Dietary heme iron is an important nutritional source of iron in carnivores and omnivores that is more readily absorbed than non-heme iron derived from vegetables and grain. Most heme is absorbed in the proximal intestine, with absorptive capacity decreasing distally. We utilized a subtractive hybridization approach to isolate a heme transporter from duodenum by taking advantage of the intestinal gradient for heme absorption. Here we show a membrane protein named HCP 1 (heme carrier protein 1), with homology to bacterial metal-tetracycline transporters, mediates heme uptake by cells in a temperature-dependent and saturable manner. HCP 1 mRNA was highly expressed in duodenum and regulated by hypoxia. HCP 1 protein was iron regulated and localized to the brush-border membrane of duodenal enterocytes in iron deficiency. Our data indicate that HCP 1 is the long-sought intestinal heme transporter.

To investigate the functional significance of mutations in Ferroportin that cause hereditary iron overload, we directly measured the iron efflux activity of the proteins expressed in Xenopus oocytes. We found that wild type and mutant Ferroportin molecules (A77D, N144H, Q248H and V162Delta) were all expressed at the plasma membrane at similar levels. All mutations caused significant reductions in (59)Fe efflux compared to wild type but all retained some residual transport activity. A77D had the strongest effect on (59)Fe efflux (remaining activity 9% of wild-type control), whereas the N144H mutation retained the highest efflux activity (42% of control). The Q248H and V162Delta mutations were intermediate between these values. Co-injection of mutant and wild-type mRNAs revealed that the A77D and N144H mutations had a dominant negative effect on the function of the WT protein.

Zinc is a vital micronutrient to all organisms and a potential toxicant to aquatic animals. It is therefore of importance to understand the mechanism of zinc regulation. In the present study, we molecularly cloned and functionally characterized a zinc transporter of the SLC39A family [commonly referred to as the ZIP (Zrt- and Irt-related protein) family] from the gill of zebrafish (Danio rerio) (DrZIP1). DrZIP1 protein was found to localize at the plasma membrane and to function as a zinc uptake transporter when being expressed in either chinook salmon (Oncorhynchus tshawytscha) embryonic 214 cells or Xenopus laevis oocytes. In comparison with pufferfish transporter proteins (FrZIP2 and FrECaC) that are known to facilitate cellular zinc uptake, DrZIP1 appears to have high affinity to bind and transport zinc, suggesting that it maybe a high-affinity zinc uptake transporter (Km < 0.5 microM) in fish. Orthologues of DrZIP1 were also identified in both freshwater and seawater pufferfish (Tetraodon nigroviridis and Takifugu rubripes), indicating that these proteins may be functionally conserved among different fish species. DrZIP1 mRNA is expressed in all the tissues examined in the present study and thus DrZIP1 may be a constitutive zinc uptake transporter in many cell types of zebrafish.

A Xenopus oocyte heterologous expression system was used to characterise iron transport properties of two members of the solute carrier 11 (slc11) protein family isolated from rainbow trout gills. One cDNA clone differed from the trout Slc11a containing an additional 52 bp in the exon between transmembrane domains (TM) 10 and 11. The 52 bp contained a stop codon, resulting in a novel isoform lacking the last two TM (termed slc11c). Slc11c and another isoform slc11b, import Fe 2+ at external pHs 6 to 7.4. Trout slc11b Fe 2+ import was more sensitive to inhibition by divalent metals. The novel vertebrate slc11c isoform functions without TM11 and 12.