Nuclear beta-catenin-dependent Wnt8 signaling in vegetal cells of the early sea urchin embryo regulates gastrulation and differentiation of endoderm and mesodermal cell lineages.
The entry of beta-catenin into vegetal cell nuclei beginning at the 16-cell stage is one of the earliest known molecular asymmetries seen along the animal-vegetal axis in the sea urchin embryo. Nuclear beta-catenin activates a vegetal signaling cascade that mediates micromere specification and specification of the endomesoderm in the remaining cells of the vegetal half of the embryo. Only a few potential target genes of nuclear beta-catenin have been functionally analyzed in the sea urchin embryo. Here, we show that SpWnt8, a Wnt8 homolog from Strongylocentrotus purpuratus, is zygotically activated specifically in 16-cell-stage micromeres in a nuclear beta-catenin-dependent manner, and its expression remains restricted to the micromeres until the 60-cell stage. At the late 60-cell stage nuclear beta-catenin-dependent SpWnt8 expression expands to the veg2 cell tier. SpWnt8 is the only signaling molecule thus far identified with expression localized to the 16-60-cell stage micromeres and the veg2 tier. Overexpression of SpWnt8 by mRNA microinjection produced embryos with multiple invagination sites and showed that, consistent with its localization, SpWnt8 is a strong inducer of endoderm. Blocking SpWnt8 function using SpWnt8 morpholino antisense oligonucleotides produced embryos that formed micromeres that could transmit the early endomesoderm-inducing signal, but these cells failed to differentiate as primary mesenchyme cells. SpWnt8-morpholino embryos also did not form endoderm, or secondary mesenchyme-derived pigment and muscle cells, indicating a role for SpWnt8 in gastrulation and in the differentiation of endomesodermal lineages. These results establish SpWnt8 as a critical component of the endomesoderm regulatory network in the sea urchin embryo.
Department of Zoology, University of Hawaii at Manoa, Honolulu, Hawaii 96822, USA. athula@hawaii.edu
Athula H
AH
Wikramanayake
Robert
R
Peterson
Jing
J
Chen
Ling
L
Huang
Joanna M
JM
Bince
David R
DR
McClay
William H
WH
Klein
D000818:Animals; D015152:Blotting, Northern; D002454:Cell Differentiation / Q000502:physiology; D019070:Cell Lineage / Q000502:physiology; D002467:Cell Nucleus / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D017931:DNA Primers; D004707:Endoderm / Q000502:physiology; D005455:Fluorescent Antibody Technique; D005775:Gastrula / Q000378:metabolism; D020869:Gene Expression Profiling; D018507:Gene Expression Regulation, Developmental; D017403:In Situ Hybridization; D008648:Mesoderm / Q000502:physiology; D008845:Microinjections; D016376:Oligonucleotides, Antisense; D010957:Plasmids / Q000235:genetics; D011506:Proteins / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012617:Sea Urchins / Q000196:embryology* / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*; D015534:Trans-Activators / Q000378:metabolism*; D051153:Wnt Proteins; D029961:Zebrafish Proteins; D051176:beta Catenin
D003598:Cytoskeletal Proteins; D017931:DNA Primers; D016376:Oligonucleotides, Antisense; D011506:Proteins; D015534:Trans-Activators; D051153:Wnt Proteins; D029961:Zebrafish Proteins; D051176:beta Catenin; C094521:wnt8a protein, zebrafish