Proteins Localized to Lipid Droplets Regulate Motion in Drosophila Melanogaster Embryos
Drosophila melanogaster embryos contain lipid droplets, which are moved along microtubules by kinesin-1 and dynein in the syncitium. As development proceeds, the droplets’ distribution changes, due to altered transport. In phase I, the droplets move bi-directionally, with no net transport. In phase II there is a net plus-end transport, with droplets moving basally, towards the center of the embryo. In phase III average transport is apical, moving droplets towards the periphery. The protein Halo contributes to these changes, but, overall how transport is controlled is unknown. We hypothesize that changes in droplet-bound proteins contribute to this altered motion. Such changes could reflect altered phosphorylation, or overall amounts of protein localized to the droplet. In order to identify the potential candidates, lipid droplets are purified and changes in droplet-bound proteins are visualized using 2-D gel electrophoresis. We purify droplets from different stages of the embryo’s development, and search for differences in their protein content by looking for spots (proteins) on the gel which have moved (phosphorylated) or increased/decreased in amount. These candidate proteins are then visualized with a luminescent stain, excised from the 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 lipid droplet aspects such as position or size. The results from this experiment are a critical step in understanding regulation of transport, relevant for many disease-related processes as well as elucidating novel roles of lipid droplets.