The Multidomain Thioredoxin-Monothiol Glutaredoxins Represent a Distinct Functional Group

Abstract

Monothiol glutaredoxins (Grxs) with a noncanonical CGFS active site are found in all kingdoms of life. They include members with a single domain and thioredoxin-Grx fusion proteins. In Saccharomyces cerevisiae, the multidomain Grx3 and Grx4 play an essential role in intracellular iron trafficking. This crucial task is mediated by an essential Fe/S cofactor. This study shows that this unique physiological role cannot be executed by single domain Grxs, because the thioredoxin domain is indispensable for function in vivo. Mutational analysis revealed that a CPxS active site motif is fully compatible with Fe/S cluster binding on Grx4, while a dithiol active site results in cofactor destabilization and a moderate impairment of in vivo function. These requirements for Fe/S cofactor stabilization on Grx4 are virtually the opposite of those previously reported for single domain Grxs. Grx4 functions as iron sensor for the iron-sensing transcription factor Aft1 in S. cerevisiae. We found that Aft1 binds to a conserved binding site at the C-terminus of Grx4. This interaction is essential for the regulation of Aft1. Collectively, our analysis demonstrates that the multidomain monothiol Grxs form a unique protein family distinct from that of the single domain Grxs.

The Monothiol Glutaredoxin Grx4 Exerts an Iron-Dependent Inhibitory Effect on Php4 Function

“…Mutational analysis of the Cys35 residue located within the TRX domain of Grx4 revealed that it is required for the establishment of a strong, iron-independent association with Fep1 (17). Consistently, a recent study of S. cerevisiae suggested that the TRX domain may serve as a docking site for the interacting partners of the multidomain monothiol glutaredoxins Grx3 and Grx4 (12). In baker's yeast, other functions for the TRX domain have been proposed, including a role in the targeting of the monothiol glutaredoxin Grx3 to the nucleus (31), as well as a regulatory role in actin cytoskeleton remodeling and a role in cellular defense against oxidative stress (40).…”

Structural and Spectroscopic Insights into BolA-Glutaredoxin Complexes

“…Importantly, the recognition of Aft1 involves at least a negatively charged region in the yeast Grx4 formed by the last 16 C-terminal residues (supplemental Fig. S2) (40). Overall, although it is not clear where the Grx-BolA complex is acting, it is very likely that the formation of the Fe-S cluster in the Grx-BolA heterodimer is crucial for Aft1 regulation.…”

Mono- and Dithiol Glutaredoxins in the Trypanothione-Based Redox Metabolism of Pathogenic Trypanosomes

“…2). The latter proteins occur almost exclusively in eukaryotes and represent a distinct functional group (35).…”

“…Instead, the capability to coordinate ISCs appears to be a common feature for 1-C-Grxs (16,40,42,43,54,67,89) with yeast Grx7 being, so far, the only known exception (59,60). The latter feature determines an evolutionary conserved and indispensable role of 1-C-Grxs in the biogenesis and assembly of iron-sulfur proteins (11,61) and other cell-specific regulatory functions such as the (in)activation of nuclear transcription factors (35,62,86).…”

“…The mechanism includes the ISC-mediated interaction of Grx3 and Grx4 with transcription factors that, depending on the iron levels, regulate operons controlling iron usage or uptake (35,62). It seems unlikely that the parasite 1-C-Grx3 plays a similar role, owing to the fact that transcription factors are almost absent in trypanosomes and gene expression is mainly controlled at post-transcriptional level (14).…”