Zhang Xiaoning, Zhou Wenwen, Zhang Peng, Gao Fengxin, Zhao Xiuling, Shum Winnie Waichi, Zeng Xuhui
Medical School, Institute of Reproductive Medicine, Nantong University, Nantong 226019, China.
Institute of Life Science, Nanchang University, Nanchang 330031, China.
Int J Mol Sci. 2021 Jan 11;22(2):652. doi: 10.3390/ijms22020652.
The calcium-binding protein spermatid-associated 1 (Cabs1) is a novel spermatid-specific protein. However, its function remains largely unknown. In this study, we found that a long noncoding RNA (lncRNA) transcripted from the gene antisense, , was also exclusively expressed in spermatids. and knockout mice were generated separately (using Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR)-Cas9 methods) to investigate their functions in spermatogenesis. The genetic loss of did not affect testicular and epididymal development; however, male mice exhibited significantly impaired sperm tail structure and subfertility. Ultrastructural analysis revealed defects in sperm flagellar differentiation leading to an abnormal annulus and disorganization of the midpiece-principal piece junction, which may explain the high proportion of sperm with a bent tail. Interestingly, the proportion of sperm with a bent tail increased during transit in the epididymis. Furthermore, Western blot and immunofluorescence analyses showed that a genetic loss of decreased Septin 4 and Krt1 and increased cyclin Y-like 1 (Ccnyl1) levels compared with the wild type, suggesting that deficiency disturbed the expression of cytoskeleton-related proteins. By contrast, mice were indistinguishable from the wild type regarding testicular and epididymal development, sperm morphology, concentration and motility, and male fertility. This study demonstrates that Cabs1 is an important component of the sperm annulus essential for proper sperm tail assembly and motility.
钙结合蛋白精子细胞相关蛋白1（Cabs1）是一种新的精子细胞特异性蛋白。然而，其功能在很大程度上仍不清楚。在本研究中，我们发现从该基因反义链转录的一种长链非编码RNA（lncRNA）也仅在精子细胞中表达。分别构建了该lncRNA和Cabs1基因敲除小鼠（采用成簇规律间隔短回文重复序列（CRISPR）-Cas9方法），以研究它们在精子发生中的功能。该lncRNA的基因缺失不影响睾丸和附睾的发育；然而，雄性小鼠精子尾部结构明显受损且生育力下降。超微结构分析显示精子鞭毛分化存在缺陷，导致环异常以及线粒体鞘-主段连接处紊乱，这可能解释了弯尾精子比例较高的原因。有趣的是，弯尾精子在附睾转运过程中的比例增加。此外，蛋白质免疫印迹和免疫荧光分析表明，与野生型相比，该lncRNA的基因缺失降低了Septin 4和角蛋白1（Krt1）的水平，并增加了细胞周期蛋白Y样蛋白1（Ccnyl1）的水平，提示该lncRNA缺陷扰乱了细胞骨架相关蛋白的表达。相比之下，Cabs1基因敲除小鼠在睾丸和附睾发育、精子形态、浓度和活力以及雄性生育力方面与野生型无差异。本研究表明，Cabs1是精子环的重要组成部分，对精子尾部的正常组装和运动至关重要。
