Expression, localization, and concentration of A-kinase anchor protein 4 (AKAP4) and its precursor (proAKAP4) in equine semen: Promising marker correlated to the total and progressive motility in thawed spermatozoa

A-kinase anchor protein 4 (AKAP4) is playing a central role in flagellar structure, chemotaxis, capacitation and sperm motility. In mammals, AKAP4 is expressed during spermatogenesis. AKAP4 is synthesized as a precursor, proAKAP4, which is cleaved into mature AKAP4 during fibrous sheath assembly. The proAKAP4 is a good indicator of sperm quality in humans and boars. The aims of this work were to study the expression, the localization and the concentration of proAKAP4 and AKAP4 in equine semen, and to evaluate the possible correlation between the total and progressive motility and the concentration of proAKAP4 measured by ELISA in post-thawed semen. Frozen sperm from 13 different stallions were used. Semen samples (n = 17) were prepared using the INRA Freeze medium to reach a concentration of 150 million spermatozoa/mL, packaged and frozen in 0.5 mL straws. The precursor proAKAP4 and the mature protein AKAP4 both localize to the fibrous sheath of the principle piece of equine sperm flagellum. The concentrations of proAKAP4 were determined in the post-thawed semen using ELISA method (Horse 4MID^® kits, 4BioDx, France). The mean concentration of proAKAP4 was then of 7.372 ± 0.79 ng/μL and was significantly correlated with the post-thawed total motility (Pearson coefficient r = 0.66, p = 0.002) and progressive motility (Pearson coefficient r = 0.76, p = 0.0002) and the amount of proAKAP4 represent the amount of spermatozoa that expressed proAKAP4. Taken together, these preliminary results confirm the interest to use proAKAP4 concentrations as a promising marker of stallion sperm quality as close correlation was observed between the proAKAP4 concentration and sperm motility parameters.