The essential role of miR-27a in craniofacial and body skeletons.

标题 miR-27 a在颅面和身体骨骼中的重要作用 摘要 MicroRNA是一种约22个核苷酸的非编码RNA，在转录后水平控制基因表达。 由于它能够同时调节大量基因，因此microRNA被认为是起作用的。 作为细胞类型特异性发育和功能的主调节剂。虽然功能增益分析 microRNA过表达可以影响转基因小鼠的这些过程，功能丧失的研究往往不能 检测表型改变，并且无法忠实地确定它们在发育和疾病中的作用。 此外，从体外分析获得的结果有时不支持在体内的功能分析。 Dicer是一种对microRNA生物合成至关重要的酶，其研究表明其在骨中的重要性 重塑然而，涉及的确切miRNA尚未确定。miR-23 a ~ 27 a ~24-2的簇 由三种由单个转录物产生的miRNAs组成。miR-23 a和miR-27 a的调节异常已经被证实。 在骨质疏松症患者中显示，但它们的调节作用仍然难以捉摸。在细胞培养研究中， miR-23 a或miR-27 a抑制成骨细胞分化，表明它们是骨形成的负调控因子。 在小鼠中，OB特异性表达miR-23 a ~ 27 a ~24-2簇揭示了其对骨细胞的影响，但对OB无影响 分化动物研究不支持细胞培养分析。迫切需要发展 功能丧失模型，以明确评估这些microRNA在体内的功能。来填补我们的知识 为了弥补这些空白，我们使用CRSPR-Cas9基因编辑创建了一个miR-27 a缺陷的小鼠模型。失去这位 单个miRNA导致颅面和身体骨骼的严重骨质疏松症，这表明 miR-27 a不能被取代。随着年龄的增长，骨丢失表型也变得更加突出。新 遗传学证据表明，miR-27 a是骨重建的正调节因子，强烈反对 以前的细胞培养研究在本申请中，我们将表征miR-27 a中的颅面和骨骼缺陷， 缺陷小鼠我们将检查与突变相关的成骨细胞和破骨细胞异常， 骨骼重塑失衡进一步阐明miR-27 a调控骨的机制 形成和再吸收，我们将确定其直接目标使用无偏筛选。完成这一 该提案具有突出的潜力，以推进我们的知识基础的表观遗传调控骨 代谢，导致新的战略，预防和治疗颅面和骨骼疾病。

Title The essential role of miR-27a in craniofacial and body skeletons Abstract MicroRNAs are ~22 nucleotides noncoding RNAs which control gene expression at post-transcriptional levels. Owing to its ability to simultaneously modulate a vast amount of genes, microRNA has been postulated to work as a master regulator for cell type-specific development and function. Although gain-of-function analysis with microRNA overexpression can affect these processes in transgenic mice, loss-of-function study often fails to detect phenotypic alterations and is unable to faithfully determine their role in development and disease. Moreover, results obtained from in vitro analysis sometimes are not supported by functional analyses in vivo. Studies of Dicer, an enzyme essential for biosynthesis of microRNAs, have implicated their importance in bone remodeling. However, the exact miRNA(s) involved has not been identified. The cluster of miR-23a~27a~24-2 consists of three miRNAs generated from a single transcript. Dysregulation of miR-23a and miR-27a has been shown in osteoporosis patients but their regulatory role remains elusive. In cell culture study, high levels of miR-23a or miR-27a inhibits OB differentiation, suggesting that they are negative regulators for bone formation. In mice, OB-specific expression of miR-23a~27a~24-2 cluster reveals its effects on osteocyte but not OB differentiation. The animal study does not support the cell culture analysis. There is an urgent need to develop loss-of-function models to definitively assess the function of these microRNAs in vivo. To fill our knowledge gaps, we created a mouse model deficient for miR-27a using CRSPR-Cas9 gene editing. The loss of this single miRNA causes severe osteoporosis in the craniofacial and body skeletons, suggesting the function of miR-27a cannot be substituted. The bone loss phenotype also becomes more prominent with age. New genetic evidence, indicating miR-27a as a positive regulator in bone remodeling, strongly argues against the previous cell culture study. In this application, we will characterize craniofacial and skeletal defects in miR-27a deficient mice. We will examine osteoblast and osteoclast abnormalities associated with the mutation to cause an imbalance in skeletal remodeling. To further elucidate the mechanism by which miR-27a regulates bone formation and resorption, we will identify its direct targets using unbiased screening. The completion of this proposal has outstanding potential to advance our knowledge base of epigenetic regulation in bone metabolism, leading to novel strategies for prevention and treatment of craniofacial and skeletal disorders.