Nuclear β‐catenin‐dependent Wnt8 signaling in vegetal cells of the early sea urchin embryo regulates gastrulation and differentiation of endoderm and mesodermal cell lineages

Abstract

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.

Global regulatory logic for specification of an embryonic cell lineage

“…This experiment, which has been repeated with molecular markers (14), means that micromeres execute signaling functions that have inductive effects on adjacent cells. We now know that between fourth and ninth cleavage (5-12 h after fertilization) the micromeres in fact express three different intercellular signaling ligands: (i) Wnt8, which enhances nuclearization of ␤-catenin in recipient cells, including themselves (15)(16)(17); (ii) a still undefined ''early signal (ES),'' which is received by adjacent cells in the fourth-to sixth-cleavage interval and is required by them for the normal process of endomesoderm specification (18); and (iii) from seventh cleavage, the Notch ligand Delta. Reception of this signal causes the ring of cells then immediately adjacent to the micromere lineage to assume mesodermal fate (19,20).…”

An evolutionary constraint: Strongly disfavored class of change in DNA sequence during divergence of cis-regulatory modules

“…The five cis-regulatory modules derive from three genes that encode transcription factors [namely, gatae (22), brachyury, and otx (23)] and two genes that encode signaling ligands [namely, delta (24) and wnt8 (25)]. The endo16 gene, also included in the analysis, encodes a terminal differentiation protein of the endoderm (26).…”

Controlled levels of canonical Wnt signaling are required for neural crest migration

“…Depending which pathway becomes activated Wnts are subdivided into two classes; canonical and non-canonical according to the ability to induce accumulation of β-catenin in the nucleus. Signaling through some Wnts such as; Wnt1 (Hinck et al, 1994;Young et al, 1998;Shimizu et al, 1997), Wnt3a (Shimizu et al, 1997) and Wnt8 (Wikramanayake et al, 2004;Kohn and Moon, 2005) has been shown to correlate with accumulation of β-catenin in the nucleus and transcriptional changes, while signaling trough others like Wnt5a (Shimizu et al, 1997;Kikuchi et al, 2012) and Wnt11 (Kohn and Moon, 2005;Tada and Smith, 2000;Pandur et al, 2002) is β-catenin-independent and involves regulation of cytoskeleton or calcium release. All the metazoan species express Wnt genes and all of those genes encode secreted proteins based on their amino acid structure and biochemical characterization (Coudreuse and Korswagen, 2007;Smolich et al, 1993).…”