Meeting Abstract
We have applied CRISPR/Cas9, a recently developed gene editing technology, to study embryos of Nematostella vectensis. Nematostella is a member of the early-branching phylum Cnidaria, the sister group to Bilateria. Bilaterians form three embryonic germ layers—ectoderm, mesoderm, and endoderm—as a key step in early development. Since cnidarians form only two embryonic germ layers, ectoderm and endoderm, this taxon is important for understanding evolutionary origins of mesoderm. We therefore chose to use CRISPR/Cas9 in Nematostella to study the developmental gene brachyury, a highly conserved T-box transcription factor which plays important roles in specification and differentiation of mesoderm in bilaterians. We show that we can excise the brachyury gene from the genome with high efficiency, and that the resulting embryos fail to express brachyury RNA. While brachyury/Cas9 embryos appear to undergo normal gastrulation movements, the endoderm is disorganized, and fails to form a normal epithelium. Expression of several genes involved in gastrulation (bmp2/4, FoxA) is disrupted in the absence of brachyury. Markers of aboral development – opposite the normal site of brachyury expression – often show expanded expression domains, suggesting that there are interactions between the oral and aboral domains during axial patterning. These results demonstrate that CRISPR/Cas9 can be applied to study early development in F0 Nematostella embryos, and that in the absence of brachyury expression, endoderm development is disrupted.
