Limits...
Epididymis response partly compensates for spermatozoa oxidative defects in snGPx4 and GPx5 double mutant mice.

Noblanc A, Peltier M, Damon-Soubeyrand C, Kerchkove N, Chabory E, Vernet P, Saez F, Cadet R, Janny L, Pons-Rejraji H, Conrad M, Drevet JR, Kocer A - PLoS ONE (2012)

Bottom Line: We show that to counteract the GPx activity losses, the epididymis of the double KO animals mounted an antioxydant response resulting in a strong increase in the global H(2)O(2)-scavenger activity especially in the cauda epididymis.Quantitative RT-PCR data show that together with the up-regulation of epididymal scavengers (of the thioredoxin/peroxiredoxin system as well as glutathione-S-transferases) the epididymis of double mutant animals increased the expression of several disulfide isomerases in an attempt to recover normal disulfide-bridging activity.Despite these compensatory mechanisms cauda-stored spermatozoa of double mutant animals show high levels of DNA oxidation, increased fragmentation and greater susceptibility to nuclear decondensation.

View Article: PubMed Central - PubMed

Affiliation: Genetics Reproduction & Development laboratory, CNRS UMR 6293 - INSERM U1103 - Clermont Université, Aubière, France.

ABSTRACT
We report here that spermatozoa of mice lacking both the sperm nucleus glutathione peroxidase 4 (snGPx4) and the epididymal glutathione peroxidase 5 (GPx5) activities display sperm nucleus structural abnormalities including delayed and defective nuclear compaction, nuclear instability and DNA damage. We show that to counteract the GPx activity losses, the epididymis of the double KO animals mounted an antioxydant response resulting in a strong increase in the global H(2)O(2)-scavenger activity especially in the cauda epididymis. Quantitative RT-PCR data show that together with the up-regulation of epididymal scavengers (of the thioredoxin/peroxiredoxin system as well as glutathione-S-transferases) the epididymis of double mutant animals increased the expression of several disulfide isomerases in an attempt to recover normal disulfide-bridging activity. Despite these compensatory mechanisms cauda-stored spermatozoa of double mutant animals show high levels of DNA oxidation, increased fragmentation and greater susceptibility to nuclear decondensation. Nevertheless, the enzymatic epididymal salvage response is sufficient to maintain full fertility of double KO males whatever their age, crossed with young WT female mice.

Show MeSH

Related in: MedlinePlus

Generation of sngpx4;gpx5-deficient mice.A and B: Diagramatic representations of mouse GPx4 (A) and GPx5 (B) gene organization (upper diagrams in A and B) and of their engineered version found in the single KO animals (lower schemes in A and B) (1, 20). Transgenic sngpx4−/− animals bear an EGFP reporter cassette (20), while transgenic gpx5−/− animals carry a deleted exon 2 (1). Grey boxes (1,a and 2 to 7 for sngpx4 and 1 to 5 for gpx5) indicate the 8 or 5 coding exons, respectively. Bold arrowheads indicate the relative positions of the various primers used thereafter in genotyping PCR amplifications. C: Typical PCR experiments carried out on genomic DNA extracted from animal fingers used to select homozygous sngpx4;gpx5−/− (DKO) animals. Size (in bp) of the expected PCR products for each primer pair used is given in the left margin. The two upper panels show sngpx4 amplifications, while the two lower panels show gpx5 amplifications. Lane 4 (DKO) stands for double knock-out animals, while lane 5 is a negative control in which no genomic DNA was added to the reactions. Primer sequences are given in Table 2. Lanes 1, 2 and 3 are positive control amplifications carried out with genomic DNA from wild type, homozygous sngpx4−/− and homozygous gpx5−/− animals, respectively.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3375294&req=5

pone-0038565-g001: Generation of sngpx4;gpx5-deficient mice.A and B: Diagramatic representations of mouse GPx4 (A) and GPx5 (B) gene organization (upper diagrams in A and B) and of their engineered version found in the single KO animals (lower schemes in A and B) (1, 20). Transgenic sngpx4−/− animals bear an EGFP reporter cassette (20), while transgenic gpx5−/− animals carry a deleted exon 2 (1). Grey boxes (1,a and 2 to 7 for sngpx4 and 1 to 5 for gpx5) indicate the 8 or 5 coding exons, respectively. Bold arrowheads indicate the relative positions of the various primers used thereafter in genotyping PCR amplifications. C: Typical PCR experiments carried out on genomic DNA extracted from animal fingers used to select homozygous sngpx4;gpx5−/− (DKO) animals. Size (in bp) of the expected PCR products for each primer pair used is given in the left margin. The two upper panels show sngpx4 amplifications, while the two lower panels show gpx5 amplifications. Lane 4 (DKO) stands for double knock-out animals, while lane 5 is a negative control in which no genomic DNA was added to the reactions. Primer sequences are given in Table 2. Lanes 1, 2 and 3 are positive control amplifications carried out with genomic DNA from wild type, homozygous sngpx4−/− and homozygous gpx5−/− animals, respectively.

Mentions: Null mice for snGPx4 and GPx5 were generated following crosses of inbred sngpx4−/− animals [24] and gpx5−/− animals [26] that were initially produced in the same C57bl/6 genetic background. Genotyping data shown in Figure 1 illustrate that the homozygous derived sngpx4;gpx5−/− animals carry the transgenic constructs and, therefore, are devoid of functional snGPx4 and GPx5 genes. As for each single KO, simultaneous sngpx4 and gpx5 inactivation had no impact on the organization of the epididymis tissue (as evidenced by histological observation) or caput or cauda sperm counts (monitored at 4 months of age), suggesting that lack of snGPx4 and GPx5 expression does not affect sperm production and epididymal transit and function (not shown).


Epididymis response partly compensates for spermatozoa oxidative defects in snGPx4 and GPx5 double mutant mice.

Noblanc A, Peltier M, Damon-Soubeyrand C, Kerchkove N, Chabory E, Vernet P, Saez F, Cadet R, Janny L, Pons-Rejraji H, Conrad M, Drevet JR, Kocer A - PLoS ONE (2012)

Generation of sngpx4;gpx5-deficient mice.A and B: Diagramatic representations of mouse GPx4 (A) and GPx5 (B) gene organization (upper diagrams in A and B) and of their engineered version found in the single KO animals (lower schemes in A and B) (1, 20). Transgenic sngpx4−/− animals bear an EGFP reporter cassette (20), while transgenic gpx5−/− animals carry a deleted exon 2 (1). Grey boxes (1,a and 2 to 7 for sngpx4 and 1 to 5 for gpx5) indicate the 8 or 5 coding exons, respectively. Bold arrowheads indicate the relative positions of the various primers used thereafter in genotyping PCR amplifications. C: Typical PCR experiments carried out on genomic DNA extracted from animal fingers used to select homozygous sngpx4;gpx5−/− (DKO) animals. Size (in bp) of the expected PCR products for each primer pair used is given in the left margin. The two upper panels show sngpx4 amplifications, while the two lower panels show gpx5 amplifications. Lane 4 (DKO) stands for double knock-out animals, while lane 5 is a negative control in which no genomic DNA was added to the reactions. Primer sequences are given in Table 2. Lanes 1, 2 and 3 are positive control amplifications carried out with genomic DNA from wild type, homozygous sngpx4−/− and homozygous gpx5−/− animals, respectively.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3375294&req=5

pone-0038565-g001: Generation of sngpx4;gpx5-deficient mice.A and B: Diagramatic representations of mouse GPx4 (A) and GPx5 (B) gene organization (upper diagrams in A and B) and of their engineered version found in the single KO animals (lower schemes in A and B) (1, 20). Transgenic sngpx4−/− animals bear an EGFP reporter cassette (20), while transgenic gpx5−/− animals carry a deleted exon 2 (1). Grey boxes (1,a and 2 to 7 for sngpx4 and 1 to 5 for gpx5) indicate the 8 or 5 coding exons, respectively. Bold arrowheads indicate the relative positions of the various primers used thereafter in genotyping PCR amplifications. C: Typical PCR experiments carried out on genomic DNA extracted from animal fingers used to select homozygous sngpx4;gpx5−/− (DKO) animals. Size (in bp) of the expected PCR products for each primer pair used is given in the left margin. The two upper panels show sngpx4 amplifications, while the two lower panels show gpx5 amplifications. Lane 4 (DKO) stands for double knock-out animals, while lane 5 is a negative control in which no genomic DNA was added to the reactions. Primer sequences are given in Table 2. Lanes 1, 2 and 3 are positive control amplifications carried out with genomic DNA from wild type, homozygous sngpx4−/− and homozygous gpx5−/− animals, respectively.
Mentions: Null mice for snGPx4 and GPx5 were generated following crosses of inbred sngpx4−/− animals [24] and gpx5−/− animals [26] that were initially produced in the same C57bl/6 genetic background. Genotyping data shown in Figure 1 illustrate that the homozygous derived sngpx4;gpx5−/− animals carry the transgenic constructs and, therefore, are devoid of functional snGPx4 and GPx5 genes. As for each single KO, simultaneous sngpx4 and gpx5 inactivation had no impact on the organization of the epididymis tissue (as evidenced by histological observation) or caput or cauda sperm counts (monitored at 4 months of age), suggesting that lack of snGPx4 and GPx5 expression does not affect sperm production and epididymal transit and function (not shown).

Bottom Line: We show that to counteract the GPx activity losses, the epididymis of the double KO animals mounted an antioxydant response resulting in a strong increase in the global H(2)O(2)-scavenger activity especially in the cauda epididymis.Quantitative RT-PCR data show that together with the up-regulation of epididymal scavengers (of the thioredoxin/peroxiredoxin system as well as glutathione-S-transferases) the epididymis of double mutant animals increased the expression of several disulfide isomerases in an attempt to recover normal disulfide-bridging activity.Despite these compensatory mechanisms cauda-stored spermatozoa of double mutant animals show high levels of DNA oxidation, increased fragmentation and greater susceptibility to nuclear decondensation.

View Article: PubMed Central - PubMed

Affiliation: Genetics Reproduction & Development laboratory, CNRS UMR 6293 - INSERM U1103 - Clermont Université, Aubière, France.

ABSTRACT
We report here that spermatozoa of mice lacking both the sperm nucleus glutathione peroxidase 4 (snGPx4) and the epididymal glutathione peroxidase 5 (GPx5) activities display sperm nucleus structural abnormalities including delayed and defective nuclear compaction, nuclear instability and DNA damage. We show that to counteract the GPx activity losses, the epididymis of the double KO animals mounted an antioxydant response resulting in a strong increase in the global H(2)O(2)-scavenger activity especially in the cauda epididymis. Quantitative RT-PCR data show that together with the up-regulation of epididymal scavengers (of the thioredoxin/peroxiredoxin system as well as glutathione-S-transferases) the epididymis of double mutant animals increased the expression of several disulfide isomerases in an attempt to recover normal disulfide-bridging activity. Despite these compensatory mechanisms cauda-stored spermatozoa of double mutant animals show high levels of DNA oxidation, increased fragmentation and greater susceptibility to nuclear decondensation. Nevertheless, the enzymatic epididymal salvage response is sufficient to maintain full fertility of double KO males whatever their age, crossed with young WT female mice.

Show MeSH
Related in: MedlinePlus