Molecular Control of MSC differentiation and Bone Formation by KDM4B

描述（由申请人提供）：本申请的长期目标是了解分子和表观遗传机制如何控制间充质干细胞/基质细胞（MSC）的成骨分化、成骨细胞功能和骨形成。骨髓间充质干细胞是具有自我更新能力和多向分化潜能的多能祖细胞，包括成骨、软骨形成和脂肪形成。虽然在理解MSC分化的转录控制方面已经取得了重大进展，但对骨形成如何受表观遗传学调控知之甚少。组蛋白甲基化是一个与基因表达的激活和抑制相关的重要过程，因此它在细胞分化的表观遗传调控中起着关键作用。虽然越来越多的证据表明组蛋白脱甲基酶在表观遗传学上调节胚胎干细胞的性质和功能，但脱甲基酶对MSC分化和骨形成的影响在很大程度上尚不清楚。为了探索脱甲基酶在MSC分化中的作用，我们系统地分析了BMP刺激的骨髓MSC中组蛋白脱甲基酶的表达，因为BMP是成骨分化的有效诱导剂。我们发现BMP快速诱导赖氨酸（K）特异性脱甲基酶（KDM 4 B;也称为JMJD 2B）的表达，该酶使赖氨酸9处的三甲基化组蛋白H3（H3 K9 me 3）脱甲基。H3 K9 me 3是参与生长和发育的基因沉默的标志。一般来说，当干细胞分化程序被触发时，一组特定的基因被激活。我们的初步研究表明，KDM 4 B促进骨髓间充质干细胞的成骨分化，同时抑制成脂分化。此外，我们发现，与年轻小鼠相比，从衰老小鼠分离的MSC中Kdm 4 b的表达显著下调。巧合的是，H3 K9 me 3标记在衰老小鼠或卵巢切除小鼠的成骨细胞中显著增加。基于这些新的发现，在本申请中，我们假设通过KDM 4 B擦除H3 K9 me 3标记在体外MSC的成骨分化以及骨形成和体内中起着不可或缺的作用。 提出了三个具体目标来检验我们的假设。目的1是确定KDM 4 B是否通过诱导DLX 5表观遗传地调节MSC谱系定型。目的2：探讨KDM 4 B清除H3 K9 me 3基因协同调控MSCs成骨分化的机制。目的3是确定KDM 4 B是否是体内骨形成所必需的，以及KDM 4 B的失调是否会损害骨质疏松症中成骨细胞的功能和骨形成。由于组蛋白去甲基化酶是化学修饰的，KDM 4 B可能作为一种新的治疗靶点，用于特异性控制再生医学中MSC的分化，并为代谢性骨疾病（如骨质疏松症）的新治疗提供线索。

DESCRIPTION (provided by applicant): The long-term objectives of this application are to understand how molecular and epigenetic mechanisms control osteogenic differentiation of mesenchymal stem/stromal cells (MSCs), osteoblast function, and bone formation. MSCs are multipotent progenitor cells with self-renewal capabilities and multilineage differentiation potentials including osteogenesis, chondrogenesis and adipogenesis. Although significant progress has been made in understanding transcriptional control of MSC differentiation, little is known about how bone formation is epigenetically regulated. Histone methylation is an important process linked to the activation and repression of gene expression, thus it plays a critical role in epigenetic regulation of cell differentiation. While growing evidence indicates tht histone demethylases epigenetically regulate embryonic stem cell properties and functions, it is largely unknown what affect demethylases have on MSC differentiation and bone formation. To explore the role of demethylases in MSC differentiation, we systemically profiled the expression of histone demethylases in BMP-stimulated MSCs from bone marrow, as BMPs are potent inducers of osteogenic differentiation. We found that BMPs rapidly induced the expression of the lysine (K)-specific demethylase (KDM4B; also known as JMJD2B) that demethylates trimethylated histone H3 at lysine 9 (H3K9me3). H3K9me3 is a hallmark for gene silencing involved in growth and development. In general, a group of specific genes are activated when the stem cell differentiation program is triggered. Our preliminary studies demonstrated that KDM4B promoted osteogenic differentiation of MSCs while inhibiting adipogenic differentiation. Moreover, we found that the expression of Kdm4b was significantly down regulated in MSCs isolated from aging mice compared to young mice. Co-incidentally, H3K9me3 marks were significantly increased in osteoblasts of aging mice or ovariectomized mice. Based on these novel discoveries, in this application, we hypothesize that erasing H3K9me3 marks by KDM4B plays integral roles in osteogenic differentiation of MSCs in vitro and bone formation and in vivo. Three specific aims are proposed to test our hypothesis. Aim 1 is to determine whether KDM4B epigenetically regulate MSC lineage commitment through induction of DLX5. Aim 2 is to explore how erasing H3K9me3 by KDM4B coordinately regulates osteogenic differentiation of MSCs. Aim 3 is to determine whether KDM4B is required for bone formation in vivo and whether dysregulation of KDM4B impairs osteoblast function and bone formation in osteoporosis. Since histone demethylases are chemically modifiable, KDM4B may present as a novel therapeutic target for specifically controlling the differentiation of MSCs in regenerative medicine, and also lead to clues for new treatment in metabolic bone diseases such as, osteoporosis.