Regulation of gastrulation movements by planar cell polarity genes in zebrafish
During vertebrate gastrulation, mesodermal and ectodermal cells undergo convergent extension (CE), a process characterised by cellular rearrangements in which polarised cells intercalate along the medio-lateral axis leading to elongation of the antero-posterior axis. This thesis aims to prove that genes which have been implicated in the establishment of planar cell polarity (PCP) in Drosophila are conserved in the non-canonical Wnt pathway that regulates CE in zebrafish during gastrulation. Firstly, I analyzed functions of Wnt5 and Wnt 11 ligands and of Fz2 and Fz7 receptors in regulating CE movements. Here, I will show that pipetail (ppt/wnt5) mutant is required for regulating CE movements in posterior mesendoderm and ectoderm while its function in the anterior mesendoderm is redundant to siberblick (slb/wntl 1). Based on gene expression analyses, loss of function and gain of function analyzes, the interaction between these Wnt ligands and Fz receptors is dependent on time, concentration and position of expression during gastrulation. Secondly, I will describe the isolation and functional characterisation of the zebrafish homologue of Drosophila prickle (pkl) during gastrulation. Zebrafish pkl functions together with Sib/Wnt 11 and Ppt/Wnt5 to regulate CE movements due to abrogation of Pkl function by morpholino that leads to defective CE movements, enhances the slb/wntl 1 and ppt/wnt5 phenotypes and suppresses the ability of wnt 11 to rescue the sib phenotype. Gain-of- function of Pkl also inhibits CE movements. Additionally, I found that Pkl could destabilise Dsh and thereby block the ability of Fz to target Dsh to the cell membrane by down-regulating levels of Dsh protein. These results suggest that Pkl acts in the non- canonical Wnt pathway to regulate CE, but it is unlikely to be a simply linear component of this pathway. I will describe the isolation of zebrafish flamingo (fmi) genes and characterise the expression pattern during gastrulation. I also describe functional characterisation of those genes during gastrulation. Initially, abrogation of Fmi function by morpholino leads to weakly defective CE movements enhances the slb/wntl 1 phenotype and defective epiboly phenotypes of the offroad (ord)/fmil mutants. In contrast, a dominant negative approach with a mutant form of Fmi, led me to conclude that Fmi is involved in the regulation of CE movements. Together with analyses of Fmi protein, I will discuss how Fmi regulates different gastrulation movements, such as convergent extension and epiboly.