Identification of proteomic markers for ram spermatozoa motility using a tandem mass tag (TMT) approach

Sperm motility is crucial for ram fertility; however, differences in the proteome of sperm with high- and low- motility in rams (Ovis aries) has yet to be achieved. Thus, the aim of this study was to identify and characterize ram spermatozoa proteins with different abundances between high- and low- motility, and identify of proteomic markers for ram spermatozoa motility using tandem mass tag (TMT) protein labeling and LC-MS/MS. In this study, the abundance of 150 proteins in high-motility (HM) ram sperm was significantly different compared with low-motility (LM) sperm. Proteins involved in sperm motility, mitochondrial activity, and spermatogenesis showed higher abundance in HM ram spermatozoa; proteins involved in protein processing and spliceosome were more abundant in LM ram spermatozoa. In conclusion, Phosphatidylethanolamine binding protein 4 is a potential proteomic marker for ram sperm motility; CCTs/HSPs are hallmarks of the LM spermatozoa in rams. Our findings highlight the functional differences between HM and LM ejaculated spermatozoa and has identified candidate proteins of interest linked to sperm motility in rams. SIGNIFICANCE: Inadequate sperm motility is one of the most important reasons for male subfertility or infertility. In this study, we found that the abundance of 150 proteins in high-motility ram sperm was significantly different compared with low-motility sperm. Proteomic biomarkers were discovered to reflect the motility variation in ram spermatozoa; these biomarkers may assist in illuminating the molecular mechanisms underlying sperm motility. This study expands the potential direction for sperm quality screening and animal breeding.