Modulation of Runx2 activity by arginine methylation

项目摘要/摘要 蛋白质精氨酸甲基化是一个迅速发展的领域，在癌症中的重要性日益得到承认，但 对于骨骼中的这种机制，人们几乎一无所知。这项建议是基于对地球的新发现。 Runx2的精氨酸甲基化。我们已经确定了两种甲基化的蛋白质精氨酸甲基转移酶 在四个已定义的精氨酸残基上运行2。通过将这些残基突变为模拟甲基化的丙氨酸- 缺乏状态，我们已经证明了在Runx2介导的Runx2精氨酸甲基化中所需的 癌细胞的运动性。在这里，我们将研究Runx2精氨酸甲基化在其在 成骨细胞分化和骨形成。目的1是为了刻画一过性精氨酸甲基化的特征 成骨细胞培养模型中Runx2的表达。我们还将比较WT Runx2和WT Runx2的成骨潜力 不可甲基化的突变体Runx2，以及操纵介导Runx2的甲基转移酶的后果 甲基化。目的2探讨Runx2精氨酸甲基化在调节生理和功能中的作用 与BMP-Smads通路的相互作用。最后，目标3是定义Runx2的精氨酸甲基化，以及 在活体颅骨发育过程中介导Runx2甲基化的甲基转移酶的表达 比较WT和不可甲基化的Runx2在颅骨器官培养中的成骨潜力。总体而言，这 该方案提出了一种假设，即在成骨过程中Runx2的活性需要精氨酸甲基化 BMP-Smad信号反应的分化和必需性。此外，我们还在研究中纳入了 设计基因组和蛋白质组分析以促进对Runx2蛋白-蛋白质的无偏见发现 受其精氨酸甲基化影响的相互作用、Runx2基因组靶点和Runx2调节的基因。 从长远来看，我们设想将从这个探索性项目中获得的知识转化为 了解骨骼发育、动态平衡和再生，并提供新的治疗方法 解决骨骼疾病的方法。

PROJECT SUMMARY / ABSTRACT Protein arginine methylation is a rapidly growing field with increasingly recognized importance in cancer, but almost nothing is known about this mechanism in bone. This proposal is based on novel discoveries on the arginine methylation of Runx2. We have identified two protein arginine methyltransferases that methylate Runx2 on four defined arginine residues. By mutating these residues to alanine that mimic the methylation- deficient state, we have demonstrated the requirement for Runx2 arginine methylation in Runx2-mediated cancer cell motility. Here, we will investigate Runx2 arginine methylation in the context of its canonical role in osteoblast differentiation and bone formation. Aim 1 is to characterize the temporal arginine methylation of Runx2 in osteogenic cell culture models. We will also compare the osteogenic potentials of WT Runx2 versus unmethylatable mutant Runx2, and the consequences of manipulating methyltransferases that mediate Runx2 methylation. Aim 2 addresses the role of Runx2 arginine methylation in modulating the physical and functional interaction with BMP-Smads pathway. Finally, Aim 3 is to define the arginine methylation of Runx2, and the expression of methyltransferases that mediate Runx2 methylation during skull development in vivo, and to compare the osteogenic potential of WT versus unmethylatable Runx2 in calvarial organ cultures. Overall, this proposal addresses the hypothesis that arginine methylation is required for Runx2 activity during osteogenic differentiation and essential for BMP-Smad signaling response. Additionally, we have incorporated in the study design genomic and proteomic analyses to facilitate unbiased discoveries of Runx2 protein-protein interactions, Runx2 genomic targets and Runx2-regulated genes that are impacted by its arginine methylation. In the long run, we envision translation of knowledge acquired in this exploratory project to improve the understanding of bone development, homeostasis and regeneration, and provide novel therapeutic approaches to tackle skeletal disorders.