Poole, Richard James; (2004) The role and regulation of Eph/Ephrins during zebrafish somitogenesis. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

Vertebrate somitogenesis involves the establishment of a segmental pattern of gene expression within the presomitic mesoderm (PSM) and the subsequent translation of this pattern into physical furrows and epithelial somites. In the first part of this thesis we have investigated the role of Eph/Ephrin signalling in this process. We show that in the fused somites mutant, lack of intersomitic boundaries and failure of paraxial mesodermal cells to undergo mesenchymal-to-epithelial transition is accompanied by a lack of Eph/Ephrin signalling interfaces. Using mosaic analysis, we provide evidence for a role for the Eph/Ephrin signalling pathway in somite boundary morphogenesis and epithelialisation. Restoration of the Eph/Ephrin interface in the paraxial mesoderm of fss/tbx24 embryos resulted in the rescue of morphological boundaries and many aspects of epithelialisation. In the second part of this thesis we took two approaches to try to understand how the early prepatterning mechanisms lead to the segmental expression of ephA4 and efnB2a in the rostral PSM. We screened a subtractive library enriched for clones expressed in the PSM. We have isolated two interesting genes from this library, a hes6 and meox1. We present here the analysis of hes6. It does not appear to have a role during somitogenesis but instead appears to regulate early cell-fate specification, gastrulation and neurogenesis. In a second approach we have analysed the promoter regions of the paralogs efnB2a and efnB2b. To facilitate this analysis we tested and then used the IScel meganuclease system. We established two transgenic lines that drive GFP under the control of efnB2a and efnB2b promoters. The efnB2a transgenic line recapitulates the endogenous expression in the PSM but the efnB2b line only drives expression during gastrulation. Based on this and a comprehensive comparison of efnB2a and efnB2b and EfnB2 expression, the evolution of efnB2 promoter regions is discussed.

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