ABCA1-induced cell surface binding sites for ApoA-I.

Arterioscler Thromb Vasc Biol 2007 Jul 3;27(7):1603-9. Epub 2007 May 3.

Division of GI/Nutrition, The Children's Hospital of Philadelphia, University of Pennsylvania School of Medicine, Philadelphia, PA 19104-4318, USA.

Objective: The purpose of this study was to understand the interactions of apoA-I with cells expressing ABCA1.

Methods And Results: The binding of wild-type (WT) and mutant forms of human apoA-I to mouse J774 macrophages was examined. Analysis of total binding at 37 degrees C of 125I-WT apoA-I to the cells and specifically to ABCA1, as determined by covalent cross-linking, revealed saturable high affinity binding in both cases. Determination of the level of cell-surface expression of ABCA1 showed that only about 10% of the apoA-I associated with the cell surface was bound directly to ABCA1. Furthermore, when 125I -apoA-I was cross-linked to ABCA1-upregulated cells and examined by SDS-PAGE, the major (approximately 90%) band migrated as monomeric apoA-I. In contrast to WT apoA-I, the C-terminal deletion mutants delta190 to 243 and delta223 to 243 that have reduced lipid affinity, exhibited marked reductions (50 and 70%, respectively) in their abilities to bind to the surface of ABCA1-upregulated cells. However, these C-terminal deletion mutants cross-linked to ABCA1 as effectively as WT apoA-I.

Conclusions: This study demonstrates that ABCA1 activity creates 2 types of high affinity apoA-I binding sites at the cell surface. The low capacity site formed by direct apoA-I/ABCA1 interaction functions in a regulatory role, whereas the much higher capacity site generated by apoA-I/lipid interactions functions in the assembly of nascent HDL particles.

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http://atvb.ahajournals.org/cgi/doi/10.1161/ATVBAHA.107.1457
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July 2007
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