This group has continued to work on metal homeostasis in Saccharomyces cerevisiae. The FRE1 gene of yeast is similar to the gp91-phox subunit of the NADPH oxidase of human granulocytes. We have studied structure-function relationships of the FRE1 gene, in particular identifying the heme-liganding residues which are probably conserved with the human homolog. We have obtained additional evidence for an iron transport complex present in the yeast plasma membrane and consisting of the multi-copper oxidase, FET3 and the permease, FTR1. CCC2, P-type ATPase similar to the Wilson disease and Menkes disease genes of humans, is required for copper delivery to FET3. The copper-loading function of CCC2 was localized to a post-Golgi compartment by making use of a panel of sec (secretory) and vps (vacuolar protein sorting) mutants with defects in iron uptake and copper loading of FET3. AFT1, the activator of iron transport was shown to interact with a specific DNA sequence, thereby transducing the signal for iron deprivation. Analogously, MAC1 was shown to interact with a distinct target sequence, transducing the signal for copper deprivation. The cellular effects of iron overload were evaluated and found to include a cell cycle arrest in both G1 and G2. Evaluation of excess iron uptake in strains deficient in DNA repair (rad1, rad6, rad52, and apn1) suggests that the G2 arrest is due to DNA damage, while the G1 arrest is due to downregulation of G1 cyclins. Homologs of the FTR1 iron permease have been identified in other organisms, including B. subtilis, S. pombe, and C. albicans. The C. albicans homolog has been cloned by complementation of the FTR1 mutant. A human homolog of the FTR1 iron permease is being sought by complementation. Finally, zinc regulated genes (both induced and repressed) have been identified by screening random lacZ fusions inserted in the yeast genome.
