Iron is an essential nutrient, but its accumulation can be highly cytotoxic. Iron homeostasis must be tightly regulated in order to satisfy cell demand without damage. In the yeast Saccharomyces cerevisiae, this regulation is mediated by two paralogous transcription factors: Aft1p and Aft2p. Under iron starvation conditions, Aft1p activates the transcription of high affinity iron transport related genes by specific binding to the DNA consensus sequence 5'-TGCACCC-3' found in the promoters of these genes. The role of Aft2p remains poorly understood. Aft2p is capable of binding in vitro to the Aft1p DNA consensus sequence and it activates the transcription of some of the Aft1p target gene under specific conditions, such as overexpression, or expression of a gain-of-function allele. We aimed at further understanding the role of Aft2p in a physiological context, by identification of its target genes and characterisation of its mode of action. For this purpose, we performed global transcriptome analyses, Northern blot and chromatin immunoprecipitation experiments using wild type and Δaft1 and/or Δaft2 mutant strains grown under iron depleted conditions. We demonstrated that Aft2p directly activates the transcription of genes that are essential for the iron intracellular sub-compartimentalization and use, while Aft1p does not. We discovered that the consensus sequence in the promoter region of genes specifically activated by At2p was not the 5'-TGCACCC-3' sequence, but the shorter 5'-(G/A)CACCC- 3'. Aft2p plays also a direct role in the transcriptional regulation of some genes involved in zinc metabolism, and it could be involved in the regulation of ergosterol and fatty acid metabolism. We also showed that Aft1p and Aft2p proteins amount is iron-regulated. The modulation of Aft1p protein amount can be attributed to its own transcriptional regulation, while the modulation of Aft2p protein amount involves post-transcriptional regulation.
