Key Laboratory for Animal Genetics, Breeding and Reproduction of Shaanxi Province, College of Animal Science and Technology, Northwest A&F University, Yangling, Shaanxi 712100, China.
Biol Reprod. 2024 Jan 13;110(1):48-62. doi: 10.1093/biolre/ioad136.
Genomic integrity is critical for sexual reproduction, ensuring correct transmission of parental genetic information to the descendant. To preserve genomic integrity, germ cells have evolved multiple DNA repair mechanisms, together termed as DNA damage response. The RNA N6-methyladenosine is the most abundant mRNA modification in eukaryotic cells, which plays important roles in DNA damage response, and YTH N6-methyladenosine RNA binding protein 2 (YTHDF2) is a well-acknowledged N6-methyladenosine reader protein regulating the mRNA decay and stress response. Despite this, the correlation between YTHDF2 and DNA damage response in germ cells, if any, remains enigmatic. Here, by employing a Ythdf2-conditional knockout mouse model as well as a Ythdf2-null GC-1 mouse spermatogonial cell line, we explored the role and the underlying mechanism for YTHDF2 in spermatogonial DNA damage response. We identified that, despite no evident testicular morphological abnormalities under the normal circumstance, conditional mutation of Ythdf2 in adult male mice sensitized germ cells, including spermatogonia, to etoposide-induced DNA damage. Consistently, Ythdf2-KO GC-1 cells displayed increased sensitivity and apoptosis in response to DNA damage, accompanied by the decreased SET domain bifurcated 1 (SETDB1, a histone methyltransferase) and H3K9me3 levels. The Setdb1 knockdown in GC-1 cells generated a similar phenotype, but its overexpression in Ythdf2-null GC-1 cells alleviated the sensitivity and apoptosis in response to DNA damage. Taken together, these results demonstrate that the N6-methyladenosine reader YTHDF2 promotes DNA damage repair by positively regulating the histone methyltransferase SETDB1 in spermatogonia, which provides novel insights into the mechanisms underlying spermatogonial genome integrity maintenance and therefore contributes to safe reproduction.
基因组完整性对于有性生殖至关重要，它确保了父母遗传信息正确传递给后代。为了保护基因组完整性，生殖细胞进化出了多种 DNA 修复机制，这些机制统称为 DNA 损伤反应。RNA N6-甲基腺苷是真核细胞中最丰富的 mRNA 修饰，它在 DNA 损伤反应中发挥重要作用，YTH N6-甲基腺苷 RNA 结合蛋白 2（YTHDF2）是一种公认的 N6-甲基腺苷阅读蛋白，调节 mRNA 降解和应激反应。尽管如此，YTHDF2 与生殖细胞中 DNA 损伤反应之间的相关性（如果有的话）仍然是个谜。在这里，我们通过使用 Ythdf2 条件性敲除小鼠模型和 Ythdf2 缺失的 GC-1 小鼠精原细胞系，探索了 YTHDF2 在精原细胞 DNA 损伤反应中的作用及其潜在机制。我们发现，尽管在正常情况下成年雄性小鼠的睾丸形态没有明显异常，但 Ythdf2 在成年雄性小鼠中的条件性突变使生殖细胞（包括精原细胞）对依托泊苷诱导的 DNA 损伤更加敏感。同样，Ythdf2-KO GC-1 细胞对 DNA 损伤的敏感性和凋亡增加，伴随着 SET 域分叉 1（SETDB1，一种组蛋白甲基转移酶）和 H3K9me3 水平降低。GC-1 细胞中 Setdb1 的敲低产生了类似的表型，但 Ythdf2 缺失的 GC-1 细胞中 Setdb1 的过表达减轻了 DNA 损伤的敏感性和凋亡。综上所述，这些结果表明，N6-甲基腺苷阅读器 YTHDF2 通过正向调节精原细胞中的组蛋白甲基转移酶 SETDB1 促进 DNA 损伤修复，为精原细胞基因组完整性维持的机制提供了新的见解，从而有助于安全繁殖。
