NRAMP1 LOCUS ENCODES A 65 KDA INTERFERON-GAMMA-INDUCIBLE PROTEIN IN MURINE MACROPHAGES

The murine Nramp1 (natural-resistance-associated macrophage protein) l ocus, formerly known as Ity/Lsh/Bcg, was isolated previously on the ba sis of chromosomal location, and as conferring natural resistance to i nfection against intracellular macrophage pathogens. The gene encodes a transporter molecule of unknown function. We have prepared polyclona l antisera against the C-terminal 35 amino acids of murine Nramp1. Thi s serum is reactive towards a 65 kDa protein, expressed in murine macr ophage cells from resistant or susceptible mice stimulated with interf eron-gamma and lipopolysaccharide, but not in non-macrophage cells. Ev idence indicates that Nramp1 is localized in a subcellular membrane ra ther than at the cell surface. This evidence includes: the identificat ion of conserved endocytic targeting motifs following inspection of hu man and murine Nramp sequences; the enrichment of Nramp1, following ma gnetic selection of phagolysosomal vesicles from activated macrophages that were allowed to phagocytose magnetic, IgG-coated beads; confocal microscopy. These studies place Nramp1 on a membrane in close proximi ty to obligate intracellular pathogens. A link between Nramp1 and diva lent-cation transport is suggested by sequence similarity with yeast S MF1. Evidence showing modulation of Nramp1 protein levels by iron chel ation provides a direct link with Nramp1 function and divalent-cation metabolism.