Human Mast Cells Secreting Leukotriene C4Express theMRP1Gene-Encoded Conjugate Export Pump

Abstract

Mast cells are known to secrete endogenously synthesized leukotriene C4 (LTC4), but the identity of the responsible export pump in human mast cells was unknown. The multidrug resistance proteins MRP1 and MRP2 have been identified as primary-active ATP-dependent export pumps for various amphiphilic anions including the glutathione conjugate LTC4. We therefore studied the expression at the RNAand protein levels of both MRP1 and MRP2 as well as the ATP-dependent LTC4 transport in the human mast cell line HMC-1. Upon stimulation by 1 microM ionomycin, intact HMC-1 cells generated 26 pmol LTC4/10(8) cells within 20 min. Transport experiments using inside-out HMC-1 membrane vesicles demonstrated an ATP-dependent LTC4 transport amounting to 1.4 pmol x (mg protein)(-1) x min(-1). Reverse transcription PCR indicated that HMC-1 cells express mRNA of MRP1, but not of MRP2 or MRP3. Cloning and sequencing of the amplified PCR fragment confirmed its identity with the human MRP1 sequence. Immunoblots using antibodies against MRP1 and MRP2 demonstrated that HMC-1 cells contain the MRP1 but not the MRP2 protein. Our results indicate that the 190 kDa integral membrane glycoprotein MRP1 mediates the ATP-dependent export of LTC4 from human mast cells to the extracellular space.