TRIM37–PARP1–TET1 axis maintains stemness and prevents osteoporosis by inhibiting DNMT1 alternative splicing via 5hmC regulation

The significance of DNA hydroxymethylation in stemness remains unknown. Here, we demonstrate 5hmC levels positively regulate mesenchymal stem cell (MSC) properties. Mechanistically, PARP1 recruits TET1 to hydrolyze methylated nucleotides on DNMT1 exons, helping CTCF to bind to exons and prevent DNMT1 alternative splicing in early MSCs. Furthermore, ATM phosphorylates TRIM37 at Th203 and promotes its entry into the nucleus, as well as the monoubiquitination of PARP1, thereby stabilizing the PARP1 protein. CTCF or TRIM37 knockdown induces replicative senescence of MSCs with loss of full-length DNMT1, while simultaneous treatment of MSCs during expansion with ATM activators, such as resveratrol, and TET1 activator, vitamin C, induces the rejuvenation of late MSCs through the TRIM37/PARP1/DNMT1 pathway. Through gene knockout, TRIM37 and PARP1 are shown to be involved in MSC aging and bone repair in vivo. This study highlights the role of DNA hydroxymethylation and its regulators in stemness, offering strategies for therapeutic interventions.DNA羟甲基化（DNA hydroxymethylation）在干细胞干性（stemness）中的调控意义迄今尚未阐明。本研究证实，5-羟甲基胞嘧啶（5hmC）水平可正向调控间充质干细胞（MSC）的干性特性。机制层面分析显示，聚腺苷二磷酸核糖聚合酶1（PARP1）招募TET1水解DNA甲基转移酶1（DNMT1）外显子上的甲基化核苷酸，辅助CCCTC结合因子（CTCF）结合该外显子，进而阻断早期MSC中DNMT1的可变剪接。此外，毛细血管扩张性共济失调突变激酶（ATM）可在苏氨酸203位点磷酸化三结构域蛋白37（TRIM37），促进其入核并介导PARP1的单泛素化修饰，进而稳定PARP1蛋白。敲低CTCF或TRIM37可诱导MSC发生复制性衰老，并伴随全长DNMT1的缺失；而在MSC扩增培养阶段联合使用ATM激活剂（如白藜芦醇）与TET1激活剂维生素C，可通过TRIM37/PARP1/DNMT1通路实现晚期MSC的年轻化。通过基因敲除实验证实，TRIM37与PARP1参与体内MSC衰老进程与骨修复过程。本研究阐明了DNA羟甲基化及其调控因子在干细胞干性维持中的关键作用，为相关治疗干预提供了新策略。