The goal of this project is to define the role of the heme export protein Feline Leukemia Virus, subgroup C,Receptor (FLVCR) and heme signaling in hematopoiesis. Cats viremic with Feline leukemia virus, subgroup C(FeLV-C) down-regulate the cell surface expression of FLVCR and develop a profound macrocytic anemia.Mice in which Flvcr is deleted have a comparable clinical phenotype, and in both species, erythroiddifferentiation arrests at the CFU-E (colony-forming unit-erythroid)/proerythroblast stage. These data led us tohypothesize that FLVCR protects cells when heme synthesis precedes or exceeds globin synthesis and that ifthese steps were poorly coordinated, ineffective erythropoiesis and anemia would result. This grant will test ourhypothesis by directly measuring free heme in developing erythroid cells with a luciferase reporter containingthe MARE sequences (Bach1 binding site) of the -globin promoter and by measuring heme-regulated eIF2 kinase (HRI) and eIF2 phosphorylation. We will show that excess heme induces anemia with rescueexperiments designed to restore the intracellular free heme balance of Flvcr-deleted mice by decreasing hemesynthesis or by increasing heme degradation using methods independent of FLVCR. In studies of rps6+/- mice,we will determine whether this pathophysiology could be relevant to the macrocytic anemia of DiamondBlackfan anemia (results from RPS19 haploinsufficiency, failed ribosome assembly, and slowed translationinitiation) and the 5q- myelodysplasia syndrome (acquired RPS14 deletion). We will also define the molecularpathways which trigger erythroid marrow failure by transcriptional profiling. In addition, we will explore the roleof FLVCR in other hematopoietic cells using comparable experimental strategies, since our recent datasuggest that FLVCR is needed for the maturation of double positive T cells to single positive T cells in thymusand may have a role in megakaryocyte endomitosis. We can thus determine whether these different lineagesshare regulatory pathways.Together these studies should provide significant insights into heme signaling and toxicities duringhematopoietic cell differentiation.
. When mice lack FLVCR, a heme export protein, they develop a severe anemia and macrocytosis (large red cells), akin to children with Diamond Blackfan anemia and adults with myelodysplasia. We will determine how and why the early red cells die and thus will gain insights into the pathophysiology, and potentially the therapy, of these diseases. We should also learn about the shared molecular mechanisms controlling red cell, T cell and platelet production.
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