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Project

The Peroxisomal Redox Orchestra: New Melodies of Old Molecular Players

Eukaryotic life, viewed from a physicochemical perspective, is a remarkable phenomenon that defies all odds, as it emerges solely through the occurrence of seemingly unfavorable chemical reactions over the course of evolution. However, this fascinating concept comes with a cost: susceptibility to environmental fluctuations. As a result, cells must continually strive to reestablish and uphold equilibrium. A paradigmatic example of this is the crucial internal balance between electrophiles and nucleophiles, commonly referred to as redox homeostasis.

Peroxisomes play a pivotal role in preserving cellular redox balance, particularly through their involvement in hydrogen peroxide metabolism. Yet, our knowledge of the molecular mechanisms underlying peroxisome-mediated redox events remains limited. This project aimed to acquire deeper insights into these processes, specifically by improving our understanding of (i) glutathione metabolism within peroxisomes, and (ii) the role of putative redox metabolite transporters residing in the peroxisomal membrane in maintaining cellular redox equilibrium, with a focus on peroxisomes.

In our study of peroxisomal glutathione metabolism, we generated a cellular knockout model for GSTK1, the sole known glutathione-consuming in mammalian peroxisomes. We observed that, following oxidative insults, the recovery of roGFP2, a redox-sensitive fluorescent sensor that equilibrates with the glutathione redox couple, was significantly slower in ΔGSTK1 cells. Since the presence of glutaredoxin-1 in peroxisomes rescued this phenotype, we concluded that GSTK1 exhibits disulfide bond oxidoreductase activity.

Next, our focus shifted to investigating the transport of redox-active molecules across the peroxisomal membrane. Initially, we employed overexpression models to study the localization of a selected group of cofactor transporters previously described in other biological membranes, in order to determine whether they also displayed a peroxisomal localization. However, none of the investigated proteins co-localized with the peroxisomal markers.

We then directed our efforts to comprehend the roles of two peroxisomal membrane proteins, SLC25A17 and PXMP4, whose functions have not yet been fully characterized. Using cellular knockout models, we found that SLC25A17 inactivation resulted in a decrease of the overall peroxisomal redox state and disrupted the interconnection between the peroxisomal and cytosolic pools of NADPH, demonstrating that SLC25A17 plays an important role in preserving peroxisomal redox homeostasis. Additionally, we obtained evidence that PXMP4 inactivation led to reduced cytosolic H2O2 levels, suggesting a role for this protein in cellular redox maintenance. Lastly, in our SLC25A17 studies, we demonstrated that the peroxisomal and cytosolic glutathione pools are maintained independently, challenging the recent view that the peroxisomal membrane is promptly permeable to reduced and oxidized glutathione.

In summary, our work reinforces the previously observed intricate interplay between cellular and peroxisomal redox metabolism. In addition, it reveals that defects in not-yet fully functionally characterized peroxisomal membrane transporters have the potential to contribute to intracellular redox imbalances. Considering that disruptions in the intracellular redox status are closely tied to both cellular health and disease, our findings may serve as a catalyst for inspiring others to explore innovative avenues to tackle these conditions, potentially uncovering new preventive and therapeutic strategies.

Date:1 Sep 2019 →  6 Dec 2023
Keywords:peroxisomes, hydrogen peroxide, membrane contact sites, redox signaling, glutathione redox state, oxidative stress
Disciplines:Cell signalling
Project type:PhD project