Center for Medical Genetics, School of Life Sciences, Central South University, Changsha, China.
Hunan Key Laboratory of Animal Models for Human Diseases, Central South University, Changsha, China.
FASEB J. 2021 Apr;35(4):e21460. doi: 10.1096/fj.202002294RR.
Spermatogenesis is a highly sophisticated process that comprises of mitosis, meiosis, and spermiogenesis. RNF216 (ring finger protein 216), an E3 ubiquitin ligase, has been reported to be essential for spermatogenesis and male fertility in mice. However, the stages affected by Rnf216 deficiency and its underlying molecular pathological mechanisms are still unknown. In this study, we generated Rnf216-deficient mice (Rnf216 ) using CRISPR-Cas9 technology. Knockout of Rnf216 led to infertility in male but not female mice. Rnf216 knockout affected the prophase of meiosis I, as no genotypic difference was observed until 12 dpp (days postpartum). Rnf216 spermatocytes were incompletely arrested at the zygotene stage and underwent apoptosis at approximately the pachytene stage. The proportion of zygotene spermatocytes was significantly increased, whereas the proportion of pachytene spermatocytes was significantly decreased in Rnf216 testes. Nevertheless, there was no significantly genotypic difference in the number of diplotene spermatocytes. We further revealed that the PKA catalytic subunit β (PRKACB) was significantly increased, which subsequently resulted in elevated PKA activity in testes from adult as well as 9 dpp Rnf216 mice. RNF216 interacts with PRKACB and promotes its degradation through the ubiquitin-lysosome pathway. Collectively, our results revealed an important role for RNF216 in regulation of meiosis and PKA stability in the testes.
精子发生是一个高度复杂的过程，包括有丝分裂、减数分裂和精子发生。RNF216（环指蛋白 216）是一种 E3 泛素连接酶，已被报道在小鼠中对精子发生和雄性生育力是必需的。然而，Rnf216 缺乏影响的阶段及其潜在的分子病理机制尚不清楚。在这项研究中，我们使用 CRISPR-Cas9 技术生成了 Rnf216 缺失的小鼠（Rnf216 ）。Rnf216 的缺失导致雄性但不是雌性小鼠不育。Rnf216 缺失影响减数分裂 I 的前期，直到 12 dpp（产后天数）才观察到基因型差异。Rnf216 缺失的精母细胞不完全停滞在合线期，大约在粗线期发生凋亡。Rnf216 睾丸中的合线期精母细胞比例显著增加，而粗线期精母细胞比例显著减少。然而，双线期精母细胞的数量没有明显的基因型差异。我们进一步揭示了 PKA 催化亚基 β（PRKACB）显著增加，这导致成年和 9 dpp Rnf216 小鼠睾丸中的 PKA 活性升高。RNF216 与 PRKACB 相互作用，并通过泛素-溶酶体途径促进其降解。总之，我们的结果揭示了 RNF216 在调节减数分裂和睾丸中 PKA 稳定性方面的重要作用。
