Nuclear Monothiol Glutaredoxins of Saccharomyces cerevisiae Can Function as Mitochondrial Glutaredoxins

Abstract

Glutaredoxins are thiol oxidoreductases that regulate protein redox state. In Saccharomyces cerevisiae, Grx1 and Grx2 are cytosolic dithiol glutaredoxins, whereas Grx3, Grx4, and Grx5 are monothiol glutaredoxins. Grx5 locates at the mitochondrial matrix and is needed for iron/sulfur cluster biogenesis. Its absence causes phenotypes such as inactivation of iron/sulfur enzymes and sensitivity to oxidative stress. Whereas Grx5 contains a single glutaredoxin domain, in Grx3 and Grx4 a thioredoxin-like domain is fused to the glutaredoxin domain. Here we have shown that Grx3 locates at the nucleus and that the thioredoxin-like domain is required for such location. We have addressed the functional divergence among glutaredoxins by targeting Grx2/3/4 molecules to the mitochondrial matrix using the Grx5 targeting sequence. The mitochondrial forms of Grx3 and Grx4 partially rescue the defects of a grx5 null mutant. On the contrary, mitochondrially targeted Grx2 does not suppress the mutant phenotype. Both the thioredoxin-like and glutaredoxin domains are needed for the mitochondrial activity of Grx3, although none of the cysteine residues at the thioredoxin-like domain is required for rescue of the grx5 phenotypes. We have concluded that dithiol glutaredoxins are functionally divergent from monothiol ones, but the latter can interchange their biological activities when compartment barriers are surpassed.

AtGRXcp, an Arabidopsis Chloroplastic Glutaredoxin, Is Critical for Protection against Protein Oxidative Damage

“…2). The dual targets of AtGRXcp have been observed for other plant and yeast proteins (45,48). For example, the plant phosphate transporter, Pht2,1, localizes to the chloroplast envelope in plants but to mitochondria when expressed in yeast (48).…”

“…For example, the plant phosphate transporter, Pht2,1, localizes to the chloroplast envelope in plants but to mitochondria when expressed in yeast (48). The nuclear localized yeast Grx3 has been shown to be able to compensate for Grx5 when localized to the mitochondria (45). Mitochondrial localization of AtGRXcp in yeast is necessary for its function, as evidenced by the fact that when the 63-amino acid signal peptide of AtGRXcp was removed, the truncated form of AtGRXcp was unable to target mitochondria and suppress the sensitivity of grx5 cells to H 2 O 2 (Fig.…”

“…For example, yeast Grx5 is mitochondria-localized (21), and Grx3 targets to nuclei (45), whereas a human PICOT is located in the cytosol (24). In order to gain insight into the function of AtGRXcp, we fused it with GFP at its C terminus and transiently expressed this fusion protein in tobacco leaf cells.…”

One Single In-frame AUG Codon Is Responsible for a Diversity of Subcellular Localizations of Glutaredoxin 2 in Saccharomyces cerevisiae

“…The functions of both systems overlap to a certain extent, although the major difference is that Grxs have a glutathione binding site and are capable of reducing GSH protein-mixed disulfides (6,13,14). Mitochondrial isoforms of each member of both systems have been characterized in yeast (15)(16)(17) and mammals (18 -20). Three Grx subfamilies have been distinguished (21).…”

“…These five Grxs also differ in regard to their subcellular localization. Grx1 is cytosolic, Grx3 and Grx4 are nuclear (17,23), Grx5 is mitochondrial (24), and Grx2 has a dual localization in the cytosol and mitochondria (16). Grx2 stands out among other Grxs for its efficiency in transferring reducing equivalents from reduced lipoamide to oxidized glutathione (25).…”