Investigating the role of individual fibroblast growth factor receptors in gene regulation during early amphibian development

There is a good understanding of the developmental processes, including neural induction, and anteroposterior patterning, and target genes regulated by FGF signalling during early Xenopus development. However, several different members of the cell-surface receptor tyrosine kinases FGFR family are expressed in early development and the roles of individual FGFRs in mediating the effects of FGF signalling in Xenopus development are not clear. The aim of this project was to investigate the unique roles of FGFRs in mediating FGF signalling, through a meta-analysis of high-throughput transcriptomic data sets and CRISPR/Cas9 genome-modification protocol. The development of a CRISPR/Cas9 protocol targeting fgfr1, fgfr4 and fgfrl1, involved optimisation of embryo injections and fragment analysis, to determine CRISPR/Cas9 targeting efficiency. This protocol enables gene expression analysis of FGF target genes, identified by a meta-analysis of high-throughput transcriptomic RNA-Seq and microarray data sets, and elucidates the unique roles of individual FGFRs in regulating the expression of FGF target genes and thus patterning X. tropicalis embryos. This thesis presents evidence towards the unique roles of FGFRs in development through bioinformatic analyses in regulating different genes, such as egr1, fos and different members of dusp and hes gene families. The CRISPR/Cas9 protocol demonstrated phenotypic defects and mutations in fgfr1, fgfr4 and fgfrl1, however this necessitates further refinement to improve targeting efficiencies. However, this gene-editing technique could be expanded to investigate the individual roles of FGF ligands in early Xenopus development.