuSC progeny isolated from myofibers iden...
Developmental Cell, Jul 1, 2023
Mechanisms of Development, Jul 1, 2017
Scientific Reports, Aug 23, 2018
Somites are paired embryonic segments that form in a regular sequence from unsegmented mesoderm d... more Somites are paired embryonic segments that form in a regular sequence from unsegmented mesoderm during vertebrate development. Although transient structures they are of fundamental importance as they generate cell lineages of the musculoskeletal system in the trunk such as cartilage, tendon, bone, endothelial cells and skeletal muscle. Surprisingly, very little is known about cellular dynamics underlying the morphological transitions during somite differentiation. Here, we address this by examining cellular rearrangements and morphogenesis in differentiating somites using live multiphoton imaging of transgenic chick embryos, where all cells express a membrane-bound GFP. We specifically focussed on the dynamic cellular changes in two principle regions within the somite, the medial and lateral domains, to investigate extensive morphological transformations. Furthermore, by using quantitative analysis and cell tracking, we capture for the first time a directed movement of dermomyotomal progenitor cells towards the rostro-medial domain of the dermomyotome, where skeletal muscle formation initiates. Embryonic morphogenesis involves dramatic tissue deformation and growth, which often occurs rapidly over short timescales. It is implicit that tissue deformations are driven by local cellular activities, including cell proliferation, changes in morphology and/or size, and cell rearrangements. However, it has been challenging to image, capture and quantify these processes in live tissues. Somites are transient, epithelial, near spherical structures that form during vertebrate development from the presomitic mesoderm (PSM) in a regular sequence and with a rostro-caudal progression 1. Somites can be staged based on morphological landmarks and age of development, using roman numerals 2. Newly formed somites consist of a ball of epithelial cells surrounding a central cavity, the somitocoel, which is filled with mesenchymal cells (stages I-III). As they differentiate, these paired body segments dissociate ventrally (from stage IV) and epithelial-to-mesenchymal transition (EMT) leads to formation of the sclerotome, the source of the axial skeleton. The dorsal somite remains epithelial and produces the dermomyotome and myotome, the source of all trunk and limb skeletal muscles 2,3. Signalling and genetic control of cell lineage specification is well characterised 4-6. For example, expression of the first myogenic marker, the transcription factor Myf5, is first detectable in the medial wall of epithelial somites 7. However, surprisingly very little is known about how individual cell dynamics and cellular rearrangements drive morphogenesis within the somite during its differentiation, for example during the emergence of the myotome. An improved and greater understanding of these processes may also benefit the derivation of musculoskeletal lineages from pluripotent stem cells 8. Along the rostro-caudal axis, each individual somite is flanked by neighbouring somites; other adjacent tissues on the medial, lateral, dorsal and ventral sides are the neural tube (future spinal cord), the intermediate and lateral plate mesoderm, the surface ectoderm and the endoderm respectively. Signalling molecules derived from many of these tissues govern the specification of somite cells towards particular fates 9-20. In addition, these flanking tissues
Development, Oct 1, 2000
The developmental signals that govern cell specification and differentiation in vertebrate somite... more The developmental signals that govern cell specification and differentiation in vertebrate somites are well understood. However, little is known about the downstream signalling pathways involved. We have shown previously that a combination of Shh protein and Wnt1 or Wnt3aexpressing fibroblasts is sufficient to activate skeletal muscle-specific gene expression in somite explants. Here, we have examined the molecular mechanisms by which the Wnt-mediated signal acts on myogenic precursor cells. We show that chick frizzled 1 (Fz1), β-catenin and Lef1 are expressed during somitogenesis. Lef1 and β-catenin transcripts become restricted to the developing myotome. Furthermore, β-catenin is expressed prior to the time at which MyoD transcripts can be detected. Expression of β-catenin mRNA is regulated by positive and negative signals derived from neural tube, notochord and lateral plate mesoderm. These signals include Bmp4, Shh and Wnt1/Wnt3a itself. In somite explants, Fz1, β-catenin and Lef1 are expressed prior to activation of myogenesis in response to Shh and Wnt signals. Thus, our data show that a combination of Shh and Wnt1 upregulates expression of Wnt pathway components in developing somites prior to myogenesis. Thus, Wnt1 could act through β-catenin on cells in the myotome.
The spalt proteins are encoded by a family of evolutionarily conserved genes found in species as ... more The spalt proteins are encoded by a family of evolutionarily conserved genes found in species as diverse as Drosophila, C. elegans and vertebrates. In humans, mutations in some of these genes are associated with several congenital disorders which underscores the importance of spalt gene function in embryonic development. Recent studies have begun to cast light on the functions of this family of proteins with increasing understanding of the developmental processes regulated and the molecular mechanisms used. Here we review what is currently known about the role of spalt genes in vertebrate development and human disease.
Seminars in Cell & Developmental Biology, Jul 1, 2022
A critical stage in the development of all vertebrate embryos is the generation of the body plan ... more A critical stage in the development of all vertebrate embryos is the generation of the body plan and its subsequent patterning and regionalisation along the main anterior-posterior axis. This includes the formation of the vertebral axial skeleton. Its organisation begins during early embryonic development with the periodic formation of paired blocks of mesoderm tissue called somites. Here, we review axial patterning of somites, with a focus on studies using amniote model systems-avian and mouse. We summarise the molecular and cellular mechanisms that generate paraxial mesoderm and review how the different anatomical regions of the vertebral column acquire their specific identity and thus shape the body plan. We also discuss the generation of organoids and embryo-like structures from embryonic stem cells, which provide insights regarding axis formation and promise to be useful for disease modelling.
Frontiers in Cell and Developmental Biology, 2017
*<p>Developmental stage indicated as Hamburger Hamilton (HH) stages <a href="http:/... more *<p>Developmental stage indicated as Hamburger Hamilton (HH) stages <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0051948#pone.0051948-Hamburger1" target="_blank">[30]</a>.</p>∧<p>Developmental stage indicated as Primitive Streak (PS) stages <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0051948#pone.0051948-LopezSanchez2" target="_blank">[31]</a>.</p><p>Table indicates the length of the primitive streak during development from stage HH3 (PS2) to HH3<sup>+</sup> (PS4) and the limits of region A and region B along the anterior-middle primitive streak. An eye piece reticule with 100 subdivisions (0 to100) was used to delineate region A and region B. The length of the primitive streak is shown in the number of subdivisions and in microns (µm). The limits of region A and region B for each developmental stage are shown as subdivisions. The primitive streak of all embryos used in the study was measured to determine developmental stage and a total of 348 embryos were labelled and analysed to determine the limits of each region. Labelling experiments were performed by either injecting vital fluorescent dyes at one to three subdivisions within each region or by grafting GFP fluorescent primitive streak tissue from most of region B of a donor embryo into the equivalent region B of a host wild type embryo.</p
MicroRNAs (miRNAs), short non-coding RNAs, which act post-transcriptionally to regulate gene expr... more MicroRNAs (miRNAs), short non-coding RNAs, which act post-transcriptionally to regulate gene expression, are of widespread significance during development and disease, including muscle disease. Advances in sequencing technology and bioinformatics led to the identification of a large number of miRNAs in vertebrates and other species, however, for many of these miRNAs specific roles have not yet been determined. LNA in situ hybridisation has revealed expression patterns of somite-enriched miRNAs, here we focus on characterising the functions of miR-128. We show that antagomir-mediated knock-down (KD) of miR-128 in developing chick somites has a negative impact on skeletal myogenesis. Computational analysis identified the transcription factor EYA4 as a candidate target consistent with the observation that miR-128 and EYA4 display similar expression profiles. Luciferase assays confirmed that miR-128 interacts with the EYA4 3’UTR. Furthermore, in vivo experiments suggest that EYA4 is reg...
Developmental Biology, 2018
Background: Studying microRNA networks during heart development is essential to obtain a better u... more Background: Studying microRNA networks during heart development is essential to obtain a better understanding of developmental defects and diseases associated with the heart and to identify novel opportunities for therapeutics. Here we highlight the advantages of chicken embryos as a vertebrate model for studying intermediate processes of heart development. Avians develop a four-chambered heart closely resembling human anatomy and they develop ex utero, which makes them easily accessible. Furthermore, embryos are available all year with a steady supply. Results: In this report we established a novel method for the knockdown of microRNA function by microinjecting AntagomiRs into the chicken heart in ovo. Our approach enables the targeted delivery of antagomirs into a locally restricted area and is not impacted by circulation. After further embryo development the successful miRNA knockdown was confirmed. Loss of function phenotypes can be evaluated rapidly, compared to more time-consuming genetic ablation experiments. The local application avoids potential systemic effects of microRNA knockdown, therefore allowing the assessment of impacts on heart development only. The method can be adjusted for different stages of chicken embryos (HH13-HH18) as well as for knockdown or targeted overexpression of coding genes. Conclusion: In conclusion our method allows targeted and locally restricted delivery of Antagomirs to the heart leading to successful knockdown of microRNA function. This method enables rapid phenotypic assessment, for example by gene expression analysis of multiple cardiac genes.