Age-dependent and iron-independent expression of two mRNA isoforms of divalent metal transporter 1 in rat brain

Abstract

The DMT1(Nramp2/DCT1) is a newly discovered proton-coupled metal-ion transport protein. The cellular localization and functional characterization of DMT1 suggest that it might play a role in physiological iron transport in the brain. In the study, we evaluated effects of dietary iron and age on iron content and DMT1 expression in four brain regions: cortex, hippocampus, striatum, substantia nigra. Total iron content in all regions was significantly lower in the low-iron diet rats and higher in the high-iron diet rats than that in the control animals, showing that dietary iron treatment for 6-weeks can alter brain iron levels. Contrary to our expectation, there was no significant alternation in DMT1(+IRE) and (−IRE) mRNA expression and protein content in all brain regions examined in spite of the existence of the altered iron levels in these regions after 6-weeks’ diet treatment although TfR mRNA expression and protein level were affected significantly, as was expected. The data demonstrates that expression of DMT1(+IRE) and (−IRE) was not regulated by iron in these regions of adult rats. The lack of response of DMT1 to iron status in the brain suggests that the IRE of brain DMT1 mRNA might be not really iron-responsive and that DMT1-mediated iron transport might be not the rate-limiting step in brain iron uptake in adult rats. Our findings also showed that development can significantly affect brain iron and DMT1(+IRE) and (−IRE) expression but the effect varies in different brain regions, indicating a regionally specific regulation in the brain.

Introduction

The DMT1, also known as Nramp2 or DCT1, is a newly discovered proton-coupled metal-ion transport protein. It was first identified on the basis of its homology to Nramp1 in 1995 [14]. In 1997, two groups [10], [16] independently identified DMT1 as the first mammalian transmembrane iron transporter. There are at least two different splice forms of DMT1 [13], [16], [19]. One splice form, called DMT1(+IRE) mRNA, contains an iron-responsive element (IRE) in the 3′-UTR and encodes a 561 amino-acid protein. Another splice form, designated DMT1(−IRE) mRNA, does not contain a classical IRE and encodes a 568 amino-acid protein [16], [24]. DMT1 is widely expressed [10], [16]. At cellular level, DMT1 can express on the endosomal membrane and acts to export iron from the endosome into cytoplasm of the cell [15], [35], [40] in addition to express on the plasma membrane. This implies that DMT1 is important in both transferrin-bound and non-transferrin-bound iron uptake and transport.

In the brain, DMT1 mRNA plus protein is consistently found in neurons and epithelial cells of the choroid plexus and presents at moderate levels in the substantia nigra [3], [5], [16], [23]. The cellular localization and functional characterization suggest that DMT1 might play a role in physiological iron transport in the brain [4], [16], [26], [47]. In the neurons of the substantial nigra in Parkinson's disease (PD), DMT1 is moderately high expressed that coincidentally correlates to the iron abnormally deposition in the same area [2]. Therefore, disruption of DMT1 expression may be involved in the increased iron accumulation in PD. In addition to PD, abnormally high level of iron in the brain has also been demonstrated in other neurodegenerative disorders [1], [21], [31], [39]. The oxidative stress induced by the increased iron has been widely believed to be involved in the cascade of events leading to neuronal death in these disorders [22], [25], [31], [34], [36]. Due to the suggested importance of DMT1 in brain iron metabolism and the putative contribution in the development of some neurodegenerative disorders, we investigated DMT1(+IRE) and (−IRE) mRNAs’ expression and protein synthesis in the cortex, hippocampus, striatum and substantia nigra of the rats with different ages or fed with high- or low-iron diet for 6-weeks in this study. Results showed that DMT1 expression in these brain regions is age-dependent but iron-independent. The lack of response of DMT1 to iron status in the brain of adult rats suggests that the IRE of brain DMT1 mRNA might be not really iron-responsive and that DMT1-mediated iron transport might be not the rate–limiting step in brain iron uptake although it might play a critical role in brain cell iron uptake.