Final Activity Report Summary - RETINA POLARISATION (Polarisation of the developing retina) The Wnt signalling pathway is involved in many aspects of vertebrate development. Most relevant to our research, it is responsible for the anterior to posterior patterning of the neuroepithelium that will become the brain. When the Wnt pathway is mis-regulated and overactive, cells that are meant to become anterior brain and eyes are re-specified to midbrain. Tcf transcription factors are the nuclear effectors of the Wnt pathway that integrate cell signalling and determine specific gene transcriptional outputs that drive the cell into a specific fate or metabolic state. Though there is plenty of information about how the Wnt pathway works at the cytosolic level, little is known about how the four members of the Tcf factor family are able to direct transcriptional outputs into a diversity of tissues. All Tcf's have the same DNA binding domain and thus the same potential to regulate the same genes so understanding how the pathway exerts different transcriptional programs is a fundamental question to address. In this context, we studied the role of tcf4 splice variants (SV) during anterior to posterior patterning of the zebrafish brain. At the developmental stage at which this happens, the embryo expresses only two tcf4 SVs, one of which includes a new exon that we identified in the course of this work. We show that only the tcf4 SVs that includes this exon have a function in Wnt mediated brain patterning. We further demonstrate that SVs that include the new exon are more prone to bind Groucho co-repressors, which could explain the functionality of the SV. Finally the functionally active tcf4 SV is able to specifically differentially trans-regulate the cdx1 gene reporter in cell luciferase assays. Our main scientific achievement is to show for first time an in vivo functional difference for tcf splice variants. This is mediated by a differential protein interaction with regulators of gene transcription, which finally would direct the transcription of a subset of Wnt pathway downstream target genes. The implications of our work are significant considering that the Wnt pathway is mis-regulated in several types of cancer pathologies. Our findings could open alternatives in the generation of more specific drugs and therapies to overcome cancer. This is also applicable to stem cell research given that the Wnt pathway is also implicated in the maintenance of stem cell identity.
