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1.
Open Biol ; 10(2): 190299, 2020 02.
Article in English | MEDLINE | ID: mdl-32102607

ABSTRACT

The early stages of development of the chick embryo, leading to primitive streak formation (the start of gastrulation), have received renewed attention recently, especially for studies of the mechanisms of large-scale cell movements and those that position the primitive streak in the radial blastodisc. Over the long history of chick embryology, the terminology used to define different regions has been changing, making it difficult to relate studies to each other. To resolve this objectively requires precise definitions of the regions based on anatomical and functional criteria, along with a systematic molecular map that can be compared directly to the functional anatomy. Here, we undertake these tasks. We describe the characteristic cell morphologies (using scanning electron microscopy and immunocytochemistry for cell polarity markers) in different regions and at successive stages. RNAseq was performed for 12 regions of the blastodisc, from which a set of putative regional markers was selected. These were studied in detail by in situ hybridization. Together this provides a comprehensive resource allowing the community to define the regions unambiguously and objectively. In addition to helping with future experimental design and interpretation, this resource will also be useful for evolutionary comparisons between different vertebrate species.


Subject(s)
Biomarkers/metabolism , Gene Expression Profiling/veterinary , Gene Regulatory Networks , Primitive Streak/anatomy & histology , Animals , Cell Polarity , Chick Embryo , Gene Expression Regulation, Developmental , Immunohistochemistry , In Situ Hybridization , Microscopy, Atomic Force , Primitive Streak/growth & development , Primitive Streak/metabolism , Sequence Analysis, RNA
2.
Development ; 147(3)2020 02 04.
Article in English | MEDLINE | ID: mdl-31964776

ABSTRACT

Directional cell intercalations of epithelial cells during gastrulation has, in several organisms, been shown to be associated with a planar cell polarity in the organisation of the actin-myosin cytoskeleton and is postulated to reflect directional tension that drives oriented cell intercalations. We have characterised and applied a recently introduced non-destructive optical manipulation technique to measure the tension in individual epithelial cell junctions of cells in various locations and orientations in the epiblast of chick embryos in the early stages of primitive streak formation. Junctional tension of mesendoderm precursors in the epiblast is higher in junctions oriented in the direction of intercalation than in junctions oriented perpendicular to the direction of intercalation and higher than in junctions of other cells in the epiblast. The kinetic data fit best with a simple viscoelastic Maxwell model, and we find that junctional tension, and to a lesser extent viscoelastic relaxation time, are dependent on myosin activity.


Subject(s)
Epithelial Cells/metabolism , Gastrulation/physiology , Intercellular Junctions/metabolism , Optical Tweezers , Primitive Streak/growth & development , Animals , Animals, Genetically Modified , Cell Movement/physiology , Cell Polarity/physiology , Chick Embryo , Gastrula/metabolism , Germ Layers/metabolism , Green Fluorescent Proteins/genetics , Green Fluorescent Proteins/metabolism , Hydrocarbons, Chlorinated/pharmacology , Microscopy, Fluorescence/methods , Myosin Type I/antagonists & inhibitors , Myosin Type I/metabolism , Myosin Type II/antagonists & inhibitors , Myosin Type II/metabolism , Pyrroles/pharmacology , Signal Transduction/physiology
3.
PLoS Genet ; 9(11): e1003957, 2013 Nov.
Article in English | MEDLINE | ID: mdl-24244203

ABSTRACT

Oct4 is a widely recognized pluripotency factor as it maintains Embryonic Stem (ES) cells in a pluripotent state, and, in vivo, prevents the inner cell mass (ICM) in murine embryos from differentiating into trophectoderm. However, its function in somatic tissue after this developmental stage is not well characterized. Using a tamoxifen-inducible Cre recombinase and floxed alleles of Oct4, we investigated the effect of depleting Oct4 in mouse embryos between the pre-streak and headfold stages, ~E6.0-E8.0, when Oct4 is found in dynamic patterns throughout the embryonic compartment of the mouse egg cylinder. We found that depletion of Oct4 ~E7.5 resulted in a severe phenotype, comprised of craniorachischisis, random heart tube orientation, failed turning, defective somitogenesis and posterior truncation. Unlike in ES cells, depletion of the pluripotency factors Sox2 and Oct4 after E7.0 does not phenocopy, suggesting that ~E7.5 Oct4 is required within a network that is altered relative to the pluripotency network. Oct4 is not required in extraembryonic tissue for these processes, but is required to maintain cell viability in the embryo and normal proliferation within the primitive streak. Impaired expansion of the primitive streak occurs coincident with Oct4 depletion ∼E7.5 and precedes deficient convergent extension which contributes to several aspects of the phenotype.


Subject(s)
Cell Differentiation/genetics , Embryonic Stem Cells/metabolism , Octamer Transcription Factor-3/metabolism , Pluripotent Stem Cells/metabolism , Animals , Cell Lineage , Cell Proliferation , Embryonic Development , Embryonic Stem Cells/cytology , Gene Expression Regulation, Developmental , Mice , Neural Tube Defects/etiology , Neural Tube Defects/genetics , Neural Tube Defects/pathology , Octamer Transcription Factor-3/antagonists & inhibitors , Octamer Transcription Factor-3/genetics , Pluripotent Stem Cells/cytology , Primitive Streak/growth & development , Primitive Streak/metabolism , SOXB1 Transcription Factors/antagonists & inhibitors , SOXB1 Transcription Factors/genetics , SOXB1 Transcription Factors/metabolism
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