THE G185R MUTATION DISRUPTS FUNCTIONS OF THE IRON TRANSPORTER NRAMP2

Microcytic anemia (mk) mice and Belgrade (b) rats have severe iron def iciency anemia due to defects in intestinal iron transport and erythro id iron utilization. Both animal mutants carry the same missense mutat ion in Nramp2 the first mammalian iron transporter to be identified. T his mutation, in which glycine 185 is changed to arginine (G185R), occ urs within predicted transmembrane domain 4 of the protein. We have pe rformed site-directed mutagenesis of murine Nramp2, focusing on amino acids of transmembrane domain 4 that are highly conserved among Nramp- like proteins. We have expressed each mutant form in transfected cells and examined iron transport function, subcellular localization, and p rotein amounts. All tested forms of Nramp2 localize to the plasma memb rane and to transferrin-containing endosomes. Most transmembrane domai n 4 mutations affect the amount of protein detected and consequently s how diminished iron transport, The G185R mutation, however, causes nea r total loss of Nramp2 function that cannot be fully explained by a de creased amount of protein, indicating that G185R disrupts iron transpo rt through an alteration in the function of Nramp2, rather than degrad ation of the protein. (C) 1998 by The American Society of Hematology.