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。 MSC 是多能祖细胞，具有自我更新能力和多谱系分化潜力，包括成骨、软骨形成和脂肪形成。尽管在理解间充质干细胞分化的转录控制方面已经取得了重大进展，但对于骨形成如何通过表观遗传调控仍知之甚少。组蛋白甲基化是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发挥着关键作用。虽然越来越多的证据表明组蛋白去甲基酶在表观遗传上调节胚胎干细胞的特性和功能，但目前尚不清楚去甲基酶对 MSC 分化和骨形成的影响。为了探讨去甲基酶在 MSC 分化中的作用，我们系统地分析了 BMP 刺激的骨髓 MSC 中组蛋白去甲基酶的表达，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) 特异性去甲基化酶（KDM4B；也称为 JMJD2B）的表达，该酶使组蛋白 H3 在赖氨酸 9 (H3K9me3) 处去甲基化。 H3K9me3 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，一组特定基因被激活。我们的初步研究表明，KDM4B 促进 MSC 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 Kdm4b 的表达显着下调。巧合的是，衰老小鼠或卵巢切除小鼠的成骨细胞中 H3K9me3 标记显着增加。基于这些新发现，在本申请中，我们假设 KDM4B 擦除 H3K9me3 标记在 MSC 体外成骨分化以及体内骨形成中发挥着不可或缺的作用。 提出了三个具体目标来检验我们的假设。目标 1 是确定 KDM4B 是否通过诱导 DLX5 在表观遗传上调节 MSC 谱系定型。目标2是探索KDM4B擦除H3K9me3如何协调调节MSCs的成骨分化。目标 3 是确定 KDM4B 是否是体内骨形成所必需的，以及 KDM4B 失调是否会损害骨质疏松症中的成骨细胞功能和骨形成。由于组蛋白去甲基酶是可化学修饰的，KDM4B可能作为再生医学中特异性控制间充质干细胞分化的新治疗靶点，也为骨质疏松症等代谢性骨疾病的新治疗提供线索。

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.