Maintaining iron homeostasis is essential for normal hematopoiesis and the prevention of organ damage due to iron overload. Although the C282Y mutation in the HFE gene is commonly found in the Caucasian population, the penetrance of hereditary hemochromatosis is quite variable (1), suggesting that other genes also participate in the regulation of iron homeostasis. The zebrafish, Danio rerio, provides an excellent genetic system for the identification of novel genes involved in iron metabolism, as evidenced by the recent identification of the novel iron transporter ferroportin1 by positional cloning of the mutation responsible for the hypochromic zebrafish mutant, weissherbst (2). Analyzing mutants generated in a large scale chemical mutagenesis screen, I have identified another mutant with hypochromic anemia, HF107, that maps to a novel locus on zebrafish chromosome 20. I propose to clone the gene responsible for this mutant and to characterize the gene product. Complementation studies and low-resolution mapping of another mutant with hypochromic anemia, HM007, resulted in the identification of a new allele of weissherbst that appears to have less severe anemia than the previously identified alleles. Determining the genetic basis for this new allele may provide insight into the regulation and function of ferroportin1. Furthermore, I propose to place ferroportin1 in a genetic pathway governing iron homeostasis by performing a screen for zebrafish mutations that overcome a defect in ferroportin1 function. We hypothesize that ferroportin1 plays a critical role in maintaining iron homeostasis and that its regulation and function depend on specific genes which are likely to be identified in the suppressor screen. These may include (1) hephaestin (2) HFE (3) transferrin receptor (4) novel genes including the postulated iron stores regulator and erythropoietic regulator (5) alternative iron transporters. In addition we expect to find gain of function mutations in the ferroportin1 gene itself. Performing a mutagenesis screen facilitates evaluation of the interactions of all these factors simultaneously and may result in the identification of novel genes involved in iron homeostasis. These novel genes may become molecular targets for the treatment of iron overload conditions, such as hereditary hemochromatosis.
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