We study the genetic control of neuronal differentiation and of pigmentary differentiation. This work has two major biological applications, one relating to the development of ocular function and the other relating to the development of ocular tumours (uveal melanoma and retinoblastoma).
Mitf (for microphthalmia) is a Myc-type transcription factor (a bHLHZip factor) expressed in melanocytes and is essential for the proliferation/differentiation of these cells. Subcellular localisation of this transcription factor encoding distinct isoforms may also be important for its function. To study this point, we have prepared vectors encoding biologically active Mitf proteins fused to GFP, making it possible to study the kinetics of Mitf transport between the nucleus and the cytoplasm. GFP can be destroyed by photobleaching in a small cellular volume within the nucleus or cytoplasm, with no effect on cell viability (Fig. 1). It is then possible to measure the fluorescence corresponding to the repopulation of the bleached zone by new Mitf-GFP fusion proteins; consequently, the diffusion dynamics can be studied (fluorescence recovery after photobleaching).
Uveal Melanoma (UM) is the most frequent and aggressive ocular primary tumor in adults with 6 new cases per million per year. The gene expression signature identifies two molecular classes of UM with low risk (Class 1) and high risk (Class 2) of metastasis essentially located in the liver. The use of cytogenetics, genomic and transcriptomic studies have conducted to the identification of genes more specifically associated with, and hopefully responsible for metastasis.
Our team is interested in understanding how cytosqueleton dynamics, proteases and DNA reparation processes are used to facilitate cell migration and invasion.
Our general objective has been to dissect the mechanisms leading to uveal melanoma metastasis. We have shown that phosphatase PTP4A3 known to promote cell migration and invasion in cancer is differentially expressed between the two class of UM and predictive of metastasis. It is now essential to identify the targets of this phosphatase to understand how the UM become metastatic.
Our plan is to analyze the effect of PTP4A3 on targets phosphorylation and their biochemical pathways to understand the control of cell migration and invasion.
Our specific approach to understanding metastasis development is based on tools that combine cell biology, molecular biology and developmental biology. We examine the effects of up and down regulation of genes of interest in cell migration in vitro by time lapse videomicroscopy and cell invasion using an in vivo model of chick embryo chorioallentoïd membrane to assess for uveal melanoma cells intravasation.