TRIM28 establishes skeletal stem cell identity and safeguards skeletogenesis (scRNA-Seq)

The identity, renewal, and multipotency of stem cells are controlled by epigenetic mechanisms. DNA methylation and trimethylation on histone H3K9 (H3K9me3) are two epigenetic marks that coordinate gene silencing in early embryogenesis. Yet whether they also play a role in regulating somatic stem cell activities governing organogenesis, particularly in the skeletal system, is unknown. Here we show that chromatin silencing, established by TRIM28 (aka. KAP1 and TIF1β) via DNA methylation and H3K9me3 during early organogenesis is crucial for skeletal development. Loss of TRIM28 in skeletal stem cells unsilences the promoter and two previously uncharacterized enhancers of the Grem1 gene, leading to GREM1 hyperexpression, which further activates AKT/mTORC1 signaling to promote skeletal stem cell renewal while restricting osteogenesis and chondrogenesis. Notably, loss of Trim28 in the growth plate also leads to the emergence of a novel stem cell cluster with neural crest cell properties and a distinctive neurogenic tendency. These collectively result in a wide range of skeletal abnormalities. Taken together, our data suggest that TRIM28 coordinates methylation of DNA and histone H3K9 to safeguard the identity and fate determination of skeletal stem cells by epigenetically silencing the GREM1/AKT/mTORC1 signaling axis. Single cell RNA profiles of E17.5 WT and Trim28 KO mouse rib cartilage cells.干细胞的身份维持、自我更新与多能性受表观遗传机制调控。DNA甲基化与组蛋白H3K9三甲基化（H3K9me3）是早期胚胎发生中协同基因沉默的两类表观遗传标记。然而，它们是否同样参与调控介导器官发生的成体干细胞活性，尤其是在骨骼系统中，目前尚不清楚。本研究表明，在早期器官发生过程中，TRIM28（又名KAP1、TIF1β）通过DNA甲基化与H3K9me3建立的染色质沉默对骨骼发育至关重要。骨骼干细胞中TRIM28的缺失会解除Grem1基因启动子与两个此前未被表征的增强子的沉默，导致GREM1过度表达，进而激活AKT/mTORC1信号通路以促进骨骼干细胞自我更新，同时抑制成骨与成软骨分化。值得注意的是，生长板中Trim28的缺失还会催生一类具有神经嵴细胞特性与独特神经发生倾向的新型干细胞簇。这些变化共同导致了广泛的骨骼异常。综上，我们的研究数据表明，TRIM28通过表观遗传沉默GREM1/AKT/mTORC1信号轴，协同调控DNA与组蛋白H3K9的甲基化，以维持骨骼干细胞的身份与命运决定。本数据集包含胚胎第17.5天（Embryonic day 17.5, E17.5）野生型（Wild Type, WT）与Trim28基因敲除（Knockout, KO）小鼠肋软骨细胞的单细胞RNA谱。