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Journal of Integrative Agriculture  2024, Vol. 23 Issue (7): 2362-2378    DOI: 10.1016/j.jia.2023.04.036
Animal Science · Veterinary Medicine Advanced Online Publication | Current Issue | Archive | Adv Search |
Dynamic transcriptome profiles and novel markers in bovine spermatogenesis revealed by single-cell sequencing
Yuan Gao1, 2, Fuxia Bai1, Qi Zhang1, Xiaoya An1, Zhaofei Wang1, Chuzhao Lei1, Ruihua Dang1#
1 Key Laboratory of Animal Genetics, Breeding and Reproduction of Shaanxi Province, College of Animal Science and Technology, Northwest A&F University, Yangling 712100, China
2 College of Life Sciences, Northwestern Polytechnical University, Xi’an 710072, China
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摘要  睾丸是精子发生的重要器官,正常的睾丸发育是种公牛优良繁殖性能的基础,此过程涉及到生殖细胞和体细胞之间复杂的相互作用。然而,睾丸细胞的异质性阻碍了牛睾丸发育机制的研究。研究以安格斯牛为研究对象,利用 scRNA-seq技术对性成熟前后安格斯牛睾丸细胞进行测序分析探讨公牛睾丸发育的转录机制。本研究利用Seurat包对数据进行过滤质控标准化降维聚类青春期前后的睾丸细胞进行差异基因分析, 利用KOBAS进行富集分析, 利用Monocle包对生精细胞进行拟时分析, 最利用FISH对标记基因进行验证。睾丸样本经过质控筛选后得到青春期前及青春期牛睾丸细胞共11083;经过降维、聚类等分析过程后,共得到12睾丸细胞亚群,其中包括9个体细胞亚群(未成熟支持细胞、成熟支持细胞、间质祖细胞、间质细胞、管周肌细胞、T细胞、巨噬细胞、平滑肌细胞、内皮细胞3个细胞亚群(精原细胞、精母细胞精子细胞),不同样本之间具有明显的细胞异质性;进一步将睾丸生精细胞进行聚类分析,获得13精原干细胞1期、精原干细胞2期、正在分化的精原细胞、已分化精原细胞、细线期、偶线期、粗线期、双线期次级精母细胞期、圆型精子、延长型精子1期延长型精子2期、未成熟精子);通过Monocle算法,成功构建了生精细胞系的拟时分化轨迹;并筛选出牛睾丸不同类群细胞的特异标记基因,例如:精原干细胞标记基因GRAF2MORC1,分化的精原细胞标记基因HJURPTCF19,代表未成熟支持细胞标记基因ARSE成熟支持细胞标记基因CLEC12B间质细胞标记基因LOC112441470等。综上所述,研究揭示了牛性成熟过程中生殖细胞的动态变化,并提出牛睾丸细胞独特的标记基因,解释牛睾丸发育和精子生成的机制提供了理论依据,为牛青春期的生殖细胞及体细胞的发育提供了新的见解,同时也为后期研究提供了宝贵的数据资源。

Abstract  
Testicular development is an important biological process in male and requires interaction between the male germ cells and somatic cells. However, the mechanisms of testicular development in livestock, particularly in cattle, are poorly understood. Furthermore, cellular heterogeneity hinders the profiling of different cell types at different developmental stages. In this study, we first performed a single-cell transcriptomic study of the bovine testis development during puberty by using 10× genomics single-cell RNA sequencing (scRNA-seq). By collecting the scRNA-seq data from 11,083 cells from prepubertal and pubertal bovine testes, a high-resolution scRNA-seq atlas was described, identifying 9 somatic and 13 spermatogenic clusters. We also distinguished several stage-specific marker genes for bovine germ cells and somatic cells, such as GRAF2 and MORC1 for SSC (spermatogonial stem cells), HJURP and TCF19 for differentiating spermatogonia, ARSE for immature Sertoli, CLEC12B for mature Sertoli, LOC112441470 for Leydig. In conclusion, we have examined the transcription levels and constructed the single-cell developmental maps of germ cells and somatic cells during testicular development in Angus cattle. The datasets provided new insights into spermatogenesis and testicular somatic cell development in cattle.
Keywords:  scRNA-seq       cattle        spermatogenesis        testis development  
Received: 16 January 2023   Accepted: 31 March 2023
Fund: 
This work was supported by the National Natural Science Foundation of China (81770514), the 2020 Scientist Plus Engineer Program of Shaanxi Province, China (K4050422468), the Key Research & Development Plan of Shaanxi Province, China (2020NY-002), the Natural Science Basis Research Plan of Shaanxi Province, China (2023-JC-QN-0242), and the China Agriculture Research System of MOF and MARA (CARS-37).
About author:  Yuan Gao, E-mail: gaoyuan2021@nwpu.edu.cn; #Correspondence Ruihua Dang, Tel: +86-29-87092120, E-mail: dangruihua@nwsuaf.edu.cn

Cite this article: 

Yuan Gao, Fuxia Bai, Qi Zhang, Xiaoya An, Zhaofei Wang, Chuzhao Lei, Ruihua Dang. 2024. Dynamic transcriptome profiles and novel markers in bovine spermatogenesis revealed by single-cell sequencing. Journal of Integrative Agriculture, 23(7): 2362-2378.

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