Intercellular signals play a fundamental role in animal development, from the establishment of the germ layers in the early embryo to the specification of distinct cell types in specialised organs. Understanding how cells interpret and respond to intercellular signals is thus of great importance. In the five years since they were discovered in vertebrates, Hedgehog family proteins have been implicated in many inductive interactions, one of the best examples being the patterning of the neural tube and somites by the axial mesoderm. Our studies have focused on notochord-somite interactions in the zebrafish and in particular on the role of Hedgehog signalling in the specification of muscle cell identity. Two distinct types of slow-twitch muscle cells can be identified in the zebrafish myotome and our studies have revealed that these specified either in response to distinct forms or distinct levels of Hedgehog signalling. The first part of this proposal aims to discriminate between these possibilities and to investigate the role of different forms of the Hedgehog re3ceptor subunit, Patched, in this process. Various mutations have been identified that block the differentiation of specific muscle cell types in the zebrafish: our studies have indicated that most of these act by eliminating or attenuating Hedgehog signalling; however, one mutation u-boot seems to act downstream of the Hedgehog pathway and thus identifies a gene required for cells to differentiate appropriately in response to the inductive cue. The second part of this proposal aims to discover the molecular nature of the u-boot gene product by positional cloning of the locus. Since Sonic Hedgehog is implicated in muscle patterning in all vertebrate specifies, the results of these studies should be of general significance.

During development, each of our cells must resolve its fate, shape and position. Revealing how these decisions are made is critical to understand how mammalian embryos form, yet their real time control in vivo remains unknown. Because fixed specimens cannot capture real time cell dynamics, I have established a cross-disciplinary research program that combines imaging technologies, quantitative and genetic methods, to study cells directly in the living mouse embryo. We have recently showed how transcription factors (TFs) bind to the DNA in single cells of living embryos to control cell fate. We also started to discover some of the mechanisms explaining how cells in the embryo change their shape, using long filopodial protrusions, and how they regulate their mechanical properties to adopt distinct positions within the embryo. Our overriding goal is to exploit the application of our single-cell imaging and quantitative technologies in the living mouse embryo, to reveal how the mechanisms that govern the first changes in cell fate, shape and position are integrated across the entire embryo to ensure normal embryogenesis.