Identification of dietary lipid cues that regulate metabolism in Drosophila.

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Dietary lipids are one major element that defines the caloric content of our diet. In addition, recent studies identified lipids as cues capable to induce a cellular and systemic metabolic response. In the wild fruit flies feed on rotting fruits, a diet rich in calories with plant and fungal components. Fungal lipids are different from plant lipids; fungal fatty acids are shorter and more saturated, and contain fungal sterols rather than plant sterols. Dietary sterols are critical for Drosophila membrane and hormone production because flies are sterol auxotrophs. To study how different lipid compositions affect metabolism in flies I have created two new recipes, plant and yeast food. Both diets share a very similar high caloric richness and have almost identical protein/sugar/lipid ratios. I have discovered that yeast lipids elevate systemic Insulin signaling. The high Insulin signaling levels result in fast development and high reproductive rates, but decrease the average lifespan of adult fruit flies. I found that lipoprotein particles are required to convey the dietary yeast lipid signal to Insulin producing neurons. To reach their neuronal targets, a specific lipoprotein particle called LTP needs to cross the Blood Brain Barrier, which controls all traffic into the brain. I have shown that nutritional yeast lipids elevate intracellular Calcium levels in Blood Brain Barrier cells and that high intracellular Calcium levels are required for the targeted receptor mediated translocation of the specific lipoprotein particle LTP (Lipid transport protein). In the brain, LTP enriches on distinct neurons that are directly connected to Insulin producing cells. My data suggest that the activity of these LTP positive neurons triggers the release of Insulin into circulation.I would now like to understand how these lipids activate BBB cells, and the mechanisms that then allow the transport of LTP across the BBB to specific neurons with the brain. To do so, I propose the following aims:1. To identify dietary lipids that activate Blood Brain Barrier cells, and the mechanisms by which they act (in flies and in a vertebrate cell culture model). 2. To map the glio-neuronal network that connects Blood Brain Barrier cells with specific neurons.3. To identify membrane traffic routes that deliver LTP across the Blood Brain Barrier (in response metabolic changes). My grant proposal outlines here detailed protocols to isolate and identify dietary lipids that activate Calcium signaling in BBB cells, and describes strategies to discriminate between structural lipids (that modulate the composition of plasma membranes to induce cell reactivity) or lipid signal cues. It describes a cutting edge genetic approach to identify the pattern and plasticity of connections between BBB cells and specific neurons. This proposal also outlines how I will use a library of endogenously YFP-tagged Rab proteins to identify membrane trafficking routes across the BBB used by LTP.