Transport Regulator Proteins in Lipid-Droplets of Drosophila melanogaster Embryos

Drosophila melanogaster embryos contain lipid droplets at all stages of development, which are moved along microtubules. As development proceeds, the droplets undergo distribution changes, due to altered transport: in Phase I, the droplets move bi-directionally, with no net transport. At a second phase there is net plus-end transport, and the droplets on average move towards the center of the embryo. In Phase III the average transport of the droplets reverses, moving toward the embryo periphery. How these changes in transport are controlled is unknown, but droplet-bound proteins certainly contribute to the process. My poster will describe the search for changing droplet-bound proteins--proteins that change their state of phosphorylation or the amount of protein localized to the droplet. To identify such proteins that are good candidates for key regulators of the changing transport, we purify lipid droplets, and visualize the droplet-bound proteins using 2-D gel electrophoresis analysis. The 2-D gel separates the proteins found on the lipid droplets by both pH and size. The gels are then compared to gels from different stages of the embryo’s development, looking for spots (proteins) which have moved over (phosphorylation) or increased/decreased in amounts. These changing proteins are then excised from these gels, and identified with mass spectrometry.  The identified candidates will then be ranked by potential importance, in part by cross-referencing with published reports on factors that affect droplet position/size, etc., in other contexts.  The results from this experiment are a critical step in understanding regulation of transport, relevant for many disease-related processes.