FUNCTIONAL INTERACTIONS BETWEEN SYNTHETIC ALKYL PHOSPHOLIPDS AND THE ABC TRANSPORTERS P-GLYCOPROTEIN, STE-6, MRP, AND PGH-1

The ABC superfamily of transporters includes the mammalian P-glycoprot ein family (Class I and Class II P-gps), the multidrug resistance-asso ciated protein (MRP), the Pgh-1 product of Plasmodium falciparum gene pfmdrl, all of which are associated with cellular pleiotropic drug res istance phenomena. STE6, the yeast transporter for the farnesylated pe ptide pheromone a, is also a member of this family. Structural similar ities in this family translate into functional homology as expression of mouse Mdr3S (P-gp), P. falciparum Pgh-1, and human MRP partially re store mating in a sterile yeast mutant lacking a functional STE6 gene. The demonstration that Class II P-gps function as phosphatidylcholine (PC) translocators raise the possibility that other ABC transporters may also interact with physiological lipids. We report the identificat ion of the synthetic lipid and PC analog ET-18-OCH3 (edelfosine) as a substrate for not only Class II P-gp but also for Class I P-gps and su rprisingly for the other ABC transporters MRP, Pgh-1, and STE6. Expres sion of these proteins in the yeast Saccharomyces cerevisiae JPY201 wa s found to confer cellular resistance to cytotoxic concentrations of t his lipid by a factor of 4-20-fold in a growth inhibition assay. The n oted activity of ABC transporters toward this synthetic lipid was spec ific as a mutant variant of Mdr3 (Mdr3F) with reduced activity could n ot convey cellular resistance to ET-18-OCH3. ET-18-OCH3 was also found capable of blocking a-peptide pheromone transport and STE6 complement ation by these ABC proteins. The inhibitory effect of ET-18-OCH3 on ce ll growth and a-factor transport could be abrogated by incubation with the lipid acceptor protein BSA or by enzymatic cleavage by microsomal alkylglycerol mono-oxygenase (MAMO). MAMO and BSA reversal of the eth er lipid effect was only seen in the presence of a functional transpor ter. These results suggest that the group of cytotoxic synthetic PC an alogs studied reveal possible structural and functional aspects common to the ABC transporters tested. Furthermore, the studies with BSA and MAMO suggest that the mechanism of transport of ET-18-OCH3 by these A BC transporters may be related to the flippase mechanism of PC transpo rt by Mdr2.