This proposal seeks to understand how genomes (the total DNA sequences of a creature) evolve to generate diversity in animal shapes. Prior to the analysis of genomes of many animals, it was assumed that animals developed differently because they had different protein-coding genes. It is now realized that even very different animals have similar sets of genes, and it is not the genes themselves, but how they are used during embryonic development, that lead to differences. How those genes are used differently during development is poorly understood, however, and is important for understanding the types of changes in animal shape that can occur during evolution. This proposal takes advantage of a clear example of a set of genes that have evolved a new function, i.e. to make a larval skeleton in the sea urchin, while most echinoderms (the group that includes the sea urchins) make a skeleton only in the adult animal. What changes in gene regulation led to this basic new ability? Many community resources will be generated including DNA sequences that will be shared on publically available databases such as NCBI and Echinobase. This proposal will also train postdoctoral researchers, graduate students, and undergraduates, including those from groups that are traditionally underrepresented in science. The broader public will also be engaged in this research through a series of public lectures in Carnegie Mellon's Osher Lifelong Learning Institute and through outreach programs with local K-12 public schools.
The gene regulatory network (GRN) for the development of the sea urchin larval skeleton is very well known. Other out-group echinoderm larvae have no or very reduced larval skeletons, yet all echinoderms make a skeleton after metamorphosis. Prior evidence suggests that the sea urchin larval skeleton was co-opted from the GRN for the formation of this adult skeleton. The first objective of this proposal is to determine the set of genes that have been co-opted from the adult skeletogenic program, and are differently expressed in the sea star mesoderm, which does not form a skeleton. Differential RNA-Seq will be used to screen for potential genes in sea urchin, sea cucumbers, and a sea star. Whole-mount in situ hybridization will be used to determine their spatial location. The next objective is to understand how cis-regulatory modules (CRMs) can function in the new larval context and to understand the function of the subcircuits of the co-opted genes in larval sea urchins. ChIP-Seq will identify CRMs and their associated genes. Functional dissection of CRMs and perturbation of identified target genes that are shared targets will determine how these CRMs can function in both the original and new circuit, and also the types of trans environments, and subcircuit topologies that permit cooption. This work will contribute to an understanding of the plasticity and evolvability of GRNs.