Abstract
In the yeast Saccharomyces cerevisiae, uptake of iron is largely regulated by the transcription factor Aft1. cDNA microarrays were used to identify new iron and AFT1-regulated genes. Four homologous genes regulated as part of the AFT1-regulon (ARN1-4) were predicted to encode members of a subfamily of the major facilitator superfamily of transporters. These genes were predicted to encode proteins with 14 membrane spanning domains and were from 26 to 53% identical at the amino acid level. ARN3 is identical to SIT1, which is reported to encode a ferrioxamine B permease. Deletion of ARN3 did not prevent yeast from using ferrioxamine B as an iron source; however, deletion of ARN3 and FET3, a component of the high affinity ferrous iron transport system, did prevent uptake of ferrioxamine-bound iron and growth on ferrioxamine as an iron source. The siderophore-mediated transport system and the high affinity ferrous iron transport system were localized to separate cellular compartments. Epitope-tagged Arn3p was expressed in intracellular vesicles that co-sediment with the endosomal protein Pep12. In contrast, Fet3p was expressed on the plasma membrane and was digested by extracellular proteases. These data indicate that S. cerevisiae has two pathways for ferrrioxamine-mediated iron uptake, one occurring at the plasma membrane and the other occurring in an intracellular compartment.
Highlights
In the yeast Saccharomyces cerevisiae, uptake of iron is largely regulated by the transcription factor Aft1. cDNA microarrays were used to identify new iron and AFT1-regulated genes
The budding yeast Saccharomyces cerevisiae has two characterized systems of elemental iron uptake, a low affinity system encoded by FET4 (1) and a high affinity system encoded by FET3 and FTR1 (2, 3)
One of the genes, ARN3 (SIT1), was previously reported to exhibit an AFT1-independent pattern of iron regulation and not to contain an AFT1 consensus binding site (13), our results show that ARN3 is ironand AFT1-regulated
Summary
In the yeast Saccharomyces cerevisiae, uptake of iron is largely regulated by the transcription factor Aft1. cDNA microarrays were used to identify new iron and AFT1-regulated genes. Four homologous genes regulated as part of the AFT1-regulon (ARN1– 4) were predicted to encode members of a subfamily of the major facilitator superfamily of transporters. The budding yeast Saccharomyces cerevisiae has two characterized systems of elemental iron uptake, a low affinity system encoded by FET4 (1) and a high affinity system encoded by FET3 and FTR1 (2, 3). Both transport systems require the action of surface reductases (FRE1 and FRE2) to reduce Fe(III) to the more soluble Fe(II) (4, 5). We further demonstrate that Fet3p is localized to the cell surface, Arn3p is found predominantly in intracellular, post-Golgi vesicles
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