Functional insights into the role of seminal plasma proteins on sperm motility of buffalo

Abstract

The objective of the present study was to describe the proteins from the seminal plasma of buffalo and correlate these proteins with sperm motility. Ejaculates from sixteen Murrah buffalo were used. Semen collection was performed by electroejaculation, and the ejaculate was evaluated by macroscopic (volume) and microscopic analysis (subjective motility and vigor, as well as sperm concentration). After the analysis, the samples were centrifuged (800g for 10 min and 10,000 for 30 min at 4 °C), and the supernatant (seminal plasma) was used to determine total protein concentration by the Bradford method. Based on total protein concentration, an aliquot (50 μg) was taken to conduct protein in-solution digestion for nano-LC-ESI-Q-TOF mass spectrometry analysis. Samples were divided into two groups, minimal (little sperm motility) and greater (typical sperm motility), based on non-hierarchical clustering considering motility and emPAI protein value. The data were analyzed by multivariate statistical analysis using principal component analysis (PCA) and partial analysis of minimum squares discrimination (PLS-DA). Forty-eight proteins were detected in the seminal plasma, and fifteen were common to two groups. There were six proteins that were significantly different between the groups. The main functions of proteins in seminal plasma were catalytic and binding activity. Spermadhesin protein, ribonuclease, 14-3-3 protein zeta/delta and acrosin inhibitor were in greater amounts in seminal plasma from the group with greater sperm motility; prosaposin and peptide YY were in greater amounts in the group with little sperm motility. The proteins detected in the greater motility group were correlated with sperm protection, including protection against oxidative stress, lipid peroxidation, protease inhibition and prevention of premature capacitation and acrosome reaction. In the group with little sperm motility, one of the identified proteins is considered to be an antifertility factor, whereas the function of other identified protein is not definitive. Results from the present study add to the knowledge base about the molecular processes related with sperm motility, and these findings can be used for determining potential markers of semen quality.

Cross comparison of seminal plasma proteins from cattle and buffalo (Bubalus bubalis)

“…This blocking action inhibits sperm motility, which delays the acrosome reaction and compromises fertilization (Shivaji & Bhargava, ). In our study, both prosaposin and peptide YY were upregulated in the buffalo, which can be compensatory response since it was also correlated with decreased motility in this species (Codognoto et al, ), although in bovine prosaposin was related to a higher pregnancy rate (Amann, Seidel, et al, ).…”

“…In our study, it was present in both groups, with over‐expression in the cattle group. It has been described in the seminal plasma of buffaloes submitted to electroejaculation and was correlated with high sperm quality (Codognoto et al, ). Spermadhesin in decreased abundance in the buffaloes can indicate lower sperm cryoresistance, since this protein is involved in the inhibition of oxidative stress and decreasing lipid peroxidation (Jobim et al, ), which are important for maintaining sperm quality during and after cryopreservation.…”

“…Acrosin inhibitor is produced by accessory sexual glands and secreted in the seminal plasma of ruminants (Cechová & Fritz, 1; Cechová, Jonáková, Sedláková, & Mach, ; Codognoto et al, ; Davidová, Jonáková, & Manásková‐Postlerová, ; Martins et al, ; Torska & Strzezek, ; Tschesche, Kupfer, Klauser, Fink, & Fritz, ), and it functions to prevent early sperm proteolysis induced by acrosin, an acrosomal enzyme responsible for zona pellucida hydrolysis during acrosomal reaction and fertilization (Adham, Nayernia, & Enzel, ; Crosby, Jones, Barros, & Carvalho, ; Gurupryaia et al, ; Martins et al, ). Acrosin inhibitor and acrosin are proteins related to in vivo and in vitro fertility in humans (Goodpasture, Zavos, Cohen, & Zaneveld, ; Kennedy, Reid, Kaminski, Jeyendran, & Zaneveld, ; Mohsenian, Syner, & Moghissi, ) and chickens (Thélie et al, ).…”

“…Seminal plasma proteins were digested in solution according to Villen and Gigy, () with modifications, as described by Codognoto et al (). Briefly, an aliquot (50 μg) was separated and added to 8 M urea, reduced with 5 mM DTT and alkylated with 14 mM iodoacetamide.…”

“…Proteomics is a technique to characterize proteins of seminal plasma that can be correlated with essential sperm cell functions (Aslam et al, ), since several proteins of this fluid are involved in sperm capacitation (Manjunath & Thérien, ; Thérien, Moreau, & Manjunath, ; Thérien, Soubeyrand, & Manjunath, ) acrosome reactions (Souza, Moura, Lima‐Souza, & Killian, ; Thérien et al, ), sperm–oocyte binding (Erikson, Way, Chapman, & Killian, ; Gonçalves, Staros, & Killian, ) and fertilization (Manjunath & Thérien, ; Moura, Chapman, Koc, & Killian, ; Thérien et al, ), in addition to acting in sperm protection (Moura et al, ). Therefore, the proteins are related to fertility (Asadpour et al, ; Baruti et al, ; Choudhary, Kavya, Jerome, & Sharma, ; Dar et al, ; Killian, Chapman, & Rogowski, ) and can be used as biomarkers in buffaloes (Codognoto et al, ; Ramteke et al, ; Selvaraju et al, ). Then, the aim of this study was to compare seminal plasma proteins from cattle and buffalo ( Bubalus bubalis ).…”