During epididymal transit, spermatozoa go through several functional maturation steps, resulting from interactions with epididymal secretomes specific to each region. In particular, the sperm membrane is under constant remodeling, with sequential attachment and shedding of various molecules provided by the epididymal lumen fluid and epididymosomes, which also deliver sncRNA cargo to sperm. As a result, the payload of sperm sncRNAs changes during the transit from the epididymis caput to the cauda. This work was designed to study the dynamics of cattle sperm sncRNAs from spermatogenesis to final maturation. Comprehensive catalogues of sperm sncRNAs were obtained from testicular parenchyma, epididymal caput, corpus and cauda, as well as ejaculated semen from three Holstein bulls. The primary cattle sncRNA sperm content is markedly remodeled as sperm mature along the epididymis. Expression of piRNAs, which are abundant in testis parenchyma, decreases dramatically at epididymis. Conversely, sperm progressively acquires miRNAs, rsRNAs, and tsRNAs along epididymis, with regional specificities. For instance, miRNAs and tsRNAs are enriched in epididymis cauda and ejaculated sperm, while rsRNA expression peaks at epididymis corpus. In addition, epididymis corpus contains mainly 20 nt long piRNAs, instead of 30 nt in all other locations. Beyond the bulk differences in abundance of sncRNAs classes, K-means clustering was performed to study their spatiotemporal expression profile, highlighting differences in specific sncRNAs and providing insights into their putative biological role at each maturation stage. For instance, Gene Ontology analyses using miRNA targets highlighted enriched processes such as cell cycle regulation, response to stress and ubiquitination processes in testicular parenchyma, protein metabolism in epididymal sperm, and embryonic morphogenesis in ejaculated sperm. Our findings confirm that the sperm sncRNAome does not simply reflect a legacy of spermatogenesis. Instead, sperm sncRNA expression shows a remarkable level of plasticity resulting probably from the combination of multiple factors such as loss of the cytoplasmic droplet, interaction with epididymosomes, and more surprisingly, the putative in situ production and/or modification of sncRNAs by sperm. Given the suggested role of sncRNA in epigenetic trans-generational inheritance, our detailed spatiotemporal analysis may pave the way for a study of sperm sncRNAs role in embryo development.

中文翻译：

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牛精子 sncRNA 在成熟过程中的动力学，从睾丸到射精的精子

在附睾转运过程中，精子经过几个​​功能成熟步骤，这是由于与每个区域特有的附睾分泌物相互作用而产生的。特别是，精子膜不断重塑，附睾管腔液和附睾体提供的各种分子依次附着和脱落，这些分子也将 sncRNA 货物运送到精子。因此，精子 sncRNA 的有效载荷在从附睾头到尾的运输过程中发生了变化。这项工作旨在研究牛精子 sncRNA 从精子发生到最终成熟的动力学。从睾丸实质、附睾头、体和尾部以及来自三头荷斯坦公牛的射精精液中获得了精子 sncRNA 的综合目录。随着精子沿附睾成熟，主要的牛 sncRNA 精子含量显着重塑。在睾丸实质中丰富的piRNA的表达在附睾中显着降低。相反，精子沿着附睾逐渐获得 miRNA、rsRNA 和 tsRNA，具有区域特异性。例如，miRNA 和 tsRNA 在附睾尾和射精的精子中富集，而 rsRNA 在附睾体中的表达达到峰值。此外，附睾语料库主要包含 20 nt 长的 piRNA，而不是所有其他位置的 30 nt。除了 sncRNAs 类别的大量差异之外，还进行了 K-means 聚类以研究它们的时空表达谱，突出特定 sncRNAs 的差异，并提供对其在每个成熟阶段推定的生物学作用的见解。例如，使用 miRNA 靶标的基因本体分析突出了丰富的过程，例如细胞周期调节、睾丸实质中对压力和泛素化过程的反应、附睾精子中的蛋白质代谢以及射精精子中的胚胎形态发生。我们的研究结果证实，精子 sncRNAome 不仅仅反映了精子发生的遗产。相反，精子 sncRNA 表达显示出显着水平的可塑性，这可能是由于多种因素的组合，例如细胞质液滴的丢失、与附睾的相互作用，更令人惊讶的是，精子对 sncRNA 的假定原位产生和/或修饰。鉴于 sncRNA 在表观遗传跨代遗传中的作用，