Functional characterization of the E399D DMT1/NRAMP2/SLC11A2 protein produced by an exon 12 mutation in a patient with microcytic anemia and iron overload.

DMT1 (Nramp2, Slc11a2) mediates iron uptake at the intestinal brush border and across the membrane of acidified endosomes. A single patient with severe microcytic anemia and iron overload was recently reported to carry a mutation in exon 12 of DMT1 (1285G>C). The mutation has two effects: it severely impairs splicing causing skipping of exon 12 and introduces an amino acid polymorphism (E399D) in the protein encoded by the remaining properly spliced transcript found in the patient. The functional properties and possible contribution to disease of the DMT1 E399D mutation are unknown and have been studied in independent mutants at that position (E399D, E399Q, E399A) expressed in LLC-PK1 kidney cells. The 3 mutants are shown to be fully functional with respect to stability, targeting and trafficking to the membrane, and are transport-competent. This indicates that DMT1G1285C is not a complete loss of function but rather that a modest amount of active DMT1 is produced in this patient. This activity may explain the distinguishing iron overload seen in this patient in addition to microcytic anemia that is absent in parallel rodent models of DMT1 deficiency.