Abstract
Peroxiredoxins (PRDXs) are antioxidant enzymes that protect cells from oxidative stress and play a role in reactive oxygen species (ROS)-mediated signaling. We reported that PRDXs are critical for human fertility by maintaining sperm viability and regulating ROS levels during capacitation. Moreover, studies on Prdx6−/− mice revealed the essential role of PRDX6 in the viability, motility, and fertility competence of spermatozoa. Although PRDXs are abundant in the testis and spermatozoa, their potential role at different phases of spermatogenesis and in perinatal germ cells is unknown. Here, we examined the expression and role of PRDXs in isolated rat neonatal gonocytes, the precursors of spermatogonia, including spermatogonial stem cells. Gene array, qPCR analyses showed that PRDX1, 2, 3, 5, and 6 transcripts are among the most abundant antioxidant genes in postnatal day (PND) 3 gonocytes, while immunofluorescence confirmed the expression of PRDX1, 2, and 6 proteins. The role of PRDXs in gonocyte viability was examined using PRDX inhibitors, revealing that the 2-Cys PRDXs and PRDX6 peroxidases activities are critical for gonocytes viability in basal condition, likely preventing an excessive accumulation of endogenous ROS in the cells. In contrast to its crucial role in spermatozoa, PRDX6 independent phospholipase A2 (iPLA2) activity was not critical in gonocytes in basal conditions. However, under conditions of H2O2-induced oxidative stress, all these enzymatic activities were critical to maintain gonocyte viability. The inhibition of PRDXs promoted a two-fold increase in lipid peroxidation and prevented gonocyte differentiation. These results suggest that ROS are produced in neonatal gonocytes, where they are maintained by PRDXs at levels that are non-toxic and permissive for cell differentiation. These findings show that PRDXs play a major role in the antioxidant machinery of gonocytes, to maintain cell viability and allow for differentiation.
Highlights
Peroxiredoxins are found in all living organisms, from bacteria, plants, yeasts to animals, where they act as scavengers of hydrogen peroxide (H2 O2 ), lipid peroxides and peroxynitrite
The present study demonstrates that PRDXs are essential for maintaining reactive oxygen species (ROS) homeostasis and cell viability in neonatal gonocytes, and that the independent phospholipase A2 (iPLA2) activity of PRDX6 in these cells is not as critical as it is in spermatozoa, suggesting differential role for these antioxidant enzymes at different phases of germ cell development
In view of the abundance of PRDXs in gonocytes, and knowing the importance of these peroxidases for spermatozoa viability and function, we examined whether inhibiting PRDXs would have an impact on the survival of PND3 gonocytes
Summary
Peroxiredoxins are found in all living organisms, from bacteria, plants, yeasts to animals, where they act as scavengers of hydrogen peroxide (H2 O2 ), lipid peroxides and peroxynitrite. PRDXs are classified depending on the cysteine residues (Cys) in their active site, that will react with peroxides. They comprise the 2-Cys PRDX1 to 4, the atypical 2-Cys PRDX5, and the 1-Cys PRDX6, which has the particularity of being bifunctional, with both peroxidase and calcium-independent phospholipase A2 (iPLA2 ) activities [4]. The 2-Cys PRDX1 to 4 are homodimers in which the thiol of a cysteine residue of one PRDX subunit gets oxidized, further reacts with the thiol group of the catalytic cysteine of the other subunit, forming a disulfide bond between the two subunits. In the atypical PRDX5, 2 Cys of the same chain react upon oxidation to form an intrasubunit disulfide bond. In the case of PRDX6, since the enzyme has only one catalytic Cys, the oxidized thiol will be reduced by the glutathione-GSH-transferase P1 (GSTP1) system [4,5,6]
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