Immunology Frontier Research Center, Osaka University, Osaka, Japan.
Department of Experimental Genome Research, Research Institute for Microbial Diseases, Osaka University, Osaka, Japan.
Andrology. 2023 May;11(4):698-709. doi: 10.1111/andr.13400. Epub 2023 Feb 14.
Adenosine deaminase domain containing 2 (ADAD2) is a testis-specific protein composed of a double-stranded RNA binding domain and a non-catalytic adenosine deaminase domain. A recent study showed that ADAD2 is indispensable for the male reproduction in mice. However, the detailed functions of ADAD2 remain elusive.
This study aimed to investigate the cause of male sterility in Adad2 mutant mice and to understand the molecular functions of ADAD2.
Adad2 homozygous mutant mouse lines, Adad2 and Adad2 , were generated by CRISPR/Cas9. Western blotting and immunohistochemistry were used to reveal the expression and subcellular localization of ADAD2. Co-immunoprecipitation tandem mass spectrometry was employed to determine the ADAD2-interacting proteins in mouse testes. RNA-sequencing analyses were carried out to analyze the transcriptome and PIWI-interacting RNA (piRNA) populations in wildtype and Adad2 mutant testes.
Adad2 and Adad2 mice exhibit male-specific sterility because of abnormal spermiogenesis. ADAD2 interacts with multiple RNA-binding proteins involved in piRNA biogenesis, including MILI, MIWI, RNF17, and YTHDC2. ADAD2 co-localizes and forms novel granules with RNF17 in spermatocytes. Ablation of ADAD2 impairs the formation of RNF17 granules, decreases the number of cluster-derived pachytene piRNAs, and increases expression of ping-pong-derived piRNAs.
In collaboration with RNF17 and other RNA-binding proteins in spermatocytes, ADAD2 directly or indirectly functions in piRNA biogenesis.
腺苷脱氨酶结构域包含蛋白 2（ADAD2）是一种睾丸特异性蛋白，由双链 RNA 结合域和非催化腺苷脱氨酶域组成。最近的一项研究表明，ADAD2 对于小鼠的雄性生殖是不可或缺的。然而，ADAD2 的详细功能仍不清楚。
本研究旨在探讨 Adad2 突变小鼠雄性不育的原因，并了解 ADAD2 的分子功能。
通过 CRISPR/Cas9 生成 Adad2 纯合突变鼠系 Adad2 和 Adad2 。Western blot 和免疫组织化学用于揭示 ADAD2 的表达和亚细胞定位。免疫共沉淀串联质谱用于确定小鼠睾丸中 ADAD2 的相互作用蛋白。RNA 测序分析用于分析野生型和 Adad2 突变睾丸的转录组和 PIWI 相互作用 RNA（piRNA）群体。
Adad2 和 Adad2 小鼠由于精子发生异常而表现出雄性特异性不育。ADAD2 与多个参与 piRNA 生物发生的 RNA 结合蛋白相互作用，包括 MILI、MIWI、RNF17 和 YTHDC2。ADAD2 在精母细胞中与 RNF17 共定位并形成新的颗粒。ADAD2 的缺失会损害 RNF17 颗粒的形成，减少簇衍生的粗线期 piRNA 的数量，并增加乒乓衍生的 piRNA 的表达。
ADAD2 在精母细胞中与 RNF17 和其他 RNA 结合蛋白一起直接或间接参与 piRNA 生物发生。
