75 research outputs found
Gene Regulatory Network Interactions in Sea Urchin Endomesoderm Induction
A major goal of contemporary studies of embryonic development is to understand large sets of regulatory changes that accompany the phenomenon of embryonic induction. The highly resolved sea urchin pregastrular endomesodermāgene regulatory network (EM-GRN) provides a unique framework to study the global regulatory interactions underlying endomesoderm induction. Vegetal micromeres of the sea urchin embryo constitute a classic endomesoderm signaling center, whose potential to induce archenteron formation from presumptive ectoderm was demonstrated almost a century ago. In this work, we ectopically activate the primary mesenchyme cellāGRN (PMC-GRN) that operates in micromere progeny by misexpressing the micromere determinant Pmar1 and identify the responding EM-GRN that is induced in animal blastomeres. Using localized loss-of -function analyses in conjunction with expression of endo16, the molecular definition of micromere-dependent endomesoderm specification, we show that the TGFĪ² cytokine, ActivinB, is an essential component of this induction in blastomeres that emit this signal, as well as in cells that respond to it. We report that normal pregastrular endomesoderm specification requires activation of the Pmar1-inducible subset of the EM-GRN by the same cytokine, strongly suggesting that early micromere-mediated endomesoderm specification, which regulates timely gastrulation in the sea urchin embryo, is also ActivinB dependent. This study unexpectedly uncovers the existence of an additional uncharacterized micromere signal to endomesoderm progenitors, significantly revising existing models. In one of the first network-level characterizations of an intercellular inductive phenomenon, we describe an important in vivo model of the requirement of ActivinB signaling in the earliest steps of embryonic endomesoderm progenitor specification
An investigation in the correlation between Ayurvedic body-constitution and food-taste preference
Integration of Canonical and Noncanonical Wnt Signaling Pathways Patterns the Neuroectoderm Along the AnteriorāPosterior Axis of Sea Urchin Embryos
<div><p>Patterning the neuroectoderm along the anteriorāposterior (AP) axis is a critical event in the early development of deuterostome embryos. However, the mechanisms that regulate the specification and patterning of the neuroectoderm are incompletely understood. Remarkably, the anterior neuroectoderm (ANE) of the deuterostome sea urchin embryo expresses many of the same transcription factors and secreted modulators of Wnt signaling, as does the early vertebrate ANE (forebrain/eye field). Moreover, as is the case in vertebrate embryos, confining the ANE to the anterior end of the embryo requires a Wnt/Ī²-catenin-dependent signaling mechanism. Here we use morpholino- or dominant negative-mediated interference to demonstrate that the early sea urchin embryo integrates information not only from Wnt/Ī²-catenin but also from Wnt/Fzl5/8-JNK and Fzl1/2/7-PKC pathways to provide precise spatiotemporal control of neuroectoderm patterning along its AP axis. Together, through the Wnt1 and Wnt8 ligands, they orchestrate a progressive posterior-to-anterior wave of re-specification that restricts the initial, ubiquitous, maternally specified, ANE regulatory state to the most anterior blastomeres. There, the Wnt receptor antagonist, Dkk1, protects this state through a negative feedback mechanism. Because these different Wnt pathways converge on the same cell fate specification process, our data suggest they may function as integrated components of an interactive Wnt signaling network. Our findings provide strong support for the idea that the sea urchin ANE regulatory state and the mechanisms that position and define its borders represent an ancient regulatory patterning system that was present in the common echinoderm/vertebrate ancestor.</p> </div
Sea Urchin FGFR Muscle-Specific Expression: Posttranscriptional Regulation in Embryos and Adults
AbstractWe have shown previously byin situhybridization that a gene encoding a fibroblast growth factor receptor (SpFGFR) is transcribed in many cell types during the initial phases of sea urchin embryogenesis (Strongylocentrotus purpuratus) (McCoonet al., J. Biol. Chem. 271,20119ā20195, 1996). Here we demonstrate by immunostaining with affinity-purified antibody that SpFGFR protein is detectable only in muscle cells of the embryo and appears at a time suggesting that its function is not in commitment to a muscle fate, but instead may be required to support the proliferation, migration, and/or differentiation of myoblasts. Surprisingly, we find thatSpFGFRtranscripts are enriched in embryo nuclei, suggesting that lack of processing and/or cytoplasmic transport in nonmuscle cells is at least part of the posttranscriptional regulatory mechanism. Western blots show that SpFGFR is also specifically expressed in adult lantern muscle, but is not detectable in other smooth muscle-containing tissues, including tube foot and intestine, or in coelomocytes, despite the presence ofSpFGFRtranscripts at similar concentrations in all these tissues. We conclude that in both embryos and adults, muscle-specific SpFGF receptor synthesis is controlled primarily at a posttranscriptional level. We show by RNase protection assays that transcripts encoding the IgS variant of the ligand binding domain of the receptor, previously shown to be enriched in embryo endomesoderm fractions, are the predominant, if not exclusive,SpFGFRtranscripts in lantern muscle. Together, these results suggest that only a minority ofSpFGFRtranscripts are processed, exported, and translated in both adult and embryonic muscle cells and these contain predominantly, if not exclusively, IgS ligand binding domain sequences
Preventing Wnt/Ī²-catenin signaling allows activation of presumptive ANE specification throughout the embryo.
<p>(A) Diagram showing the regulatory factors shared between the sea urchin ANE and the vertebrate forebrain/eye field. Both territories are restricted by mechanisms dependent on posterior Wnt signaling. (B, D, F, H) <i>foxq2</i> expression in glycerol control (B, D) or Axin mRNA-injected embryos (F, H) at the 32-cell stage (B, F) and late blastula stage (D, H). <i>six3</i> expression in glycerol control (C) and TCF-Eng mRNA-injected early blastulae (G). (E) A normal 3.5-d pluteus stage embryo. (I) A 3.5-d embryo misexpressing Axin mRNA. White box outlines the ANE. Serotonergic neurons (green), DAPI (nuclei, blue). (J) Schematic showing the stages of ANE restriction in 32-cell stage (6 hpf) (a), early blastula stage (7ā15 hpf) (b), and mesenchyme blastula stage (24 hpf) embryos (c). ANE (blue), nĪ²-catenin (orange nuclei at 32-cell stage), endomesoderm territory (orange at blastula stages), posterior ectoderm (gray at blastula stages), and unknown restriction mechanism activated by posterior nĪ²-catenin (orange arrows).</p
Wnt1 and Wnt8 signaling are necessary for Fzl5/8-JNK-mediated ANE restriction.
<p>(A) Three-color in situ hybridization for <i>wnt1</i> (red), <i>wnt8</i> (green), and <i>foxq2</i> (magenta) transcripts during ANE restriction. <i>wnt1 mRNA</i> appears yellow when overlapping with <i>wnt8 mRNA</i>. (B) The <i>foxq2</i> expression domain is not restricted in embryos injected with a Wnt1 (b) or Wnt8 (c) morpholino. (C) <i>foxq2</i> expression is completely eliminated in embryos injected with either Wnt1 (b) or Wnt8 (d) mRNA. The Wnt1- and Wnt8-mediated inhibition of <i>foxq2</i> expression requires functional Fzl5/8 (c, e); the Wnt8-mediated inhibition of <i>foxq2</i> expression requires JNK activity (f). MO, morpholino.</p
Fzl1/2/7 signaling and PKC activity are necessary for ANE specification.
<p>(A) <i>foxq2</i> expression at stages from 32-cell to hatched blastula, as well as <i>fzl5/8</i> and <i>six3</i> expression at mesenchyme blastula stage in control embryos (Aaāf), Fzl1/2/7 morpholino-injected embryos (Ahām), and embryos treated with PKC inhibitor (Aoāt). (Ag, n, u) Neurons in control, Fzl1/2/7 morpholino-injected, and PKC inhibitor-treated 96-hpf pluteus larvae. White boxes outline the ANE. Serotonergic neurons (green), DAPI (nuclei, blue), synaptotagminB/1e11 (pan-neural, magenta). (B) Western blot showing that phosphorylation of PKC is blocked to similar extents with a PKC inhibitor and Fzl1/2/7 MO. (C, D) qPCR measurements from three different cultures of embryos showing that the early ANE regulatory genes are down-regulated in the absence of Fzl1/2/7 signaling at the 60-cell stage (C) and late blastula stages (D). The <i>y</i>-axis shows the fold change in gene expression level in Fzl1/2/7 morpholino-containing embryos relative to controls. The dotted line marks a 3-fold change in expression. <i>nodal</i> expression is used as an internal control.</p
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