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miR-199-5p regulates spermiogenesis at the posttranscriptional level via targeting Tekt1 in allotriploid crucian carp
Journal of Animal Science and Biotechnology ( IF 7 ) Pub Date : 2022-04-13 , DOI: 10.1186/s40104-022-00693-4
Shengnan Li 1 , Qiubei Wang 1 , Lu Huang 1 , Siyu Fan 1 , Ting Li 1 , Yuqing Shu 1 , Chun Zhang 1, 2 , Yi Zhou 1, 2 , Qingfeng Liu 1 , Kaikun Luo 1 , Min Tao 1, 2 , Shaojun Liu 1, 2
Affiliation  

Sperm abnormalities are one of the primary factors leading to male sterility, but their pathogenesis is still unclear. Although miRNAs are suggested to exert important roles in the regulation of spermatogenesis at both transcriptional and posttranscriptional levels, little is currently known regarding the regulation of sperm flagella assembly by microRNAs (miRNAs). The role of miRNAs in the development of sperm abnormalities in sterile triploid fish has not been studied. In this study, we found that miR-199-5p was widely expressed in all detected tissues of different-ploidy crucian carp. As one of the testis-specific candidate markers, Tekt1 was predominantly expressed in the testis. Quantitative real-time PCR (qRT-PCR) analyses showed that the expression trend of miR-199-5p was exactly opposite to that of Tekt1. Through bioinformatics analysis, we identified a putative miR-199-5p binding site in the Tekt1 mRNA. We further identified Tekt1 as a target of miR-199-5p using luciferase reporter assay. Finally, we confirmed that miR-199-5p was necessary for sperm flagellar assembly and spermatogenesis in vivo via intraperitoneal injection of miR-199-5p antagomir or agomir in diploid red crucian carp. Moreover, miR-199-5p gain-of-function could lead to spermatids apoptosis and abnormal spermatozoa structure, which is similar to that of allotriploid crucian carp. Our studies suggested that abnormally elevated miR-199-5p inhibited the sperm flagella formation in spermiogenesis by negatively regulating the expression of Tekt1, thereby causing sperm abnormalities of male allotriploid crucian carp.

中文翻译:

miR-199-5p 通过靶向异源三倍体鲫鱼中的 Tekt1 在转录后水平调节精子发生

精子异常是导致男性不育的主要因素之一,但其发病机制尚不清楚。尽管 miRNA 被认为在转录和转录后水平的精子发生调节中发挥重要作用,但目前关于 microRNA (miRNA) 对精子鞭毛组装的调节知之甚少。尚未研究 miRNA 在不育三倍体鱼精子异常发育中的作用。在这项研究中,我们发现 miR-199-5p 在所有检测到的不同倍性鲫鱼组织中广泛表达。作为睾丸特异性候选标记之一,Tekt1 主要在睾丸中表达。定量实时PCR(qRT-PCR)分析表明miR-199-5p的表达趋势与Tekt1的表达趋势完全相反。通过生物信息学分析,我们在 Tekt1 mRNA 中鉴定了一个推定的 miR-199-5p 结合位点。我们使用荧光素酶报告基因分析进一步将 Tekt1 鉴定为 miR-199-5p 的靶标。最后,我们通过腹膜内注射 miR-199-5p antagomir 或 agomir 在二倍体红鲫鱼体内证实了 miR-199-5p 是体内精子鞭毛组装和精子发生所必需的。此外,miR-199-5p功能获得可导致精子细胞凋亡和精子结构异常,这与异源三倍体鲫鱼相似。我们的研究表明,异常升高的 miR-199-5p 通过负调控 Tekt1 的表达来抑制精子发生过程中精子鞭毛的形成,从而导致雄性异源三倍体鲫鱼精子异常。我们使用荧光素酶报告基因分析进一步将 Tekt1 鉴定为 miR-199-5p 的靶标。最后,我们通过腹膜内注射 miR-199-5p antagomir 或 agomir 在二倍体红鲫鱼体内证实了 miR-199-5p 是体内精子鞭毛组装和精子发生所必需的。此外,miR-199-5p功能获得可导致精子细胞凋亡和精子结构异常,这与异源三倍体鲫鱼相似。我们的研究表明,异常升高的 miR-199-5p 通过负调控 Tekt1 的表达来抑制精子发生过程中精子鞭毛的形成,从而导致雄性异源三倍体鲫鱼精子异常。我们使用荧光素酶报告基因分析进一步将 Tekt1 鉴定为 miR-199-5p 的靶标。最后,我们通过腹膜内注射 miR-199-5p antagomir 或 agomir 在二倍体红鲫鱼体内证实了 miR-199-5p 是体内精子鞭毛组装和精子发生所必需的。此外,miR-199-5p功能获得可导致精子细胞凋亡和精子结构异常,这与异源三倍体鲫鱼相似。我们的研究表明,异常升高的 miR-199-5p 通过负调控 Tekt1 的表达来抑制精子发生过程中精子鞭毛的形成,从而导致雄性异源三倍体鲫鱼精子异常。我们通过在二倍体红鲫中腹膜内注射 miR-199-5p antagomir 或 agomir 证实了 miR-199-5p 是体内精子鞭毛组装和精子发生所必需的。此外,miR-199-5p功能获得可导致精子细胞凋亡和精子结构异常,这与异源三倍体鲫鱼相似。我们的研究表明,异常升高的 miR-199-5p 通过负调控 Tekt1 的表达来抑制精子发生过程中精子鞭毛的形成,从而导致雄性异源三倍体鲫鱼精子异常。我们通过在二倍体红鲫中腹膜内注射 miR-199-5p antagomir 或 agomir 证实了 miR-199-5p 是体内精子鞭毛组装和精子发生所必需的。此外,miR-199-5p功能获得可导致精子细胞凋亡和精子结构异常,这与异源三倍体鲫鱼相似。我们的研究表明,异常升高的 miR-199-5p 通过负调控 Tekt1 的表达来抑制精子发生过程中精子鞭毛的形成,从而导致雄性异源三倍体鲫鱼精子异常。
更新日期:2022-04-13
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