ABCA1 mutants reveal an interdependency between lipid export function, apoA-I binding activity, and Janus kinase 2 activation.

ABCA1 exports cholesterol and phospholipids from cells by a multistep pathway that involves forming cell surface lipid domains, solubilizing these lipids by apolipoproteins, binding of apolipoproteins to ABCA1, and activating signaling processes. Here we used a mutational analysis approach to evaluate the relationship between these events. We prepared seven naturally occurring mutants and one artificial missense mutant of ABCA1 with varying degrees of impaired function, expressed them to similar levels as wild-type ABCA1 on the cell surface of BHK cells, and measured ABCA1-dependent lipid export, apolipoprotein A-I (apoA-I) binding, and signaling activities. Linear regression analyses showed that cholesterol and phospholipid efflux and cellular apoA-I binding correlated significantly with the ability of ABCA1 to form cell surface lipid domains. Lipid export and cellular apoA-I binding activities and formation of lipid domains also correlated with the amount of apoA-I that could be cross-linked to ABCA1. Moreover, each of these lipid export and apoA-I binding activities correlated with apoA-I-induced Janus kinase 2 (JAK2) activation. Thus, these missense mutations in ABCA1 impair lipid export, apoA-I binding, and apoA-I-stimulated JAK2 activities to similar extents, indicating that these processes are highly interactive components of a pathway that functions to export lipids from cells.