Dre2, a Conserved Eukaryotic Fe/S Cluster Protein, Functions in Cytosolic Fe/S Protein Biogenesis

Abstract

In a forward genetic screen for interaction with mitochondrial iron carrier proteins in Saccharomyces cerevisiae, a hypomorphic mutation of the essential DRE2 gene was found to confer lethality when combined with Δmrs3 and Δmrs4. The dre2 mutant or Dre2-depleted cells were deficient in cytosolic Fe/S cluster protein activities while maintaining mitochondrial Fe/S clusters. The Dre2 amino acid sequence was evolutionarily conserved, and cysteine motifs (CX₂CXC and twin CX₂C) in human and yeast proteins were perfectly aligned. The human Dre2 homolog (implicated in blocking apoptosis and called CIAPIN1 or anamorsin) was able to complement the nonviability of a Δdre2 deletion strain. The Dre2 protein with triple hemagglutinin tag was located in the cytoplasm and in the mitochondrial intermembrane space. Yeast Dre2 overexpressed and purified from bacteria was brown and exhibited signature absorption and electron paramagnetic resonance spectra, indicating the presence of both [2Fe-2S] and [4Fe-4S] clusters. Thus, Dre2 is an essential conserved Fe/S cluster protein implicated in extramitochondrial Fe/S cluster assembly, similar to other components of the so-called CIA (cytoplasmic Fe/S cluster assembly) pathway although partially localized to the mitochondrial intermembrane space.

ACKNOWLEDGMENTS

We thank Eduard Hurt of the Biochemie-Zentrum der Universitat Heidelberg (BZH), Heidelberg, Germany, for the plasmids pRS316-Rpl25-eGFP and pRS316-Rps2-eGFP. We thank Tracey Rouault of the Cell Biology and Metabolism Branch of NICHD, NIH, Bethesda, MD, for the antibody to IRP1. We thank Gunter B. Kohlhaw of Purdue University, West Lafayette, IN, for helpful discussions regarding the isopropylmalate isomerase assay and for sharing enzyme substrate with us. We thank John Stevens, Tufts University, and Qifan (Jenny) Zhang, Stuyvesant High School, New York, NY, for assistance with the genetic screening.

D. Pain is supported by AHA 0655946T and NIH R01 AG030504; F. Daldal is supported by NIH GM 38239 and DOE ER20052; E. Bi is supported by NIH R01 GM59216; T. Ohnishi is supported by NIH R01 GM30736; A. Dancis is supported by NIH R01 DK53953.