Transcription factor gene regulatory networks and biomechanics of germ layer formation

The establishment of the three germ layers (ectoderm, endoderm and mesoderm) during gastulation marks one of the most important steps in the development of an organism as it makes the embryo multi-layered and often goes hand in hand with formation and elongation of the AP axis. In particular, the formation of the mesoderm is characterised by highly dynamic morphogenetic processes, which are often associated with an epithelial-to- mesenchymal transition (EMT) and cell migration. EMT is a multi-step process, which results from the downregulation of the cell adhesion molecule Cadherin, eventually leading to the detachment of individual epithelial cells. A crucial conserved regulator of EMT during gastrulation in vertebrates is the transcription factor Snail. However, the link and transition between the transcriptional gene regulatory networks and the morphogenetic cellular level is still poorly understood. It is the aim of this study to reveal the molecular and cellular mechanisms of the morphogenetic movements during germ layer formation in a simple diploblastic and evolutionary basal model system, the sea anemone Nematostella vectensis. In contrast to published reports, our preliminary functional analyses have indicated an important role for the Snail homolog during gastrulation in Nematostella, which would push the conserved function to the common ancestor 600 Mio years ago. Furthermore, we have identified two classical Cadherins, which undergo a switch of expression during onset of gastrulation and show differential expression in both berm layers. Using state-of-the-art methods in functional genomics (RNA-seq, gene knockdown, ChIP-seq) in combination with analyses of biomechanics and morphogenesis (4D microscopy, quantitative imaging) we want to: 1) investigate the function of Snail during gastrulation, to identify in a genome- wide approach (ChIP-seq) all its target genes and to compare them with Snail targets in Bilateria, 2) analyse in detail the morphogenetic movements and forces during gastrulation and to understand the role of the differentially expressed cadherins as well as other targets of Snail to be revealed by the ChIP-seq analysis. By revealing the link of the transcriptional control network and the biomechanical-morphogenetic level, we hope to provide a framework for the understanding of conservation and divergence of an fundamental developmental process.

Most animals are built of three germ layers, the outer ectoderm, the inner endoderm and the middle mesoderm. These germ layers are formed during early embryogenesis in a process called gastrulation. Depending on the species it involves invagination or immigration of parts of the embryo to the inner side. Notably, the Cnidaria (sea anemones, corals, jelly fish, freshwater polyps), which branched off already about 600 Mio years ago, are composed of only two germ layers, ectoderm and endoderm. Yet, their gastrulation mode is reminiscent of that of other organisms. In this project, we therefore investigated the role of conserved proteins that regulate gastrulation in all other animals, which have also been identified in the sea anemone. We found that interfering with their function also disrupts gastrulation in the sea anemone, suggesting that the general function of these very ancient genes is very similar to that of humans and other animals. Thus, the genetic regulation of gastrulation has been evolved in the common ancestor, at least 600 Mio years ago. Despite this deep conservation of the genetic cascade, we also found evidence for a conserved epigenetic regulation: we found evidence that mechanical stress such as stretching or pressure can induce gene expression and morphogenetic movements using a similar cascade as in vertebrates. We proposed that the interplay between mechanotransduction (i.e. the activation of gene activity by mechanical stress) and genetic regulation contributes to the robustness of animal development, which also goes back to the common ancestor. Since the evolution of three germ layers was probably a very crucial innovation in evolution, we wanted to understand whether the two cell layers in the sea anemones show already any kind of segregation. Indeed, we found that mesodermal functions (e.g. gonads, nutrient storage, muscles) and endodermal functions (digestion, Insulin secretion) are already topologically separated in the diploblastic animal. We propose that the evolution of the mesoderm simply involved a segregation of these distinct territories into three tissue layers.