MDR3 P-glycoprotein, a phosphatidylcholine translocase, transports severalcytotoxic drugs and directly interacts with drugs as judged by interference with nucleotide trapping

The human MDR3 gene is a member of the multidrug resistance (MDR) gene fami ly. The MDR3 P-glycoprotein is a transmembrane protein that translocates ph osphatidylcholine. The MDR1 P-glycoprotein related transports cytotoxic dru gs. Its overexpression can make cells resistant to a variety of drugs. Atte mpts to show that MDR3 P-glycoprotein can cause MDR have been unsuccessful thus far. Here, we report an increased directional transport of several MDR 1 P-glycoprotein substrates, such as digoxin, paclitaxel, and vinblastine, through polarized monolayers of MDR3-transfected cells. Transport of other good MDR1 P-glycoprotein substrates, including cyclosporin A and dexamethas one, was not detectably increased. MDR3 P-glycoprotein-dependent transport of a short-chain phosphatidylcholine analog and drugs was inhibited by seve ral MDR reversal agents and other drugs, indicating an interaction between these compounds and MDR3 P-gp, Insect cell membranes from Sf9 cells overexp ressing MDR3 showed specific MgATP binding and a vanadate-dependent, N-ethy lmaleimide-sensitive nucleotide trapping activity, visualized by covalent b inding with [alpha-P-32]8 azido-ATP. Nucleotide trapping was (nearly) aboli shed by paclitaxel, vinblastine, and the MDR reversal agents verapamil, cyc losporin A, and PSC 833. We conclude that MDR3 P-glycoprotein can bind and transport a subset of MDR1 P-glycoprotein substrates. The rate of MDR3 P-wg lycoprotein-mediated transport is low for most drugs, explaining why this p rotein is not detectably involved in multidrug resistance. It remains possi ble, however, that drug binding to MDR3 P-glycoprotein could adversely affe ct phospholipid or toxin secretion under conditions of stress (e.g. in preg nant heterozygotes with one MDR3 null allele).