CCMEBAZ · Molecular basis of cell communication during a migratory event that establishes brain asymmetry in zebrafish

Molecular basis of cell communication during a migratory event that establishes brain asymmetry in zebrafish

The long-term aim of the undertaken study is to resolve the genetic and cell biological mechanisms that contribute to the establishment of vertebrate brain asymmetry. In the zebrafish forebrain, an event crucial in establishing and elaborating asymmetry is the migration of the parapineal primordium. Parapineal cells are specified bilaterally at the midline. They subsequently assemble into a rosette-like structure and migrate to the left side of the brain. The molecular basis of the cellular communication amongst the parapineal cells during migration is still poorly understood. Previous studies in Dr Wilson's group showed that Fgf8, a growth factor involved in many important processes during vertebrate development, is also essential for the migration of the parapineal and the establishment of brain asymmetry. Fgf8 ligand is expressed widely in the surrounding to the parapineal neuronal tissue and there has been identified one receptor FGFR4 to potentially bind to this ligand in the parapineal cells. In this project we identified the molecular pathways that are activated upon binding of the ligand to the receptor in the parapineal cells. We also identified what is the effect of each pathway in the formation and migration of the parapineal. Additionally, we validated as a tool and utilised a transgenic Fgf signalling reporter line to visualise pathway activation in the parapineal cells in the living embryo. It is well established that upon binding of Fgf ligands to Fgf receptors distinct intracellular molecular pathways may be activated that mediate the effects of Fgf signalling. These include Ras-MAPK, PI3K -Akt and PLC gamma. The most extensively studied and also commonly activated pathway is the RAS-MAPK pathway. We found that the RAS-MAPK pathway does not have a major role in parapineal formation and migration. On the contrary we show that the PI3 - Akt pathway may have a more direct role in parapineal migration and regulation of Fgf reporter activation in the parapineal. We examined the role of these pathways using pharmacological treatments in wild type zebrafish developing embryos as well as using the Fgf reporter line mentioned above. During the course of this study we also examined the cross-talk of Fgf pathway activation with other molecular pathways with important roles during development. We examined the role of Notch signalling pathway by generating a fish line that has compromised Notch signalling pathway activation and also expreses the Fgf reporter transgene. The number of Green fluorescent protein (GFP) positive cells has been counted in both sibling and mutant embryos at 35hpf. This analysis reveals that there are more (almost twice as many) GFP positive cells in the mutant embryos, which is consistent with the hypothesis that Notch signalling pathway has a role in focal activation of Fgf signalling in the parapineal. We are currently preparing a manuscript to be published in a high impact journal describing this study. The above study was undertaken in collaboration with Dr Myriam Roussigne in Centre de Biologie du Dévelopement Université Paul Sabatier, Toulouse, France. During this project the Marie Curie Research Fellow Matina Tsalavouta participated in a screen undertaken in the host laboratory to identify zebrafish carrying mutations in the Wnt pathway that may affect parapineal migration, communication with the associated habenular nuclei and therefore establishment of asymmetry in the zebrafish brain. A relevant novel mutant was identified and characterised. This study is now complete and submitted for publication in a high impact journal. It is currently under review. Finally, we have also initiated a novel complementary project studying the role of Hedgehog (Hh) signalling in regulating Fgf8 expression in the epithalamus. We found that Hh regulates Fgf8 in a temporally restricted manner and we have a novel hypothesis to test further and expand on those findings. Training of the researcher The researcher has been receiving adequate training not only in terms of technical and intellectual skills to execute the project but also in terms of personal and professional development. Technical training on embryological and other developmental biology techniques has been provided to the researcher by other colleagues in the host laboratory. Training in acquiring high resolution confocal imaging expertise has been provided by the staff members of the UCL cell and developmental biology imaging facility. Presentations The researcher has been reporting progress regularly in lab meetings. Three more formal presentations were given at a joint meeting with KCL. A project abstract was submitted for the 7th European Zebrafish meeting, Edinburgh, Scotland, 5-9 July 2011. A project abstract was submitted and a poster presented for the morphogenesis and dynamics of multicellular systems, EMBO series conference, EMBL Heidelberg, Germany 7-9 September 2012. Other activities that contribute to the training of the researcher The researcher has supervised two undergraduate students during their final year research project. The researcher has attended a training course on science communication and has also undertaken the organisation of numerous outreach activities (including hosting school students for short shadowing projects, organising school visits to the laboratory, hosting short visits of university students) introducing them to the research that is undertaken in our lab and this project in particular. The researcher attended a series of professional development courses organised by UCL. These courses include personal development and practical training, career development and academic public speaking and communication skills. Socio-economic impact of the study The undertaken study is concerned with a fundamental research question and contributes significantly to the advancement of knowledge in this area maintaining the United Kingdom at the forefront of this area of research. In particular during the course of this study collaborations have been established with researchers in other European countries further contributing to the development of European Research Area (ERA). Further information on the particular project and the researcher can be found in Dr Wilson's laboratory web page at: - http://www.ucl.ac.uk/zebrafish-group/research/asymmetry.php - http://www.ucl.ac.uk/zebrafish-group/personSingle.php?id=97 - http://www.ucl.ac.uk/zebrafish-group/outreach/

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