Abstract
The process of spermatogenesis is complex and systemic, requiring the cooperation of many regulators. However, little is known about how micro RNAs (miRNAs) regulate spermatogenesis in poultry. In this study, we investigated key miRNAs and their target genes that are involved in spermatogenesis in chickens. Next-generation sequencing was conducted to determine miRNA expression profiles in five cell types: primordial germ cells (PGCs), spermatogonial stem cells (SSCs), spermatogonia (Spa), and chicken sperm. Next, we analyzed and identified several key miRNAs that regulate spermatogenesis in the four germline cell miRNA profiles. Among the enriched miRNAs, miRNA-301a-5p was the key miRNA in PGCs, SSCs, and Spa. Through reverse transcription quantitative PCR (RT-qPCR), dual-luciferase, and miRNA salience, we confirmed that miR-301a-5p binds to transforming growth factor-beta 2 (TGFβ2) and is involved in the transforming growth factor-beta (TGF-β) signaling pathway and germ cell development. To the best of our knowledge, this is the first demonstration of miR-301a-5p involvement in spermatogenesis by direct binding to TGFβ2, a key gene in the TGF-β signaling pathway. This finding contributes to the insights into the molecular mechanism through which miRNAs regulate germline cell differentiation and spermatogenesis in chickens.
Highlights
Spermatogenesis is a complex and asynchronous germ cell formation process that includes primordial germ cells that develop through several immature stages by undergoing mitosis, meiosis, and differentiation into haploid spermatozoa and mature sperm [1,2,3]
Recent studies on spermatogenesis have demonstrated that micro RNAs (miRNAs) dysregulation has been implicated in male fertility, leading to sperm abnormality and spermatogenesis disruption, suggesting that miRNAs are functionally important in the spermatogenesis process in mammalian and poultry [13,14,15,17,18]
Using target gene prediction in three databases, we found that TGFB2 was among the target genes of miR-301a-5p
Summary
Spermatogenesis is a complex and asynchronous germ cell formation process that includes primordial germ cells that develop through several immature stages by undergoing mitosis, meiosis, and differentiation into haploid spermatozoa and mature sperm [1,2,3]. Many studies have shown that several regulator types which include methylation, ubiquitination, acetylation, transcription factor and et al, play major roles in germline cell development and spermatogenesis [7,8,9,10,11,12]. Few studies have reported the regulation of the TGF-β pathway in spermatogenesis by miRNAs. In the present study, we sequenced the small RNA of four cell types PGCs, SSCs, Spas, and sperm in the chicken and filtered the specific miRNAs of each cell type. We want to propose an epigenetic explanation for this process
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