15 SPERM STORAGE IN FEMALE REPRODUCTIVE TRACT: STUDY OF MOLECULES INVOLVED

Abstract

Because of prolonged sperm storage in their oviduct, domestic hens can produce fertile eggs for up to 3 weeks following a single AI. The oviduct secretions may have an effect on sperm survival, but its composition during fertilization is unknown. In the present study, we compared the proteomic content of uterine fluid collected from two lines of hens divergent by their duration of fertility period (DFP), which defined sperm-storage duration. The first line displays a shorter period of sperm storage (10 days, line DFP–), whereas the second displays a longer period of sperm storage (21 days, DFP+). The aim was to identify proteins or peptides that may be involved in spermatozoa survival. Uterine fluid was collected 10 h after oviposition either before (n = 5/line) or 24 h after (n = 5/line) AI. Samples were pooled by condition: DFP+ before AI, DFP+ after AI, DFP– before AI, and DFP– after AI. Bottom-up approach using sodium dodecyl sulfate-polyacrylamide gel electrophoresis and nano LC-MS/MS was performed (3 replicates). Data were matched against the NCBInr database (2014) using Mascot, and identifications were validated by the peptide and protein Prophet algorithm using Scaffold software. To determine the differences in protein expression, spectral counting and XIC quantitative methods were employed using Scaffold Q+ (P < 0.05, ratio > 2). Two proteins were up-regulated and one was down-regulated in oviductal secretion of both lines in response to AI. However, AI induced a significantly different abundance between protein content of DFP– and DFP+ fluids. A panel of 8 proteins, included one DFP+-specific protein, was more abundant in DFP+ line than in DFP–. Only one protein was less abundant in DFP+ line than in DFP–. In conclusion, the presence of sperm in the genital tract induced quantitative differences of the protein content of the uterine fluid in DFP– and DFP+ hen lines. These differences imply proteins that are known as male proteins (sperm, seminal plasma, testis). Analysis of sperm protein modifications after storage will help us to understand the functional implication of these candidates.