小鼠雄性生殖干细胞转录组分析揭示成熟精原干细胞特征

doi:10.16288/j.yczz.22-047

遗传 ›› 2022, Vol. 44 ›› Issue (7): 591-608.doi: 10.16288/j.yczz.22-047

小鼠雄性生殖干细胞转录组分析揭示成熟精原干细胞特征

郭彦¹(), 杨乐乐²(), 戚华宇²()

1. 中国科学技术大学,生命科学与医学部生命科学学院医药生物技术系,合肥 230026
2. 中国科学院广州生物医药与健康研究院,细胞谱系与发育研究中心,广州 510530

收稿日期:2022-02-23 修回日期:2022-04-26 出版日期:2022-07-20 发布日期:2022-07-06
通讯作者: 杨乐乐,戚华宇 E-mail:rxm749426464@qq.com;yang_lele@gibh.ac.cn;qi_huayu@gibh.ac.cn
作者简介:郭彦,在读硕士研究生,专业方向：生物工程。E-mail: rxm749426464@qq.com
基金资助:
广东省自然科学基金面上项目编号(2020A1515010882);广东省科技计划项目编号(2020B1212060052);生物岛实验室前沿探索项目资助编号(2018GZR110105021)

Transcriptome analysis of mouse male germline stem cells reveals characteristics of mature spermatogonial stem cells

Yan Guo¹(), Lele Yang²(), Huayu Qi²()

1. Department of Pharmaceutical Biotechnology, School of Life Sciences, Department of Life Sciences and Medicine, University of Science and Technology of China, Hefei 230026, China
2. Center for Cell Lineage and Development,Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510530, China

Received:2022-02-23 Revised:2022-04-26 Online:2022-07-20 Published:2022-07-06
Contact: Yang Lele,Qi Huayu E-mail:rxm749426464@qq.com;yang_lele@gibh.ac.cn;qi_huayu@gibh.ac.cn
Supported by:
Supported by the Natural Science Foundation of Guangdong Province No(2020A1515010882);Science and Technology Planning Project of Guangdong Province No(2020B1212060052);the Frontier Research Program of Guangzhou Regenerative Medicine and Health, Bioland Laboratory No(2018GZR110105021)

摘要/Abstract

摘要：

精原干细胞(spermatogonial stem cells, SSCs)是成年动物睾丸中的成体干细胞,具有自我更新与分化的能力。小鼠(Mus musculus)精原干细胞来源于胚胎期的原始生殖细胞(primodial germ cells, PGCs),小鼠出生前原始生殖细胞处于有丝分裂静止状态,出生后恢复增殖并由曲细精管中央迁移至管壁基质,建立稳定的精原干细胞克隆。成熟小鼠的精原干细胞周期性地启动精子发生以维持雄性动物长期稳定的生殖能力。精原干细胞在其建立和成熟后是否具有特征上的差异目前尚不清楚。本研究在前期建立的不同年龄小鼠精原干细胞(表达多能性基因Pou5f1编码的OCT4)转录组数据基础上,对小鼠新生期(出生后3天)、幼年期(出生后7天)和成熟期(2~3月龄)精原干细胞的基因表达差异进行了生物信息学分析,包括差异表达基因(differentially expression genes, DEGs)的筛选、DEGs编码的蛋白相互作用网络(protein-protein interaction, PPI)的建立、功能聚类富集(Gene Ontology, GO)和通路分析(Kyoto Encyclopedia of Genes and Genomes, KEGG),以及使用基于GO、KEGG和HALLMARK的基因集富集分析(gene set enrichment analysis, GSEA)。结果显示,OCT4阳性精原干细胞在小鼠新生期、幼年期和成熟期存在大量差异表达基因,所编码的蛋白主要生物学功能集中在生物合成和能量代谢、免疫反应、细胞连接和迁移以及细胞分化等方面。精原干细胞细胞膜成分的显著变化可能影响精原干细胞的超敏反应、细胞间相互作用以及对细胞外环境因子的应答反应。在能量代谢方式上,随着年龄的增加,OCT4阳性精原干细胞逐渐从线粒体氧化磷酸化作用转变为糖酵解作用,同时也显著减少了细胞内核糖体形成相关基因的转录。这些结果为进一步研究雄性生殖干细胞形成和成熟的调控机制提供了新的思路。

关键词: 精原干细胞, OCT4, 转录组, 差异表达基因

Abstract:

Spermatogonial stem cells (SSCs) are adult stem cells in the testis of male animals and have the ability in self-renewal and differentiation. SSCs are derived from primordial germ cells (PGCs) that are mitotically arrested in the embryo before birth. Following the birth of the animal, PGCs resume mitosis and migrate from the centre of the seminiferous tubules to the basement membrane. The descendent of PGCs (also called gonocytes) establish stable SSC colonies in about a week postnatally in order to support the life-long spermatogenesis. Whether SSCs at different developmental stages differ in their molecular and cellular characteristics is currently unclear. In the presented study, we conducted bioinformatics analyses using transcriptomics data established previously in the laboratory on OCT4 (encoded by the pluripotent gene Pou5f1) expressing SSCs from the neonatal (3 days-post-partum, 3-dpp), juvenile (7-dpp) and adult (2~3-month) mice, including screen of differentially expressed genes (DEGs), protein-protein interaction (PPI) network analysis of DEGs and clustering of sub-networks from PPI. GO (Gene Ontology) and KEGG (Kyoto Encyclopedia of Genes and Genomes) analyses were also performed on clustered sub-networks of the PPI. In addition, all genes were analyzed using GSEA (gene set enrichment analysis) based on GO, KEGG and HALLMARK gene sets. The results showed that SSCs have a large number of DEGs among OCT4-positive SSCs from neonatal, juvenile and adult mice. The distinguishable biological functions encoded by these DEGs include biosynthesis and energy metabolism, immune response, cell junction and expression of migration and cell differentiation-related genes. Significant changes in the cell membrane composition of OCT4-positive SSCs may not only cause hypersensitive immune reactions but also affect the cell-cell contact and responses to secreted cytokines in the extracellular environment. The results also suggest that OCT4-positive SSCs may shift metabolic state from oxidative phosphorylation to glycolysis and significantly reduce the transcription of genes related to ribosome formation during aging. These results provide new clues for future research on the regulatory mechanisms of male germline stem cell development, growth and aging.

Key words: spermatogonial stem cells, OCT4, transcriptome, differentially expressed genes

郭彦, 杨乐乐, 戚华宇. 小鼠雄性生殖干细胞转录组分析揭示成熟精原干细胞特征[J]. 遗传, 2022, 44(7): 591-608.

Yan Guo, Lele Yang, Huayu Qi. Transcriptome analysis of mouse male germline stem cells reveals characteristics of mature spermatogonial stem cells[J]. Hereditas(Beijing), 2022, 44(7): 591-608.

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序号	基因	上游引物(5′→3′)	下游引物(5′→3′)
1	Actin	TGTACCCAGGCATTGCTGACAG	CTGCTGGAAGGTGGACAGTG
2	Id4	CACTCACCGCGCTCAACA	CTCCGGTGGCTTGTTTCTCTT
3	Ybx2	GGAGTTTGATGTCGTGGAAGG	CGTCGATTAGGGGCATAGCG
4	Ddr1	ATGCTGACATGAAGGGACATTT	GGTGTAGCCTACGAAGGTCCA
5	Nefm	ATGACGAGCCATTTCCCACT	TGCAGTCCAAGAGCATCGAG
6	Fxyd6	TCTCCGTTGGGATACTTCTCAT	CTCCGCAGCGTTTGTAGTGAT
7	Gfra1	AACTGCCAGCCAGAGTCAAG	GGCTGCTGGAGTCTATGTAG
8	Rarg	TGCCTGGTTTTACAGGGCTC	TCCGAGAATGTCATAGTGTCCT

小鼠雄性生殖干细胞转录组分析揭示成熟精原干细胞特征

Transcriptome analysis of mouse male germline stem cells reveals characteristics of mature spermatogonial stem cells

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