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

miR-27a 在颅面和身体骨骼中的重要作用。

• 批准号: 9900768
• 负责人: Takamitsu Maruyama
• 金额: $ 16.12万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-04-01 至 2021-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9900768
• 关键词: Affect; Age; Anabolism; Animals; Bioinformatics; Bone Resorption; Bone remodeling; CRISPR/Cas technology; Cell Culture Techniques; Cells; Data; Defect; Development; Dicer Enzyme; Disease; Elderly; Elements; Epigenetic Process; Expression Profiling; Gene Expression; Genes; Genetic; Genetic Models; Genetic Transcription; Health; Hematopoietic; Homeostasis; Human; In Vitro; Knowledge; Maintenance; Measures; Mediating; Metabolic Diseases; MicroRNAs; Modeling; Molecular; Mouse Strains; Mus; Mutation; Nucleotides; Osteoblasts; Osteoclasts; Osteocytes; Osteogenesis; Osteoporosis; Pathogenesis; Patients; Phenotype; Prevention strategy; Process; Regulation; Role; Skeletal Development; Skeleton; TNFSF11 gene; Testing; Therapeutic; Transcript; Transgenic Mice; Untranslated RNA; Woman; Work; bone; bone fragility; bone loss; bone metabolism; cell type; comparative; craniofacial; craniofacial disorder; design; epigenetic regulation; fracture risk; gain of function; in vivo; insight; knowledge base; loss of function; men; mouse genetics; mouse model; novel; novel strategies; osteoblast differentiation; osteoclastogenesis; overexpression; paracrine; progenitor; screening; skeletal; skeletal disorder; stem cells; transcriptome; transcriptome sequencing

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.

标题 MiR-27A在头面部和身体骨骼中的重要作用 摘要 MicroRNAs是一种~22个核苷酸的非编码RNA，在转录后水平控制基因的表达。 由于它能够同时调节大量的基因，所以微核糖核酸被认为是起作用的 作为细胞类型特定发育和功能的主调节因子。尽管函数增益分析具有 在转基因小鼠中，microRNA的过度表达可以影响这些过程，功能丧失研究往往无法 检测表型变化，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能得到体内功能分析的支持。 DICER是一种对microRNAs的生物合成至关重要的酶，它的研究暗示了它们在骨骼中的重要性 改建。然而，涉及的确切miRNA(S)尚未确定。MiR-23A~27A~24-2星系团 由一个转录本产生的三个miRNA组成。MiR-23a和miR-27a的调节失调 在骨质疏松症患者中表现出来，但它们的调节作用仍然难以捉摸。在细胞培养研究中，高水平的 MiR-23a或miR-27a抑制成骨细胞分化，提示它们是骨形成的负调控因子。 在小鼠中，miR-23a~27A~24-2簇的OB特异性表达揭示了其对骨细胞的影响，但对OB没有影响 差异化。动物实验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microRNAs在体内的功能。为了填满我们的知识 为了弥补缺口，我们使用CRSPR-Cas9基因编辑创建了一个miR-27A缺陷的小鼠模型。失去这一点 单个miRNA会导致头面部和身体骨骼严重的骨质疏松，提示miRNA的功能 MIR-27A是不可替代的。随着年龄的增长，骨丢失的表型也变得更加突出。新的 遗传证据表明miR-27A在骨重建中是一个积极的调节因素，强烈反对 先前的细胞培养研究。在这个应用中，我们将在miR-27A中描述颅面和骨骼缺陷的特征 有缺陷的小鼠。我们将检查与突变相关的成骨细胞和破骨细胞异常，以引起 骨骼重塑的不平衡。为了进一步阐明miR-27A调节骨骼的机制 对于形成和吸收，我们将使用无偏筛选来确定其直接靶点。这项工程的完成 该提案具有显著的潜力，可以提高我们关于骨骼表观遗传调控的知识基础 新陈代谢，导致了预防和治疗头面部和骨骼疾病的新策略。