Mobility of Capicua in Live Fly Embryos
Capicua (CIC) is a protein that acts as a transcription repressor in developing tissues. The class of CIC proteins is well conserved among metazoan organisms and has been associated with human diseases, including cancer and neurodegeneration. In Drosophila melanogaster, CIC is essential for early embryogenesis because of its function in the Torso pathway, which controls the specifications of terminal regions in the fly embryo. Previous studies have identified that CIC functions through the repression of genes involved in RTK-pathways, however, CIC’s molecular interactions are not well understood and require further investigation. To examine CIC’s mobility at physicological conditions, we measured the diffusion of CIC using Fluorescent Correlation Spectroscopy (FCS) in live fly embryos that express a fusion protein of CIC and superfolder Green Fluorescent Protein (GFP). FCS experiments were conducted in the nuclei and cytoplasm, for various nuclear cycles, and at different positions along the anterior-posterior axis in several embryos. It was found that the best fitting model was a two-component 3D diffusion model, in which one species of CIC is rapidly diffusing (~30 µm2/s) and the remaining fraction is diffusing very slowly (~0.8 µm2/s). This suggests that when CIC binds to its DNA targets it diffuses very slowly and when it is unbound it diffuses freely until it finds a new target. This study provides new information on the CIC’s dynamics in a living organism and better informs us on its molecular interactions as a transcriptional repressor.