Department of Urology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, China.
Center for Reproductive Medicine, Cheeloo College of Medicine, Shandong University, Jinan, China.
Cell Cycle. 2023 Oct;22(19):2194-2209. doi: 10.1080/15384101.2023.2280170. Epub 2023 Dec 5.
Spermatozoa released from the testis cannot fertilize an egg before becoming mature and motile in the epididymis. Based on three bulk and one single-cell RNA-seq (scRNA-seq) data series, we compared mRNA or miRNA expression between epididymal segment-specific samples and the other samples. Hereby, we identified 570 differentially expressed mRNAs (DE-mRNAs) and 23 differentially expressed miRNAs (DE-miRNAs) in the caput, 175 DE-mRNAs and 15 DE-miRNAs in the corpus, 946 DE-mRNAs and 12 DE-miRNAs in the cauda. In accordance with respective DE-miRNAs, we predicted upstream transcription factors (TFs) and downstream target genes. Subsequently, we intersected target genes of respective DE-miRNAs with corresponding DE-mRNAs, thereby obtaining 127 upregulated genes in the caput and 92 upregulated genes in cauda. Enriched upregulated pathways included cell motility-related pathways for the caput, smooth muscle-related pathways for the corpus, and immune-associated pathways for the cauda. Protein-protein interaction (PPI) network was constructed to extract key module for the caput and cauda, followed by identifying hub genes through cytohubba. Epididymis tissues from six mice were applied to validate hub genes expression using qRT-PCR, and 7 of the 10 genes displayed identical expression trends in mice caput/cauda. These hub genes were found to be predominantly distributed in spermatozoa using scRNA-seq data. In addition, target genes of DE-miRNAs were intersected with genes in the PPI network for each segment. Subsequently, the miRNA and mRNA regulatory networks for the caput and cauda were constructed. Conclusively, we uncover segment-specific miRNA-mRNA regulatory network, upstream TFs, and downstream pathways of the human epididymis, warranting further investigation into epididymal segment-specific functions.
从睾丸中释放出来的精子在成熟并在附睾中运动之前不能使卵子受精。基于三个 bulk 和一个单细胞 RNA-seq（scRNA-seq）数据系列，我们比较了附睾特定节段样本与其他样本之间的 mRNA 或 miRNA 表达。在此，我们在附睾头部鉴定了 570 个差异表达的 mRNAs（DE-mRNAs）和 23 个差异表达的 miRNAs（DE-miRNAs），在附睾体部鉴定了 175 个 DE-mRNAs 和 15 个 DE-miRNAs，在附睾尾部鉴定了 946 个 DE-mRNAs 和 12 个 DE-miRNAs。根据各自的 DE-miRNAs，我们预测了上游转录因子（TFs）和下游靶基因。随后，我们将各自的 DE-miRNAs 的靶基因与相应的 DE-mRNAs 进行了交集，从而获得了在附睾头部上调的 127 个基因和在附睾尾部上调的 92 个基因。上调途径包括头部与细胞运动相关的途径、体部与平滑肌相关的途径和尾部与免疫相关的途径。构建蛋白-蛋白相互作用（PPI）网络以提取头部和尾部的关键模块，并通过 cytohubba 识别枢纽基因。使用 qRT-PCR 验证了来自 6 只小鼠的附睾组织中枢纽基因的表达情况，其中 10 个基因中的 7 个在小鼠头部/尾部显示出相同的表达趋势。这些枢纽基因通过 scRNA-seq 数据发现主要分布在精子中。此外，在每个节段中，DE-miRNAs 的靶基因与 PPI 网络中的基因进行了交集。随后，构建了头部和尾部的 miRNA 和 mRNA 调控网络。总之，我们揭示了人附睾的节段特异性 miRNA-mRNA 调控网络、上游 TFs 和下游途径，值得进一步研究附睾的节段特异性功能。
