Genetic and Biochemical Analysis of High Iron Toxicity in Yeast

Abstract

Iron storage in yeast requires the activity of the vacuolar iron transporter Ccc1. Yeast with an intact CCC1 are resistant to iron toxicity, but deletion of CCC1 renders yeast susceptible to iron toxicity. We used genetic and biochemical analysis to identify suppressors of high iron toxicity in Δccc1 cells to probe the mechanism of high iron toxicity. All genes identified as suppressors of high iron toxicity in aerobically grown Δccc1 cells encode organelle iron transporters including mitochondrial iron transporters MRS3, MRS4, and RIM2. Overexpression of MRS3 suppressed high iron toxicity by decreasing cytosolic iron through mitochondrial iron accumulation. Under anaerobic conditions, Δccc1 cells were still sensitive to high iron toxicity, but overexpression of MRS3 did not suppress iron toxicity and did not result in mitochondrial iron accumulation. We conclude that Mrs3/Mrs4 can sequester iron within mitochondria under aerobic conditions but not anaerobic conditions. We show that iron toxicity in Δccc1 cells occurred under both aerobic and anaerobic conditions. Microarray analysis showed no evidence of oxidative damage under anaerobic conditions, suggesting that iron toxicity may not be solely due to oxidative damage. Deletion of TSA1, which encodes a peroxiredoxin, exacerbated iron toxicity in Δccc1 cells under both aerobic and anaerobic conditions, suggesting a unique role for Tsa1 in iron toxicity.

The J anus transcription factor H ap X controls fungal adaptation to both iron starvation and iron excess

“…Paradoxically, the same redox property makes this metal potentially toxic by causing oxidative stress (Halliwell & Gutteridge, 1984; Lin et al , 2011). Thus, iron homeostasis requires precise regulation of iron uptake and storage to satisfy the cellular needs but to avoid toxic iron excess.…”

Iron toxicity in yeast: transcriptional regulation of the vacuolar iron importer Ccc1

“…Dysregulation of this balance results in iron toxicity and cell death. Studies have shown that removing cytosolic iron either by sequestration in the vacuole or in the mitochondria can act as a protective mechanism against iron toxicity (Chen and Kaplan 2000; Li et al 2001; Lin et al 2011). Ccc1 plays a key role in this balance by importing iron into the vacuole.…”

Responses of Saccharomyces cerevisiae Strains from Different Origins to Elevated Iron Concentrations

“…Despite this complexity, we decided to perform a first approach to analyze iron homeostasis in these strains at the molecular level. For this purpose, we cultivated yeast cells in liquid SC medium to reach the exponential growth phase and used quantitative reverse transcription-PCR (qPCR) to determine the mRNA levels of four genes that are crucial for iron uptake, distribution, and utilization: CTH2, which optimizes iron utilization within cells by regulating multiple iron-dependent metabolic pathways (12); FET3, a copper-dependent ferroxidase essential for high-affinity iron uptake (11); FET4, a low-affinity iron transporter located at the plasma membrane (7,8); and CCC1, responsible for iron transport and storage in the vacuole (15,16). It is worth highlighting that transcriptional activator Aft1 upregulates the expression of genes CTH2, FET3, and FET4 when yeast cells sense low iron (12,28), whereas Yap5 transcription factor increases CCC1 mRNA levels upon iron excess (14).…”

“…When environmental iron levels rise, the yeast Yap5 transcription factor activates specific genes that protect cells from the potential danger of high iron (14). The most relevant transcriptional activation by Yap5 is exerted on CCC1, which encodes a protein that protects cells from excess iron by transporting the metal from the cytoplasm to the vacuole, where it is stored (15,16). Recent studies have demonstrated that S. cerevi-siae does not directly sense extracellular or cytosolic iron concentrations.…”