Male Fertility Potential Molecular Mechanisms Revealed by iTRAQ-Based Quantitative Proteomic Analysis of the Epididymis from Wip1-/- Mice

Proteomics and postgenomic technologies have found growing recent applications in reproductive biology and fertility research. Mice deficient in the wild-type p53-induced phosphatase 1 (WIP1) exhibit defects in reproductive organs. In this study, we aimed to understand how Wip1 deficiency affects spermatogenesis and sperm maturation using the Wip1(-/-) mouse model and gel-free isobaric tags for relative and absolute quantitation liquid chromatography-tandem mass spectrometry quantitative proteomic analysis of the whole epididymis, including somatic tissue and sperm. A total of 8763 proteins were identified, of which 91 were significantly differentially expressed proteins (DEPs) in Wip1(-/-) mice. Of note, four reproduction-related DEPs (PRM2, ODF1, PIWIL1, and KLHL10) were confirmed by western blotting. Pathway analysis suggested that the Smac/Diablo-mediated apoptotic pathway and the SERPINA3-mediated inflammatory process may contribute to the atrophy and marked reduction of sperm in the epididymis. Network analysis of reproduction-related DEPs revealed possible interactions of WIP1 that may affect sperm maturation, such as reduced ODF1 and PRM2 expression and increased PIWIL1 expression by p53. Histological analysis showed a spermatid deficiency in the epididymis and was further confirmed in testis in Wip1(-/-) mice. Immunohistochemistry showed that testicular expressions of PRM2 and PIWIL1 were down- and upregulated, respectively. In summary, WIP1 deficiency seems to cause impaired spermatogenesis in the testis and damaged sperm maturation in the epididymis. These may be attributed in part to regulation of PRM2, ODF1, PIWIL1, and associated pathways as well as the inflammatory and apoptotic pathways. Further preclinical and clinical studies of male fertility using proteomics and multiomics research are called for.