piR-121380 Is Involved in Cryo-Capacitation and Regulates Post-Thawed Boar Sperm Quality Through Phosphorylation of ERK2 via Targeting PTPN7

Abstract

Cryopreservation induces capacitation-like (cryo-capacitation) changes, similar to natural capacitation, and affects the fertility potential of post-thawed sperm. The molecular mechanism of sperm cryo-capacitation during cryopreservation remains unknown. PIWI-interacting RNAs (piRNAs) have been reported to be involved in cryo-capacitation of post-thawed sperm and regulation of sperm motility, capacitation, and chemotaxis. In this study, protein tyrosine phosphatase nonreceptor type 7 (PTPN7) was positively targeted by piR-121380 after a dual luciferase assay. The mRNA expression of PTPN7 and piR-121380 was significantly decreased (p < 0.01); however, PTPN7 protein was significantly increased (p < 0.01) in post-thawed boar sperm. Furthermore, E1RK1/2 phosphorylation was reduced during cryopreservation. Six hours after transfection with piR-121380 mimic and inhibitor, the phosphorylation of ERK2 was significantly increased and decreased (p < 0.01), respectively. Furthermore, the highest and lowest total sperm motility, forward motility, and capacitation rate were observed after piR-121380 mimic and inhibitor treatments, respectively. The concentration of intracellular calcium ([Ca2+]i) showed no significant difference after transfection with either piR-121380 mimic or inhibitor at 1, 3, and 6 h. In conclusion, we demonstrated that piR-121380 modulates ERK2 phosphorylation by targeting PTPN7, which induces sperm cryo-capacitation, and eventually affects the motility and fertility potential of post-thawed sperm.

Cryopreservation Induces Acetylation of Metabolism-Related Proteins in Boar Sperm

“…After protein extraction and concentration measurement, Western blot analysis was performed to determine the expression of proteins in fresh and frozen sperm. Western blotting was performed as described by [ 84 ] with a few modifications described briefly as follows: protein samples were mixed with Laemmle blue (containing 1 percent BME as a reducing agent) stain in a 3:1 ratio. After gentle mixing, the samples were placed at 95 °C for 5 min.…”

The gene regulatory role of non-coding RNAs in non-obstructive azoospermia

“…Its low expression can alleviate the decrease of cell viability, affect cell cycle and reduce the rate of apoptosis ( 55 ). PiR_121380 regulates the phosphorylation of ERK2 by targeting PTPN7, which affects sperm motility and fertility potential after thawing ( 56 ).…”