Proteomic Study On Spermation Of Mouse Testes

N-glycosylation is one of the most important and common post-translational modification in vivo. It plays important roles in many biological processes, including protein folding and protein localting. Although N-glycosylation is known to be important for spermatogenesis and fertilization, little is known about the composition and structure of N-linked glycoproteins, especially the glycosylation sites of these proteins on human testis. Testis, as an important endocrine organ, mainly produces sperm and androgen. The complex structure of the testis and its important role in the male reproductive system have been demonstrated that it has important impact on the research of male fertility. On the other hand, proteomics technology has been widely used in the qualitative and quantitative research of the whole proteins of the tissue and cell level; in recent years, Max Planck laboratory’s used lectin to enrich glycosylated protein and advanced mass spectrometry to identify glycosylated protein. In the same time deglycosylation is performed in 180-water, which makes that proteomics can be used for the system and comprehensive analysis of the specific modification sites in glycoproteins.At present, there is no research about mouse testis glycoproteome to be reported. In present study, we used glycopeptide enrichment technology with tandem mass spectrometry(MS/MS) method to establish mouse testis glycoproteome, and identified 1463 N-glycosylation sites and 740 N-glycosylated protein groups. We interpreted the sequence features of these sites and the function and regulatory pathways of the N-glycoproteins of these sites through bioinformatics methods. We found that in addition to the NXS and NXT motifs, the most enriched motif in the mouse testis N-glycosylation sites was NCX. Go and KEGG functional enrichment analysis revealed these proteins were mainly localized in cell membrane, lysosome, golgi apparatus and other parts; most of these N-glycoproteins had hydrolase and aminopeptidase activity, mainly involved in cell adhesion, protein hydrolysis, protein positioning and other events. Finally, we further screened those non-secretory and testis specific glycoproteins to analyse in detail, and the complex interaction and gene regulatory networks were constructed, these glycoproteins were found to closely related to the process of spermatogenesis. Our study will provide a theoretical basis for the research on the molecular mechanism of the role of glycoproteins in spermatogenesis.