Pattern of ferroportin-1 and Nramp2 gene expression in normal hematopoiesis

N.M. Sposi PhD, A. Rossini PhD, L. Cianetti PhD, E. Saulle PhD, E. Petrucci, M. Gabbianelli PhD, C. Peschle M.D. and U. Testa M.D.

Department of Hematology-Oncology,Istituto Superiore di Sanita, Rome, Italy

Recently, new genes have been identified that are involved in iron metabolism such as the divalent cation transporter-1( DCT1 o Nramp2) and ferroportin-1( o Ireg-1). Nramp2 and Ireg-1 are involved in the import and processing of non-transferrin bound iron and in iron export out of the cells, respectively. Two different Nramp2 mRNAs are generated by alternative polyadenylation/splicing that have or lack a 3¡¦ iron responsive element ( +IRE or {IRE, respectively ). We have investigated the expression of Nramp2 and ferroportin-1 genes by semiquantitative RT-PCR analysis on early hematopoietic progenitor cells purified from human adult peripheral blood and induced in liquid suspension culture to unilineage erythroid, granulocytic, monocytic and megakaryocytic differentiation. The two mRNA isoforms of Nramp2 are undetectable in quiescent hematopoietic progenitor cells and rapidly upmodulated during early erythroid differentiation. At later stages of erythroid maturation Nramp2(+IRE) declines, while Nramp2(-IRE) continues to increase. In contrast the two isoforms of Nramp2 were virtually undetectable in all other hemopoietic lineages. Very interesting was the observation that ferroportin-1 mRNA levels are scarcely expressed in quiescent hematopoietic progenitor cells and are markedly increased during initial stages of hemopoietic differentiation in all four lineages. In parallel we extended our analysis on other types of hemopoietic cells, i.e. in monocytes maturing to macrophages. Northern blot analysis showed that ferroportin-1 gene is expressed at low level in freshly isolated monocytes, but markedly increased when these cells were induced to differentiate to macrophages. Differentiated macrophages incubated in the presence of iron salts showed a sharp increase of ferroportin-1 expression at both mRNA and protein level( by RT-PCR and immunohistochemistry analysis). Gel shift experiments carried out with Ireg-1- and Nramp2- specific IRE probes showed a low, but significant level of RNA-binding activity in monocytes, whose level increased during monocyte to macrophage differentiation. In contrast in iron-treated macrophage cells we observed a decline of RNA-binding activity. These observations indicate that: i) both two mRNA isoforms of Nramp2 are selectively expressed during erythroid differentiation and not in other hemopoietic lineages; ii) Nramp2(-IRE) mRNA is preferentially expressed during the later stages of erythroid maturation; iii) ferroportin-1 mRNA is greatly expressed during initial stages of hemopoietic differentiation and during monocyte to macrophage maturation. We suggest that ferroportin-1 can protect early hematopoietic progenitor cells from iron toxicity and can be involved in iron recycling at level of monocyte/macrophage cells. The regulation of ferroportin-1 expression by iron may involve novel mechanisms of gene regulation.

KEY WORDS: Ireg-1, monocyte, erythroid, progenitor.

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Paper presented at the International Symposium on Predictive Oncology and Intervention Strategies; Paris, France; February 9 - 12, 2002; in the section on Gene Expression, Part 2.