Lipids, stored as triacylglycerol (TAG), play an essential role as a reserve of metabolic energy. In holometabolous insects, these reserves mainly accumulate during the larval stage in the form of TAG in the fat body. The digestion of fatty acid (FA) containing lipids and their processing in the midgut and transport to the fat body for storage are critical to insect development. Dietary cholesterol, or sterols that can be converted to cholesterol, are critical for insects because they cannot make sterols de riovo. Therefore, the absorption of sterols and their transport to tissues is also critical to insect development. Over the past several years we have made substantial progress in elucidating the general outline of the metabolic pathways involved in these processes. The critical unresolved issues include: 1) The mechanism by which FA and sterols are absorbed from the lumen into midgut enterocytes. 2) The mechanisms involved in lipid transfer between the hemolymph lipoprotein lipophorin and tissues. 3) The regulation of lipid export from midgut and lipid storage in the fat body. We propose to elucidate the mechanisms involved in lipid absorption from the midgut lumen and transport to sites of storage in the fat body using Manduca sexta larvae as a model insect through pursuit of the following specific aims: Specific aim 1. To clone and characterize fatty acid and sterol carrier proteins from the midgut. We will elucidate the role of FA and sterol transport proteins in lipid absorption from the midgut lumen into midgut epithelium.Specific aim 2. To establish the mechanism of DAG transport. We will critically test the role of lipophorin receptors and LTP in the transfer of DAG between tissues and lipophorin and we will develop and test quantitatively a model for DAG transport.Specific aim 3. To establish the mechanism of cholesterol transport. We will critically test the mass action hypothesis for the regulation of cholesterol homeostasis in larval M. sexta.Specific aim 4. To clone and characterize lipophorin receptors from the midgut and the fat body. We will elucidate the role of lipophorin receptors in lipid transport.