Transcriptome analysis of inseminated sperm storage tubules throughout the duration of fertility

Authors: Brady, Kristen; KRASNEC, K - Former ARS Employee; Long, Julie

Submitted to: Poultry Science
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/8/2022
Publication Date: 2/6/2022
Citation: Brady, K.M., Krasnec, K., Long, J.A. 2022. Transcriptome analysis of inseminated sperm storage tubules throughout the duration of fertility. Poultry Science. 101:101704-101722. https://doi.org/10.1016/j.psj.2022.101704.
DOI: https://doi.org/10.1016/j.psj.2022.101704

Interpretive Summary: Avian sperm storage is extremely limited once semen is collected for artificial insemination but can occur for prolonged periods of time in the reproductive tract of the hen within specialized structure called sperm storage tubules, located in the uterovaginal junction portion of the oviduct. In addition, the sperm storage tubules appear to play a role in selecting only optimal sperm for storage and clearing suboptimal sperm from the reproductive tract. Despite the key role that sperm storage tubules play in determining the hen fertility, the mechanisms regulating sperm entrance, maintenance, and exit from sperm storage tubules remain largely unknown. This study used a global gene expression approach to compare gene expression profiles at five different timepoints throughout the sperm storage period to gain a deeper understanding of how sperm storage tubules function and regulate sperm dynamics. New findings related to innate and acquired immune responses, sperm mobility suppression, lipid synthesis and transfer, steroid hormone regulation, ion balance homeostasis in sperm storage tubules were obtained through this study. This study provides direction for further research aims, with the ultimate goal of exploiting the sperm storage mechanisms exhibited in sperm storage tubules to improve the capacity for semen storage prior to artificial insemination.

Technical Abstract: Sperm storage tubules (SSTs) are specialized invaginations of the oviductal epithelium that permit avian species to store spermatozoa for extended periods of time, without compromising sperm fertilization capacity. The molecular and physiological mechanisms behind sperm storage tubule differentiation, sperm protection, and regression remain largely unknown and have potential implications in drastically improving hen fertility, sperm storage, and semen cryopreservation in commercial poultry species. RNA sequencing was performed on sperm storage tubules isolated from the oviductal epithelium from inseminated hens at day 1, 7, 30, 60, and 90 post-insemination (n=4 per timepoint). Read mapping and differential expression analysis were performed using CLC Genomics Workbench. A total of 2340 differentially expressed genes were subjected to pathway analysis through Ingenuity Pathway Analysis (IPA). Through functional annotation of differentially expressed genes during early, peak, and late egg production, novel insights into the role of innate and acquired immunoresponses to sperm, lipid synthesis and transfer, steroid hormone signalling, cytoskeletal reorganization, and regulation of ion homeostasis in SSTs were obtained. Additionally, potential pathways suppressing sperm mobility were identified. Upstream analysis identified potential regulatory roles for 18 upstream regulators that could regulate sperm storage tubule function and sperm mobility reduction. Understanding sperm storage tubule function throughout the laying cycle and in terms of sperm preservation, may allow for the development of industry-based protocols for semen storage and cryopreservation that manipulate the sperm preservation capabilities of sperm storage tubules.