Lipid Translocation from the Cytosolic Leaflet of the Plasma Membrane to the Cell Surface by MultidrugTransporters

Abstract

Many molecules that are exposed on the outside of the cell are involved in recognition and signaling events. This is true for receptor proteins but also for various classes of membrane lipids like the phospholipid phosphatidylserine and various glycosphingolipids. To obtain a better understanding of the function of lipids in these processes we and others have studied the general mechanisms by which lipid molecules reach the cell surface, i. e. the outer leaflet of the plasma membrane. Based on the pioneering work by Pagano and colleagues (Pagano et al., 1983; Lipsky and Pagano, 1985) we have extensively used short-chain analogs of membrane lipids. More specifically, we have added to the cells a ceramide or a diacylglycerol with a C₆-NBD-fluorescent fatty acyl chain at the C2 position of the sphingosine and at the sn2 position of the diacylglycerol (Figure 1). The general idea is that these precursors will be converted to (short-chain) membrane lipids in the cell and will be transported to the plasma membrane. Because of the short-acyl chain these products will display a higher off-rate from membranes than the regular membrane lipids. Conveniently, as an assay for their appearance in the outer leaflet of the plasma membrane, they can be extracted from that surface by albumin in the bathing medium. Obviously, we hoped that the short-chain analogs would follow the same transport pathways as their natural counterparts. However, it has been clear from the start that any finding obtained by the use of short-chain lipids must finally be confirmed for the natural lipids, which generally possess two long acyl chains.