Monothiol CGFS glutaredoxins and BolA-like proteins: [2Fe-2S] binding partners in iron homeostasis

Abstract

Monothiol glutaredoxins (Grxs) with a signature CGFS active site and BolA-like proteins have recently emerged as novel players in iron homeostasis. Elegant genetic and biochemical studies examining the functional and physical interactions of CGFS Grxs in the fungi Saccharomyces cerevisiae and Schizosaccharomyces pombe have unveiled their essential roles in intracellular iron signaling, iron trafficking, and the maturation of Fe-S cluster proteins. Biophysical and biochemical analyses of the [2Fe-2S] bridging interaction between CGFS Grxs and a BolA-like protein in S. cerevisiae provided the first molecular-level understanding of the iron regulation mechanism in this model eukaryote and established the ubiquitous CGFS Grxs and BolA-like proteins as novel Fe-S cluster-binding regulatory partners. Parallel studies focused on Escherichia coli and human homologues for CGFS Grxs and BolA-like proteins have supported the studies in yeast and provided additional clues about their involvement in cellular iron metabolism. Herein, we review recent progress in uncovering the cellular and molecular mechanisms by which CGFS Grxs and BolA-like proteins help regulate iron metabolism in both eukaryotic and prokaryotic organisms.

Figure 1

Domain structure of CGFS monothiol Grxs from E. coli, S. cerevisiae, S. pombe, and H. sapiens. The Trx-like domains and Grx-like domains are shown as purple and blue boxes, respectively. The conserved cysteines in the active sites of the Trx and Grx domains are numbered and shown in yellow. Predicted or known mitochondrial targeting signals are shown as pink boxes.

Figure 2

X-ray crystal structure of E. coli Grx4 (PDB 2WCI) (33) with close-up view of the GSH-ligated [2Fe-2S] cluster.

Figure 3

Domain structure of BolA-like proteins from E. coli, S. cerevisiae, S. pombe, and H. sapiens. The protein names of members of the BolA1, BolA2, and BolA3 subfamilies are shown in red, blue, and cyan, respectively. E. coli YrbA is not grouped in any of these subfamilies based on phylogenic analysis (59). The BolA-like domain is shown as a green box, while predicted or known mitochondrial targeting signals are shown as pink boxes. The conserved histidines identified as Fe-S ligands in some family members are numbered and shown in yellow.

Figure 4

Proposed model for S. cerevisiae iron regulation under iron replete (A) and iron depleted (B) conditions. Solid and dotted lines indicate active and attenuated pathways, respectively. Co-activators for Aft1/Aft2 are not included in the model for the sake of simplicity. See text for details.

Figure 5

Domain structure of iron-responsive transcriptional regulators Aft1 and Aft2 from S. cerevisiae (A) and Fep1 and Php4 from S. pombe (B). Conserved, putative Zn finger (ZF) ligands are shown in yellow and putative iron-responsive cysteine residues are shown in cyan. The dark green boxes indicate the positions of nuclear export signals, while the light blue boxes show the locations of nuclear import signals. The gray box in the Php4 CCAAT-binding complex (CBC) binding domain depicts the specific location of the Php2/3/5 binding residues.

Figure 6

Proposed model for S. pombe iron regulation under iron replete (A) and iron depleted (B) conditions. Solid and dotted lines on the left panels indicate active and attenuated pathways, respectively. The right panels in A and B demonstrate the reciprocal transcriptional regulation between Php4 and Fep1 under iron replete and iron depleted conditions. The Fep1 co-repressors Tup11 and Tup12 are not included in the model for the sake of simplicity. See text for details.

Figure 7

Models for [2Fe-2S]²⁺ Grx homodimers (left) and [2Fe-2S]²⁺ Grx-BolA heterocomplexes (right) characterized from E. coli, S. cerevisiae, and H. sapiens. In each case, Grx-BolA heterocomplexes can be formed by titration of Grx homodimers with the apo BolA-like protein. In all CGFS Grx homodimers, the active site cysteines in the Grx-like domains and 2 GSH molecules ligate the [2Fe-2S] clusters. For yeast and human Grx-BolA heterocomplexes, each [2Fe-2S] cluster is ligated by one Grx domain active site cysteine, one GSH, a histidine from the BolA-like protein and an unidentified 4^th ligand. For the E. coli [2Fe-2S]²⁺ Grx4-BolA heterodimer, the ligands provided by BolA have not been identified.