Ferritins are the main iron storage proteins found in animals, plants, and bacteria. The capacity to store iron in ferritin is essential for life in mammals, but the mechanism by which cytosolic iron is delivered to ferritin is unknown. Human ferritins expressed in yeast contain little iron. Human Poly r(C)-Binding Protein 1 (PCBP1) increased the amount of iron loaded into ferritin when expressed in yeast. PCBP1 bound to ferritin in vivo, and bound iron and facilitated iron loading into ferritin in vitro. Depletion of PCBP1 in human cells inhibited ferritin iron loading and increased cytosolic iron pools. Thus, PCBP1 can function as a cytosolic iron chaperone in the delivery of iron to ferritin.Ferritins are iron storage proteins that are ubiquitously expressed in animals, plants, and bacteria. They serve both to sequester excess iron taken up by the cell and to release stored iron to meet the cell's metabolic needs during iron scarcity (1). In animals, ferritin is a cytosolic heteropolymer consisting of 24 subunits of H-and L-isoforms that assemble into a hollow sphere into which iron is deposited. Ferritin H-chains contain the iron-binding and ferroxidase activities that are required for mineralization of the ferritin core. Deletion of the H-ferritin gene is lethal in mice (2) and in flies (3).In cells, metallochaperones deliver metals to their cognate enzymes and transporters. Although cytosolic copper and nickel chaperones have been described (4-7), no cytosolic iron chaperones have been identified, despite the presence of numerous iron-dependent enzymes in the cytosol. Frataxin, the protein lacking in the neurological disease Friedreich's ataxia, functions as a mitochondrial iron chaperone for iron-sulfur cluster and heme biosynthesis (8,9).Fungi are anomalous among eukaryotes in that they do not express ferritins. We expressed human H-and L-ferritins in the yeast Saccharomyces cerevisiae. The peptides assembled into multimeric complexes with properties similar to native human ferritins, but contained only small amounts of iron ( fig. S1, A and B). We hypothesized that yeast might also lack the requisite iron chaperones needed for delivery of iron to ferritin and designed a genetic screen to identify human genes that, when expressed in yeast, could increase the amount of iron loaded into ferritin. We introduced an iron-regulated FeRE/HIS3 reporter construct (10) into a yeast strain expressing H-and L-ferritin (Fig. 1A). This construct confers histidine prototrophy to cells when the reporter is bound and transcriptionally activated by Aft1p, the major irondependent transcription factor in yeast. Aft1p is activated during periods of cytosolic iron depletion (11), which could occur if substantial amounts of cytosolic iron were diverted into ferritin. †To whom correspondence should be addressed.
All living organisms require nutrient minerals for growth and have developed mechanisms to acquire, utilize, and store nutrient minerals effectively. In the aqueous cellular environment, these elements exist as charged ions that, together with protons and hydroxide ions, facilitate biochemical reactions and establish the electrochemical gradients across membranes that drive cellular processes such as transport and ATP synthesis. Metal ions serve as essential enzyme cofactors and perform both structural and signaling roles within cells. However, because these ions can also be toxic, cells have developed sophisticated homeostatic mechanisms to regulate their levels and avoid toxicity. Studies in Saccharomyces cerevisiae have characterized many of the gene products and processes responsible for acquiring, utilizing, storing, and regulating levels of these ions. Findings in this model organism have often allowed the corresponding machinery in humans to be identified and have provided insights into diseases that result from defects in ion homeostasis. This review summarizes our current understanding of how cation balance is achieved and modulated in baker's yeast. Control of intracellular pH is discussed, as well as uptake, storage, and efflux mechanisms for the alkali metal cations, Na + and K + , the divalent cations, Ca 2+ and Mg 2+ , and the trace metal ions, Fe 2+ , Zn 2+ , Cu 2+ , and Mn 2+ . Signal transduction pathways that are regulated by pH and Ca 2+ are reviewed, as well as the mechanisms that allow cells to maintain appropriate intracellular cation concentrations when challenged by extreme conditions, i.e., either limited availability or toxic levels in the environment. TABLE OF CONTENTS Abstract 677Introduction 678
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