Mssbauer and LC-ICP-MS investigation of iron trafficking between vacuoles and mitochondria in vma2Saccharomyces cerevisiae.
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Vacuoles are acidic organelles that store FeIII polyphosphate, participate in iron homeostasis, and have been proposed to deliver iron to mitochondria for iron-sulfur cluster (ISC) and heme biosynthesis. Vma2 cells have dysfunctional V-ATPases, rendering their vacuoles nonacidic. These cells have mitochondria that are iron-dysregulated, suggesting disruption of a putative vacuole-to-mitochondria iron trafficking pathway. To investigate this potential pathway, we examined the iron content of a vma2 mutant derived from W303cells using Mssbauer and EPR spectroscopies and liquid chromatography interfaced with inductively-coupled-plasma mass spectrometry. Relative to WT cells, vma2 cells contained WT concentrations of iron but nonheme FeII dominated the iron content of fermenting and respiring vma2 cells, indicating that the vacuolar FeIII ions present in WT cells had been reduced. However, vma2 cells synthesized WT levels of ISCs/hemes and had normal aconitase activity. The iron content of vma2 mitochondria was similar to WT, all suggesting that iron delivery to mitochondria was not disrupted. Chromatograms of cytosolic flow-through solutions exhibited iron species with apparent masses of 600 and 800Da for WT and vma2, respectively. Mutant cells contained high copper concentrations and high concentrations of a species assigned to metallothionein, indicating copper dysregulation. vma2 cells from previously studied strain BY4741 exhibited iron-associated properties more consistent with prior studies, suggesting subtle strain differences. Vacuoles with functional V-ATPases appear unnecessary in W303cells for iron to enter mitochondria and be used in ISC/heme biosynthesis; thus, there appears to be no direct or dedicated vacuole-to-mitochondria iron trafficking pathway. The vma2 phenotype may arise from alterations in trafficking of iron directly from cytosol to mitochondria.