Proteome Analysis Of Sperm Between High-and Low-motility In Hu Sheep Using TMT Approach

Fertilization of eggs by spermatozoa sets the stage for mammalian development,and viable sperm are a prerequisite for successful fertilization and embryo development.Motility is a key element of good quality sperm cells,as they need to pass through the female reproductive tract to the site of fertilization.Sperm motility is very important for animal reproduction.Scholars have also done a lot of research on sperm motility and found some genes and proteins related to sperm motility,but there is less research on spermatozoa of Hu sheep.In this study,we used the mini-Percoll density gradient centrifugation method to separate Hu sheep ejaculation,and high-motility group sperm and low-motility group sperm were obtained.And the quantitative proteomics of TMT marker technique was used to quantitatively analyze the proteome of Hu sheep,and the biological analysis of the identified protein was carried out.This study provides valuable information for further exploring the mechanism of the the biological markers of protein related to sperm motility.The result shows that:1)In this study,mini-Percoll density gradient centrifugation was used to separate Hu sheep fresh essence.The total motility rate of high-motility group and low-motility group was 75.7% and 28.0%,there were extremely significant differences between high-motility group and low-motility group.The progressive motility rate of high-motility group and low-motility group was 28.0% and 1.7%,there were extremely significant differences between high-motility group and low-motility group.2)A totle of 1,727 proteins were identified in Hu sheep high-and low-motility sperm group,including 1,554 proteins were quantified.Among these quantified proteins,there are 97 differentially expressed proteins,5 proteins were up-regulated in the high-motility group,while 95 proteins expression increased in the low-motility group.Proteins up-regulated in the high-motility group were: PEBP4,SPATA18,GPD1,CPVL,FAM161 A.3)The differentially expressed proteins were analyzed by GO annotation,in the biological process,most of identified proteins involved in protein folding,cell redox homeostasis,mRNA splicing,via spliceosome and spermatid development;in the cellular components,most of the proteins were located in the extracellular exosome,membrane and nucleoplasm;most of identified proteins involved in poly(A)RNA binding,nucleotide binding,mRNA binding,and translation elongation factor activity.4)KEGG pathway analysis revealed that differentially expressed proteins mainly involved in Spliceosome,Protein processing in endoplasmic reticulum,and Ribosome.In this study,we have analyzed the differential proteome analysis of sperm between high-and low-motility in Hu sheep.The results provide a reference for discovering the biomarkers of the Hu sheep sperm motility and exploring the molecular mechanism of the sperm motility.