Proteomics and single-cell RNA analysis of Akap4-knockout mice model confirm indispensable role of Akap4 in spermatogenesis
Proteomics and single-cell RNA analysis of Akap4-knockout mice model confirm indispensable role of Akap4 in spermatogenesis
复制标题
Akap4 敲除小鼠模型的蛋白质组学和单细胞 RNA 分析证实了 Akap4 在精子发生中不可或缺的作用
DOI：
10.1016/j.ydbio.2019.06.017
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发表时间：
2019-10-15
影响因子：
2.7
通讯作者：
Sun, Ling
中科院分区：
文献类型：
--
作者：
Fang, Xiang;Huang, Ling-Long;Sun, Ling
精子纤维鞘是一种独特的细胞骨架结构，参与精子成熟、运动和获能等多种生理功能。AKAP4已被描述为纤维鞘的结构和功能完整性所必需的。我们利用CRISPR-Cas9系统建立了AKAP4基因敲除小鼠系。对精子和睾丸的细胞形态和活力进行了研究，证实了AKAP4基因的缺失导致精子形态异常、活力异常和不育。研究了睾丸的蛋白质组分，发现在AKAP4基因敲除的小鼠中，AKAP4显著减少。睾丸单细胞RNA测序和分析显示，AKAP4、Haspin和Ccdc38三个基因在总细胞群中发生了显著变化。单细胞RNA表达谱还表明，AKAP4基因敲除的睾丸与野生型睾丸的主要差异存在于伸长细胞团，在AKAP4基因敲除的睾丸中，带有细胞黏附和迁移标记基因的伸长细胞亚群增加，而以线粒体鞘基因标记的伸长细胞亚群减少。我们的结果揭示了精子发生的复杂和协调的过程，并证实了AKAP4的S在精子鞭毛的完整性和精子的逐步成熟中所起的不可或缺的作用。
Sperm fibrous sheath, a unique cytoskeletal structure, is implicated in various sperm physiological functions, such as sperm maturation, motility and capacitation. AKAP4 has been described to be required for structural and functional integrity of the fibrous sheath. We generated Akap4-knockout mice line using CRISPR-Cas9 system. Cytomorphology and motility of sperm and testes were studied, confirming loss of Akap4 led to abnormal sperm morphology, motility and infertility. The proteomic components of testes were studied and Akap4 was found to be significantly decreased in the Akap4-knockout mice. Testis single-cell RNA sequencing and analysis revealed three genes with significant change in the general cell population, i.e., Akap4, Haspin, and Ccdc38. The single-cell RNA expression profiles also showed that the major difference between Akap4-knockout and wild-type testes existed in the elongating cell cluster, where in the Akap4-knockout testes, a subgroup of elongating cells with marker genes involved in cell adhesion and migration were increased, while a subgroup of elongating cells marked by mitochondrial sheath genes were decreased. Our results revealed the complex and well-coordinated procedures of spermatogenesis, and substantiated Akap4's indispensable roles in the integrity of sperm flagellum and the step-wise maturation of spermatozoa.