Nematostella BMP signaling

During the last decade it became clear that the molecular basis for axis formation is basically conserved within Bilateria. For instance the dorso-ventral axis is established mainly by opposing gradients of the signaling molecules of the family of Bone Morphogenetic Proteins (BMPs) in vertebrates (Dpp in the fruitfly Drosophila, respectively) at the non-neural side and the BMP antogonists (e.g. Chordin) on the neural side. In order to elucidate the evolutionary origin of the dorso-ventral axis we investigate the role of BMP signaling in a representative of a pre- bilaterian animal lineage, the cnidarian sea anemone Nematostella vectensis. Nematostella has one apparent body axis, but we have recently shown that genes coding for homologs BMP2/4 and the antagonists Chordin and gremlin not only exist in Cnidaria, but are asymmetrically expressed with respect to the primary oral-aboral axis, indicating a second, molecularly defined body axis. However, unlike in most Bilateria, BMP2/4 and chordin in Nematostella do not form opposing gradients but are expressed on the same side. The key question is therefore, whether the same molecular system was used independently in Cnidaria and Bilateria to establish asymmetries in the body axis in two different ways or whether the dorso-ventral body axis and the directive axis have a common evolutionary origin. In order to shed light into this fundamental question we recently established functional approaches based on gene knockdown and transgenics. In a recent publication we showed that BMP signaling is required for a symmetry break during gastrulation leading to the asymmetric expression. However, many questions remain unsolved. It is still unclear, how the gradients of BMP expression are interpreted, i.e. how this translates into a gradient of nuclear phosphorylated Smad1, the effector protein of the BMP4 signaling pathway. We intend to address this question by double antibody stainings and in situ hybridisation as well as by generating specific reporter lines for the pSmad localisation. Further, we would like to elucidate the role of two other asymmetrically expressed genes, the BMP like molecule GDF5 and the BMP antagonist Gremlin, which are expressed on the opposite side of BMP2/4 and chordin. In this project, we propose to investigate this question functionally by Morpholino-mediated specific gene knockdowns, ectopic overexpression in transgenics and molecular analysis of numerous marker genes. We expect that our results will contribute significantly to our understanding how the dors-ventral body axis in Bilateria evolved and which role the orthologous molecules play in an animal that appears to have only one obvious body axis.

Most animals have a back-belly body axis, which is established by an activity gradient of a conserved BMP signaling pathway. Interestingly, the superficially radial-looking sea anemones also use the same signaling molecules to establish a second body axis, perpendicular to their main axis. This would not have been that surprising, had not there been roughly 600 million years of independent evolution between cnidarians (sea anemones, corals, jellyfish, Hydra) and bilaterians (animals with head-tail and back-belly axis, such as flies, worms or humans). We assayed the effect of switching all the crucial genes involved in the BMP signaling network off in a sea anemone Nematostella vectensis and thus elucidated the topology of this network. We show that the second body axis of the sea anemone is maintained by interplay of two signaling centres, each producing its own set of BMP ligands and their antagonists. Thus a stable gradient of BMP signaling is established and maintained, keeping one end of the secondary body axis different from the other. We find striking similarities but also significant disparities to the way the same signaling system functions in back-belly axis patterning in vertebrates. Strikingly, mathematical modelling reveals that the stability of the signaling system is very sensitive to changes in the production, diffusion and degradation rates of the shared components of the sea anemone and vertebrate BMP signaling, whereas the parameters describing Nematostella-specific components can vary widely, and thus can probably be easily replaced in the evolution. This hypothesis might explain the divergence of BMP networks observed among animals and suggests that the molecular basis for BMP-dependent axis formation has a common origin 600 Myr ago. Our study also shows a new, BMP signaling-dependent way of regulating the activity of the Hox genes in sea anemones. These important regulatory genes define which structures should develop at which positions along the head-tail axis in bilaterians. Their function in Nematostella still awaits investigation.