The role of microRNAs in Schwann cell development and disease

microRNA 在雪旺细胞发育和疾病中的作用

基本信息

批准号：
8653995
负责人：
Rajeshwar B Awatramani
金额：
$ 31.88万
依托单位：
NORTHWESTERN UNIVERSITY AT CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-07-01 至 2015-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8653995
关键词：
Ablation Architecture Axon Biological Assay Cell Culture Techniques Cell Differentiation process Cell Lineage Cells Co-Immunoprecipitations Coculture Techniques Commit Complex Demyelinating Diseases Development Developmental Gene Dicer Enzyme Disease Enzymes Equilibrium Event Gene Expression Gene Expression Profiling Genes Genetic Genetic Programming Grant Human Image JUN gene Light Luciferases Malignant Neoplasms Mediating MicroRNAs Modeling Molecular Mus Myelin Myelin Sheath Natural regeneration Nerve Nervous system structure Neural Crest Cell Neurofibromatoses Neuroglia Neurons Neuropathy Nucleotides Pathway interactions Peripheral Peripheral Nerves Peripheral Nervous System Phenocopy Phenotype Play Process RNA Regulator Genes Regulatory Pathway Reporter Repression Role Schwann Cells Staging System Testing base cell dedifferentiation cohort design dysmyelination gene repression human DICER1 protein image processing mutant myelination nerve injury novel therapeutics overexpression periaxin postnatal programs public health relevance research study role model transcription factor

项目摘要

DESCRIPTION (provided by applicant): Schwann cell differentiation is thus a multistage process, the progression from one stage to the next being driven by dynamic alterations in underlying genetic programs. We hypothesize that microRNAs, being important developmental regulators in other systems, will be critical for SC developmental transitions. To evaluate the function of microRNAs in Schwann cell (SC) development we have conditionally ablated the microRNA processing enzyme, dicer. SC lacking dicer are stalled during differentiation and do not produce myelin, phenocopying models of congenital hypomyelination. Here in Specific Aim1, we will perform a detailed ultrastructural and molecular characterization of these SC lacking dicer. Preliminary analyses suggest that these SC are arrested at the promyelinating-myelinating transition, maintain the expression of Sox2, and lack the SC master regulator gene Egr2 (Krox20). To prove that loss of Egr2 is the underlying reason for dysmyelination, we will determine if forced expression of Egr2 can circumvent the phenotype of SC lacking dicer. Finally, we will generate a mosaic deletion of dicer, to verify that the changes observed in the dicer cKO are cell autonomous. In Specific Aim 2, we will perform microRNA profiling in developmental and pathological scenarios. We will identify key microRNAs that are dynamically expressed and then bioinformatically determine their candidate targets. We will then verify microRNA-target interactions using luciferase reporter assays in heterologous cells, argonaute co-immunoprecipitation, and overexpression in SC-neuron coculture systems. Finally we will determine whether some key microRNAs are themselves activated by Egr2, testing the hypothesis that Egr2 represses antecedent gene programs in SC with the help of microRNAs. Together these studies will be important in understanding the role of microRNAs in development. Further, since the processes underlying SC differentiation are considered a "mirror image" of SC dedifferentiation, these studies will not only shed light on developmental transitions but also on converse transitions that occur in peripheral myelin disorders and nerve injury.

描述（由申请人提供）：施万细胞分化因此是一个多阶段过程，从一个阶段到下一阶段的进展是由潜在遗传程序的动态改变驱动的。我们假设 microRNA 作为其他系统中重要的发育调节因子，对于 SC 发育转变至关重要。为了评估 microRNA 在雪旺细胞 (SC) 发育中的功能，我们有条件地消除了 microRNA 加工酶 dicer。缺乏切丁酶的 SC 在分化过程中停滞不前，不产生髓磷脂，这是先天性髓鞘形成不足的表型模型。在具体目标 1 中，我们将对这些缺乏切块机的 SC 进行详细的超微结构和分子表征。初步分析表明，这些 SC 在髓鞘形成-髓鞘形成转变过程中被阻滞，维持 Sox2 的表达，并且缺乏 SC 主调节基因 Egr2 (Krox20)。为了证明Egr2的缺失是髓鞘形成障碍的根本原因，我们将确定Egr2的强制表达是否可以规避缺乏dicer的SC表型。最后，我们将生成切丁机的马赛克删除，以验证切丁机cKO中观察到的变化是细胞自主的。在具体目标 2 中，我们将在发育和病理情况下进行 microRNA 分析。我们将识别动态表达的关键 microRNA，然后通过生物信息学确定其候选靶标。然后，我们将使用异源细胞中的荧光素酶报告基因测定、argonaute 免疫共沉淀和 SC 神经元共培养系统中的过表达来验证 microRNA 与靶点的相互作用。最后，我们将确定一些关键的 microRNA 本身是否被 Egr2 激活，检验 Egr2 在 microRNA 的帮助下抑制 SC 中先行基因程序的假设。这些研究对于理解 microRNA 在发育中的作用非常重要。此外，由于 SC 分化的过程被认为是 SC 去分化的“镜像”，因此这些研究不仅将揭示发育转变，而且还将揭示外周髓鞘疾病和神经损伤中发生的逆转变。