The ABCA1 transporter orchestrates cellular lipid homeostasis by promoting the release of cholesterol to plasmatic acceptors. The molecular mechanism is, however, unknown. We report here on the biophysical analysis in living HeLa cells of the ABCA1 lipid microenvironment at the plasma membrane. The modifications of membrane attributes induced by ABCA1 were assessed at both the outer and inner leaflet by monitoring either the lifetime of membrane inserted fluorescent lipid analogues by fluorescence lifetime imaging microscopy (FLIM) or, respectively, the membrane translocation of cationic sensors. Analysis of the partitioning of dedicated probes in plasma membrane blebs vesiculated from these cells allowed visualization of ABCA1 partitioning into the liquid disordered-like phase and corroborated the idea that ABCA1 destabilizes the lipid arrangement at the membrane. Specificity was demonstrated by comparison with cells expressing an inactive transporter. The physiological relevance of these modifications was finally demonstrated by the reduced membrane mobility and function of transferrin receptors under the influence of an active ABCA1. Collectively, these data assess that the control of both transversal and lateral lipid distribution at the membrane is the primary function of ABCA1 and positions the effluxes of cholesterol from cell membranes downstream to the redistribution of the sterol into readily extractable membrane pools.

The ABCA1 transporter is the prototype of the A class of mammalian adenosine triphosphate binding cassette transporters and one of the largest members of this family. ABCA1 has been originally identified as an engulfment receptor on macrophages and, more recently, it has been shown to play an essential role in the handling of cellular lipids. Indeed by promoting the effluxes of membrane phospholipids and cholesterol to lipid-poor apoprotein acceptors, ABCA1 controls the formation of high-density lipoproteins and thus the whole process of reverse cholesterol transport. A number of additional phenotypes have been found in the mouse model of invalidation of the ABCA1 gene. In spite of their clinical diversity, they all are extremely sensitive to variations in the physicochemical properties of the cell membrane, which ABCA1 controls as a lipid translocator.