COMPARISON OF TRANSPORT-PROPERTIES OF THE REDUCED FOLATE CARRIER AND FOLATE RECEPTOR IN MURINE L1210 LEUKEMIA-CELLS

This laboratory previously described an L1210 murine leukemia cell lin e with a functional defect in the reduced folate carrier and increased expression of folate receptor-beta (F2-MTX(r)A). This cell line was u sed to characterize methotrexate (MTX) influx mediated by folate recep tor-beta and to compare this with influx mediated by the reduced folat e carrier in L1210 parental cells. Influx of 0.2 mu M MTX in F2-MTX(r) A cells was one-third that of L1210 cells and was abolished by very lo w concentrations of folic acid. Kinetic analysis revealed that MTX tra nsport mediated by folate receptor-beta exhibited an influx K-t one-th ird, and an influx V-max one-fourth, that of the reduced folate carrie r. Metabolic inhibitors markedly suppressed influx in F2-MTX(r)A cells but had no effect on MTX influx in L1210 cells. MTX influx in both ce ll lines was inhibited by the organic anions probenecid, sulfobromopht halein, and CI-920, but to a lesser extent in F2-MTX(r)A cells. The in hibitory effects of these anions on transport in F2-MTX(r)A cells coul d be attributed to their inhibition of MTX binding to the folate recep tor. Although MTX influx in both cell lines was not sodium dependent, removal of extracellular chloride increased influx 2-fold in L1210 cel ls while markedly inhibiting influx in F2-MTX(r)A cells. Substitution of Cl- with isethionate or NO3- partially restored influx in the latte r cells, whereas SO42- was inhibitory. Anions enhanced MTX binding to folate receptor-beta with isethionate > SO42- > Cl-. Decreasing the bu ffer pH to 6.2 produced a 69% reduction, and a 260% increase, in MTX i nflux in L1210 cells and F2-MTX(r)A cells, respectively. The data indi cate that folate receptor-beta-mediated MTX influx has properties fund amentally different from transport mediated by the reduced folate carr ier in terms of energy, ion, and pH dependence. There was no evidence indicating that these processes are functionally linked.