Docosahexaenoic acid (DHA), the longest and most unsaturated omega-3 abundant in animal cells, has garnered considerable interest because of its participation in a wide variety of human conditions including cancer. To justify the considerable biological energy nature spends to synthesize and tenaciously retain this unusual fatty acid, DHA must have a fundamental and essential function common to all cells. Mounting evidence points to a vital role for DHA in governing membrane structure and function. To harness DHA's potential action as a biological, non-toxic anti-cancer agent, it is imperative to understand DHA's role in membrane structure. The experiments proposed herein will test the hypothesis that the interaction between DHA-containing phospholipids and cholesterol in membranes drives the production of DHA-rich, cholesterol-poor and DHA- poor, cholesterol-rich lipid domains. Subsequent segregation of membrane proteins into the domains will then result in distinct patches of membrane, each possessing a different composition and function. By this hypothesis the altered membrane structure increases the tumor's susceptibility to eradication. The proposed project will greatly clarify DHA's role in dictating membrane structure and function with the following experimental attack. First, domains will be induced in simple bilayers (lipid vesicles) and monolayers with DHA, and domain structure studied with a series of fluorescent membrane structural probes and by fluorescence digital imaging microscopy. Next, the complexity of the system will be increased by adding a purified membrane protein (MHC I, a protein of considerable importance to immunologic tumor surveillance) to the lipid vesicles, and investigating the protein's lipid preference and domain effects on protein clustering, conformation and vertical displacement. The work will then be extended to complex natural membranes (tumor cells). To explore DHA's role in domain formation, the composition and structure of DHA-induced exfoliated vesicles (domains) and parent plasma membranes will be measured. Finally, it will be determined whether DHA-induced domain formation limits tumor cell deformability and thus extravasation potential during metastasis. Successfully unraveling DHA's mode of action should suggest practical clinical, dietary or therapeutic applications for this potentially exciting, useful natural substance.
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