One of the most fundamental and critical functions of embryological development is the control and regulation of differential genes and gene networks. The study of the gene networks involved in development is a mechanism for understanding the developmental process at its most basic level. An evolutionary change in a morphological feature or features must depend on a reorganization or co-option of one or more developmental gene regulatory network just as retention of an ancestral morphological trait must rely on retention of a common gene regulatory network. Studying two closely related classes in the same phylum with the same essential morphological feature yet with unique developmental characteristics provides insight into the evolution of these evolutionarily resolute gene regulatory networks. We have developed a new model system using brittle stars to further these studies. In this investigation I have identified key genes of the gene regulatory network (GRN) found in embryonic endo-mesoderm development in the sea urchin, responsible for embryonic skeletogenesis, and compared these key genes with homologues in the brittle star. From the examination of two closely related gene regulatory networks found in two related classes of Echinoderms insight can be gained into the foundation of morphological change over time.

The gastrula stage represents the point in development at which the three primary germ layers diverge. At this point the gene regulatory networks that specify the germ layers are established and the genes that define the differentiated states of the tissues have begun to be activated. These networks have been well characterized in sea urchins, but not in other echinoderms. Embryos of the brittle star Ophiocoma wendtii share a number of developmental features with sea urchin embryos, including the ingression of mesenchyme cells that give rise to an embryonic skeleton. Notable differences are that no micromeres are formed during cleavage divisions and no pigment cells are formed during development to the pluteus larva stage. More subtle changes in timing of developmental events also occur. To explore the molecular basis for the similarities and differences between these two echinoderms, the gastrula transcriptome of Ophiocoma wendtii was sequenced and characterized. I identified brittle star transcripts that correspond to 3385 genes in existing databases, including 1863 genes shared with the sea urchin Strongylocentrotus purpuratus gastrula transcriptome. I have characterized the functional classes of genes present in the transcriptome and compared them to those found in sea urchin. I then examined which members of the germ-layer specific gene regulatory networks (GRNs) of S. purpuratus are expressed in the O. wendtii gastrula. The results indicate that there is a shared "genetic toolkit" central to the echinoderm gastrula, a key stage in embryonic development, though there are also differences that reflect changes in developmental processes. The brittle star expresses genes representing all functional classes at the gastrula stage. Brittle stars and sea urchins have comparable numbers of each class of genes, and share many of the genes expressed at gastrula. Examination of the brittle star genes whose sea urchin orthologs are utilized in germ layer specification reveals a relatively higher level of conservation of key regulatory components compared to the overall transcriptome. I also identify genes that were either lost or whose temporal expression has diverged from that of sea urchins. Overall, the data suggest that embryonic skeleton formation in sea urchins and brittle stars represents convergent evolution by independent cooptation of a shared pathway utilized in adult skeleton formation. Transcription factors are of central importance to both development and evolution. Patterns of their expression and interactions form the gene regulatory networks which control the building of the embryonic body. Alterations in these patterns can result in the construction of altered bodies. To help increase understanding of this process, I compared the transcription factor mRNAs present in early gastrula-stage embryos of the brittle star Ophiocoma wendtii to those found in two species of sea urchins and a starfish. Brittle star homologs were found for one third of the transcription factors in the sea urchin genome and half of those that are expressed at equivalent developmental stages in sea urchins and starfish. Overall, the patterns of transcription factors found and not found in brittle star resemble those of other echinoderms, with the differences largely consistent with morphological differences. This study provides further evidence for the existence of deeply conserved developmental genetic processes, with various elements shared among echinoderms, deuterostomes, and metazoans.

Ophioderma longicauda (Bruzelius, 1805) est un complexe d’espèces cryptiques incluant six lignées mitochondriales (L1-L6), dont certaines (L2-L3-L4) incubent leur descendance, alors que d'autres se reproduisent probablement via des larves lécithotrophes. Afin de définir les limites d’espèces dans le complexe O. longicauda, le statut reproductif des lignées L1 et L3 a été étudié. L’analyse morphologique et génétique a montré qu’il s’agissait d’espèces biologiques différentes, avec notamment différentes périodes de reproduction. De plus, l’analyse par DAPC de 31 marqueurs génétiques a montré que le complexe O. longicauda était constitué de six groupes génétiques distincts. Deuxièmement, l’influence des traits d’histoire de vie sur la connectivité et la diversité génétique a été étudiée. Pour ce faire, 10 marqueurs ont été séquencés pour six populations sympatriques des lignées L1 et L3 en Grèce. La structure génétique était très marquée pour l’espèce incubante L3, tandis que l’espèce dispersante L1 n’a pas montré de structure génétique à cette échelle. L’analyse de la diversité génétique pour ces 10 marqueurs a montré que celle des dispersantes était trois à quatre fois plus élevée que celle des incubantes. De plus, l’analyse de la diversité génétique dans les transcriptomes des L1 et L3 a montré qu’elle était 1.5 à 2 fois plus élevée chez les dispersantes que chez les incubantes. Finalement, deux canaux ioniques impliqués dans la mobilité des spermatozoïdes ont montré une évolution sous sélection positive. Ces résultats suggèrent que la compétition des spermatozoïdes pourrait être un mécanisme d’isolement pré-zygotique chez Ophioderma longicauda. / Ophioderma longicauda (Bruzelius, 1805) is a cryptic species complex including six mitochondrial lineages (L1-L6), of which three (L2-L3-L4) brood their juveniles, whereas the other lineages most likely reproduce using lecithotrophic larvae. In order to define the species limits in the O. longicauda complex, the reproductive status of lineages L1 and L3 was studied. The morphological and genetic study showed that they were different biological species, with notably different reproductive periods. In addition, the analysis of 31 genetic markers using DAPC showed that the O. longicauda complex included six distinct genetic groups. Secondly, the influence of life-history traits on connectivity and genetic diversity was studied. To do so, 10 markers were sequenced for six sympatric populations of lineages L1 and L3 from Greece. The genetic structure was high for the brooding species, whereas the broadcasting species did not display any genetic structure at that scale. The analysis of genetic diversity for these 10 markers showed that diversity was three to four times higher in broadcasters than in brooders. In addition, the analysis of genetic diversity in the L1 and L3 transcriptomes showed that diversity was 1.5 to 2 times higher in broadcasters than in brooders. Finally, two ion channels involved in sperm motility showed an evolution under positive selection. These results suggest that sperm competition might be a mechanism of pre-zygotic isolation in Ophioderma longicauda.