The Drosophila model will be used for two complementary and unbiased genetic screens, designed to identify novel genes that affect iron homeostasis in multi-cellular organisms. S. cerevisiae has been used successfully for the same purpose, however this unicellular organism does not express neither ferritin (the major iron storage protein) nor iron regulatory proteins (the major regulators of intracellular iron homeostasis in mammals). Knockout mouse models of known iron metabolism genes have been developed to study relevant human disorders, but have limited value for identification of novel genes. To study ferritin expression in Drosophila, we have previously characterized a fly strain expressing GFP-Fer1HCH, which is incorporated into endogenous ferritin complexes that sequester iron and show subcellular and tissue specific expression patterns that are identical to wild type ferritins. Green fluorescence is readily observed in the intestine, brain and in large characteristic pericardial cells of first instar larvae. If fed on iron, ferritin expression is induced in the anterior portion of the intestine. For the first screen EMS-mutagenized chromosomes will be crossed in trans with the GFP-ferritin carrying flies, looking for altered expression patterns. The second screen is based on the observation that ubiquitous overexpression of ferritin can be lethal under iron limiting conditions. Ferritin-induced lethality is rescued by iron supplementation, suggesting that the cause of lethality is related to iron sequestration by excess ferritin. We expect that, under low iron conditions, flies overexpressing ferritin will only grow if we genetically reduce the expression of factors important in ferritin assembly or iron incorporation into ferritin. Combining the two screening strategies we expect to unravel novel genes that impact on iron homeostasis and introduce the Drosophila model in BioIron research.
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