Milk lipids, largely composed of triacylglycerols, exist as a dispersion of globules surrounded by a membrane derived from the apical plasma membrane of milk secreting mammary epithelial cells. Within mammary epithelial cells, cytoplasmic lipid droplets appear to be the immediate precursors of milk lipid globules. Cytoplasmic lipid droplets appear to originate as very small, triacylglycerol- rich droplets which bleb or bud from endoplasmic reticulum. These droplets are surrounded by a protein and polar lipid coat material derived from endoplasmic reticulum. Droplets appear to grow by fusion with each other, and the ultimate size distribution of lipid globules in milk may be controlled by the extent of this fusion. Calcium, a cytosol protein fraction, and gangliosides of the lipid droplet surface appear to be involved in fusion of these droplets. Objectives of the proposed study are to establish the mechanism by which droplets are formed and released from endoplasmic reticulum, to determine the nature and mode of action of factors which promote lipid droplet fusion, and to establish how lipid droplets are attracted to and interact with apical plasma membrane. Cell-free systems in which lipid droplets can be generated from endoplasmic reticulum and in which lipid droplet fusions can be induced have been developed. A cell-free system in which lipid droplet-plasma membrane interaction occurs will be developed. Through application of biochemical methods, origin, fusion and interactions of lipid droplets in these cell-free systems will be studied. Results obtained will be correlated with observations made through ultrastructural and immunocytochemical studies of tissue specimens. These studies are directed toward one aspect of the overall goal of this research program, which is to understand what controls the rate of synthesis and secretion of the constituents of milk.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
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Cellular Biology and Physiology Subcommittee 1 (CBY)
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Virginia Polytechnic Institute and State University
Schools of Earth Sciences/Natur
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Keenan, T W; Zierdt, C H (1994) Lipid biosynthesis by axenic strains of Blastocystis hominis. Comp Biochem Physiol Biochem Mol Biol 107:525-31
Ghosal, D; Shappell, N W; Keenan, T W (1994) Endoplasmic reticulum lumenal proteins of rat mammary gland. Potential involvement in lipid droplet assembly during lactation. Biochim Biophys Acta 1200:175-81
Keon, B H; Ankrapp, D P; Keenan, T W (1994) Cytosolic lipoprotein particles from milk-secreting cells contain fatty acid synthase and interact with endoplasmic reticulum. Biochim Biophys Acta 1215:327-36
Moreau, P; Cassagne, C; Keenan, T W et al. (1993) Ceramide excluded from cell-free vesicular lipid transfer from endoplasmic reticulum to Golgi apparatus. Evidence for lipid sorting. Biochim Biophys Acta 1146:9-16
Ghosal, D; Ankrapp, D; Keenan, T W (1993) Low molecular mass GTP-binding proteins are secreted from mammary epithelial cells in association with lipid globules. Biochim Biophys Acta 1168:299-306
Keon, B H; Ghosal, D; Keenan, T W (1993) Association of cytosolic lipids with fatty acid synthase from lactating mammary gland. Int J Biochem 25:533-43
Taranto, P A; Keenan, T W; Potts, M (1993) Rehydration induces rapid onset of lipid biosynthesis in desiccated Nostoc commune (Cyanobacteria). Biochim Biophys Acta 1168:228-37
Keenan, T W; Huang, C M; Zierdt, C H (1992) Comparative analysis of lipid composition in axenic strains of Blastocystis hominis. Comp Biochem Physiol B 102:611-5
Keenan, T W; Dylewski, D P; Ghosal, D et al. (1992) Milk lipid globule precursor release from endoplasmic reticulum reconstituted in a cell-free system. Eur J Cell Biol 57:21-9
Szczepaniak, A; Huang, D; Keenan, T W et al. (1991) Electrostatic destabilization of the cytochrome b6f complex in the thylakoid membrane. EMBO J 10:2757-64

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