The yeast Saccharomyces cerevisiae contains a pair of paralogous iron-responsive 
transcription activators, Aft1 and Aft2. Aft1 activates the cell surface iron 
uptake systems in iron depletion, while the role of Aft2 remains poorly 
understood. This study compares the functions of Aft1 and Aft2 in regulating the 
transcription of genes involved in iron homeostasis, with reference to the 
presence/absence of the paralog. Cluster analysis of DNA microarray data 
identified the classes of genes regulated by Aft1 or Aft2, or both. Aft2 
activates the transcription of genes involved in intracellular iron use in the 
absence of Aft1. Northern blot analyses, combined with chromatin 
immunoprecipitation experiments on selected genes from each class, demonstrated 
that Aft2 directly activates the genes SMF3 and MRS4 involved in mitochondrial 
and vacuolar iron homeostasis, while Aft1 does not. Computer analysis found 
different cis-regulatory elements for Aft1 and Aft2, and transcription analysis 
using variants of the FET3 promoter indicated that Aft1 is more specific for the 
canonical iron-responsive element TGCACCC than is Aft2. Finally, the absence of 
either Aft1 or Aft2 showed an iron-dependent increase in the amount of the 
remaining paralog. This may provide additional control of cellular iron 
homeostasis.