Genomics and expression profiles of the Hedgehog and Notch signaling pathways in sea urchin development

Abstract

The Hedgehog (Hh) and Notch signal transduction pathways control a variety of developmental processes including cell fate choice, differentiation, proliferation, patterning and boundary formation. Because many components of these pathways are conserved, it was predicted and confirmed that pathway components are largely intact in the sea urchin genome. Spatial and temporal location of these pathways in the embryo, and their function in development offer added insight into their mechanistic contributions. Accordingly, all major components of both pathways were identified and annotated in the sea urchin Strongylocentrotus purpuratus genome and the embryonic expression of key components was explored. Relationships of the pathway components, and modifiers predicted from the annotation of S. purpuratus, were compared against cnidarians, arthropods, urochordates, and vertebrates. These analyses support the prediction that the pathways are highly conserved through metazoan evolution. Further, the location of these two pathways appears to be conserved among deuterostomes, and in the case of Notch at least, display similar capacities in endomesoderm gene regulatory networks. RNA expression profiles by quantitative PCR and RNA in situ hybridization reveal that Hedgehog is produced by the endoderm beginning just prior to invagination, and signals to the secondary mesenchyme-derived tissues at least until the pluteus larva stage. RNA in situ hybridization of Notch pathway members confirms that Notch functions sequentially in the vegetal-most secondary mesenchyme cells and later in the endoderm. Functional analyses in future studies will embed these pathways into the growing knowledge of gene regulatory networks that govern early specification and morphogenesis.

Gene regulatory network subcircuit controlling a dynamic spatial pattern of signaling in the sea urchin embryo

“…Following the gradual disappearance of maternal Notch protein (9), the notch gene is zygotically expressed like hesC in the Veg2 endoderm ring (10), where it is responsible for transducing the NSM Delta signal. Mutation of a cis-regulatory Blimp1 site in the Notch gene (Fig.…”

“…The Notch receptor is initially expressed maternally and is globally distributed, although by early blastula stage it is regionally localized (9,10). Before this, the location of intracellular Notch signaling is determined entirely by spatial control of transcriptional expression of the delta gene, encoding a Notch ligand.…”

The dynamic gene expression patterns of transcription factors constituting the sea urchin aboral ectoderm gene regulatory network

“…For Sp-Klf7, because of its low expression level, a dinitrophenol (DNP)-labeled probe was made using the LabelIT DNP Labeling Kit (Mirus, Madison, WI). In situ hybridization was performed as previously described (Walton et al, 2006;Yu and Holland, 2009) with the following modifications. Embryos were fixed in 4% paraformaldehyde in FSW with 10 mM 4-(2-Hydroxyethyl)-1-piperazinepropanesulfonic acid (EPPS; Sigma E1894, St. Louis, MO), pH 8, at 4 C overnight.…”

Logic of gene regulatory networks

“…At 7 th cleavage the skeletogenic cells at the center of the vegetal plate express the Delta ligand and present it on their surface. It is received through the Notch receptor by the adjacent ring of cells [10,11,12 • ], which are in consequence specified as mesoderm. When the Delta ligand is bound, the intracellular domain of Notch (N ic ) is cleaved from the receptor.…”

“…Reception of the Delta signal by mesodermal cells kicks off the subsequent specification events. The Notch receptor, through which the Delta signal is received, is expressed widely in the early embryo [10,11]. However, the Delta ligand stays attached to the cells that express it, effectively limiting its range to its immediate neighbors [7].…”