Contribution of the H63D mutation in HFE to murine hereditary hemochromatosis

Abstract

Hereditary hemochromatosis (HH) is an autosomal recessive disease characterized by iron accumulation in several organs, followed by organ damage and failure. The C282Y mutation in the HFE gene explains 80 -90% of all diagnosed cases of HH in populations of northwestern European ancestry. Targeted disruption of the mouse Hfe gene (or introduction of the murine mutation analogous to the C282Y human mutation) produces a murine model of HH. Another mutation in the HFE gene, H63D, is more prevalent than C282Y. However, the physiological consequences of the H63D mutation (as well as C282Y͞H63D compound heterozygosity) on iron homeostasis are less well established. To evaluate the phenotypic consequences of the C282Y͞H63D and H63D͞H63D genotypes, we produced H67D (corresponding to H63D in humans) and C294Y (corresponding to C282Y in humans) knock-in mice. H67D homozygous mice, C294Y homozygous mice, and H67D͞ C294Y compound heterozygous mice each demonstrated hepatic iron loading. Even on a standard diet, by 10 weeks of age, hepatic iron levels in mice of these three genotypes were significantly higher than those of wild-type littermates. The relative severity of hepatic iron loading was C294Y͞C294Y > C294Y͞H67D > H67D͞ H67D. We conclude that the H67D allele, when homozygous or combined with a more consequential mutation like C294Y, leads to hepatic iron loading. These observations indicate that the H67D mutation leads to partial loss of Hfe function and can contribute to murine HH.knock-in mice ͉ C282Y mutation ͉ H63D mutation ͉ Cre-loxP H ereditary hemochromatosis (HH) is an autosomal recessive disorder characterized by elevated transferrin saturations, hepatic iron overload, and variable clinical manifestations including liver cirrhosis, hypogonadism, heart failure, and arthropathy. Prevalence of mutations in the gene responsible for HH is estimated to be 2-5 per 1,000 in Caucasians. The gene mutated in HH, the hemochromatosis gene (HFE), encodes an MHC class I-like protein (1) that heterodimerizes with ␤ 2 -microglobulin (␤ 2 M). Most cases of HH arise from a founder mutation in HFE (845G to -A) that results in the amino acid substitution C282Y and prevents the association of HFE with ␤ 2 M (1-3). Functional significance of HFE-␤ 2 M complex in iron homeostasis was supported from the observations that HFE-␤ 2 M is physically associated with transferrin receptor 1 (TfR1) in human placenta (4), in crypt enterocytes of duodenum (5), and in cultured human cell lines (6, 7). The effect of HFE on transferrin-mediated iron uptake seems to depend on the particular cell type studied as well as the magnitude of expression of ␤ 2 M (8).Although the most common HFE mutation associated with HH is C282Y, the most common HFE mutation overall is H63D. Among Caucasian populations, the allele frequency of C282Y is 2-5% and that of H63D is 15-20% (9, 10). The H63D allele frequency has been reported to be as high as 30% in the Basque population of Spain (11). The H63D mutation seems to have evolutionarily predated C282Y and to have ar...

Aquisition, Mobilization and Utilization of Cellular Iron and Heme: Endless Findings and Growing Evidence of Tight Regulation

“…Importantly, 10-15% of patients worldwide demonstrate none of these three mutations, but are not clinically distinguished from others (Sheth and Brittenham 2000;Tomatsu et al 2003).…”

“…Mice lacking β 2 -microglobulin are reported to exhibit an iron-overload similar to hemochromatosis Sheth and Brittenham 2000). The HFE protein forms a stable complex with transferrin receptor 1, but not transferrin receptor 2, and apparently reduces the affinity of the receptor for transferrin (Tomatsu et al 2003;Brissot et al 2004). The complex forms at the cell surface and along the endocytotic pathway for the uptake of transferrin, where HFE and transferrin receptors are co-localized.…”

“…However, in patients with hemochromatosis and mice deficient in β 2 -microglobulin, the crypt cells behave as if starved of iron (Sheth and Brittenham 2000;Ahmad et al 2002;Waheed et al 2002). The expression of the molecules required for intestinal iron absorption is upregulated in the mature enterocytes and reticuloendothelial cells from HFEdeficient humans and mice did not accumulate iron in vivo (Sheth and Brittenham 2000;Waheed et al 2002;Tomatsu et al 2003). These observations are inconsistent with the dominant function of HFE as a competitive inhibitor of iron uptake.…”

Host H67D Genotype Affects Tumor Growth in Mouse Melanoma

“…Our laboratory maintains a colony of C57BL/6 mice with a combination of WT and H67D alleles for the HFE gene [17]. This colony has previously been shown to exhibit characteristics of hemochromatosis (increased hepatic iron loading, brain iron dyshomeostasis) and is superior to the commonly used HFE knockout models because it more closely approximates the human disease [18].…”

Effect of hemochromatosis on manganese neurotoxicity

“…Therefore, several mice models have been generated to mimic Hfe mutations in humans. Among these models, I selected mice with Hfe H67D mutation since this model is a homologous to the H63D mutation in the HFE gene in humans [207]. I added this model in addition to Hfe -/-mice to reflect hereditary hemochromatosis in humans and to further verify the significance of Hfe gene in altering iron metabolism.…”