Integrative Proteomic and Phosphoproteomic Profiling of Testis from Wip1 Phosphatase-Knockout Mice: Insights into Mechanisms of Reduced Fertility*

Abstract

Mice lacking wild-type p53-induced phosphatase 1 (Wip1) display male reproductive defects including smaller testes, subfertility and spermatogenesis defects at the round- and elongating-spermatid stages. However, the molecular mechanisms underlying these abnormalities remain unclear. Here we examined the proteome and phosphoproteome of testes from Wip1-knockout mice using a quantitative proteomic approach. From a total of 6872 proteins and 4280 phosphorylation sites identified, 58 proteins and 159 phosphorylation sites were found to be differentially regulated compared with wild type mice. Pathway enrichment analyses revealed that these regulated proteins and phosphosites were mainly involved in adherens/tight junctions, apoptosis, inflammatory response, spermatogenesis, sperm motility, and cytoskeletal assembly and depolymerization. Wip1-knockout mice showed decreased expression of junction-associated proteins (occludin, ZO-1, and N-cadherin) and impaired integrity of the blood-testis barrier. In addition, Wip1 deficiency was associated with elevated levels of cytokines and germ cell apoptosis in the testis. These results suggest that proinflammatory cytokines may impair the blood-testis barrier dynamics by decreasing the expression of junction-associated proteins, which could lead to subfertility and spermatogenesis defects. Collectively, these findings help to explain the low reproductive function caused by Wip1 deletion and provide novel insights into our understanding of causes of male infertility.