Differential iron transport into phagosomes isolated from the RAW264.7 macrophage cell lines transfected with Nramp1Gly169 or Nramp1Asp169

The transport of iron by RAW264.7 macrophage cell lines transfected with either Nramp1(Gly169) (resistant) or Nramp1(Asp169) (susceptible) alleles was assessed, We found no difference between resistant and susceptible cells in the rate of Fe import or export when Fe transport was measured in intact cells. In contrast, the rate of Fe import by later-bead phagosomes isolated from resistant cells was more than double the rate by lates-bead phagosomes from susceptible cells. Similarly, phagosomes isolated fi om resistant cells that had been pre-labeled with Fe-55-citrate before phagocytosis contained up to four times as much Fe as the corresponding phagosomes from susceptible cells. Phagocytosis of Mycobacterium avium was accompanied by an increase in the production of hydroxyl radicals by Nramp1(Gly169)-transfected macrophages but not Ly macrophages transfected with the susceptible allele, These results are consistent with the hypothesis that Nramp1 functions to transport Fe into the bacterium-containing phagosome where it serves as a catalyst for the Haber-Weiss reaction, which accounts for the increased capacity of these cells to limit mycobacterial growth.