Targeting aging genes and pathways to promote optic nerve regeneration

针对衰老基因和途径促进视神经再生

基本信息

批准号：
10547815
负责人：
Mei Wan
金额：
$ 40.94万
依托单位：
JOHNS HOPKINS UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-01-01 至 2024-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10547815
关键词：
ATAC-seq Aging Axon Bioinformatics Biomedical Research Blood Vessels Brain Caenorhabditis elegans Catalytic Domain Cell Aging Cells Central Nervous System Chromatin Chromosome Fragility Consensus Corticospinal Tracts Cytoskeleton DNA Repair Data Development Enzymes Epigenetic Process Evolution FOXO3A gene FRAP1 gene Failure Genes Genetic Techniques Genetic Transcription Genomic Instability Heterochromatin Histones IGF-1 Signaling Pathway Insulin Insulin-Like Growth Factor I Knock-out Longevity Lysine Mediating Methylation Molecular Molecular Target Myosin Type II Natural regeneration Nerve Regeneration Neurodegenerative Disorders Neurons Noise Nonmuscle Myosin Type IIA Nutrient Optic Nerve Optic Nerve Injuries PIK3CG gene Pathway interactions Process Production Protocols documentation Publishing RNA-Directed DNA Polymerase Regenerative capacity Regulation Regulator Genes Regulatory Pathway Rejuvenation Retinal Ganglion Cells Role SIRT1 gene SOX11 gene STK11 gene Sensory Signal Transduction Spinal cord injury TP53 gene Telomerase Traumatic Brain Injury age effect aging gene axon growth axon injury axon regeneration c-myc Genes cell regeneration designer receptors exclusively activated by designer drugs detection of nutrient experimental study gene repression genetic manipulation histone methylation human disease in vivo regeneration induced pluripotent stem cell inhibitor innovation insight neural novel optic nerve regeneration overexpression screening sensor spinal cord regeneration telomere transcription factor transcriptome sequencing transcriptomics

项目摘要

Summary To date, targeting the genes regulating intrinsic axon growth ability have produced by far the most promising results in optic nerve regeneration. Recent studies, including ours, have provided strong evidence that neuronal aging might be a key converging process underlying the loss of intrinsic axon growth ability of CNS neurons. Indeed, many genes that act to regulate axon regeneration are also hallmark genes of aging (genomic instability, telomere attrition, epigenetic alteration, and nutrient sensing, etc.). First, recent studies and our preliminary results showed that c-Myc and p53, two well-known genes involved in DNA repair and genomic instability during aging, act to support optic nerve regeneration. Second, our preliminary study showed that telomerase reverse transcriptase (TERT) was necessary for sensory axon regeneration in vivo. Third, aging is often associated with decreased methylation of histone 3 at lysine 27 (H3K27) and increased methylation of H3K4, resulting in reduced amount of heterochromatin. In support, the level of H3K27 demethylase UTX increases during aging and knocking out UTX in c. elegans promotes longevity. Our unpublished study showed that knocking out UTX and its targeted gene, Magi3, in RGCs drastically promoted optic nerve regeneration. Fourth, the insulin and IGF-1 signaling (IIS) pathways, the key regulators of nutrient sensing, are the most conserved aging controlling pathway in evolution. IGF-1 and many IIS downstream targets, such as Pten/PI3K, Akt, and mTor, are all important regulators of optic nerve regeneration. Our published study and a recent study have shown that Sirt1 and LKB1, two important nutrient sensors, function to regulate sensory axon and spinal cord regeneration, respectively. Foxo3, another key target of Akt signaling, has recently been shown to promote vascular cell regeneration through Sirt1. Lastly, recent findings indicated that cellular reprogramming process can reverse aging and rejuvenate the cells. Importantly, manipulations of several reprogramming factors, such as KLF4 and Lin28, have been shown to promote optic nerve regeneration. Therefore, we hypothesize that aging regulatory genes/pathways can be manipulated to promote optic nerve regeneration through rejuvenation of mature CNS neurons. In Aim 1, we will determine if manipulation of miR-138/Sirt1, TERT, and Foxo3 in RGCs can promote optic nerve regeneration. In Aim 2, we will first determine if combination of these aging genes with myosin II knockout or enhanced neural activity would have synergistic effects on regeneration. We will then use RNA-seq and ATAC-seq of purified RGCs to explore how these aging genes regulate optic nerve regeneration. In Aim 3, by performing RNA-seq and ATAC-seq of purified RGCs at different developing, maturation, and aging stages, we will first use advanced integrative bioinformatics analyses to identify top candidate aging genes and their associated transcription factors, both of which act to orchestrate RGCs aging. We will then perform functional screening experiments to determine their roles in regulation of axon growth and optic nerve regeneration.

概括迄今为止，针对调节内在轴突生长能力的基因已经产生了迄今为止最有希望的导致视神经再生。最近的研究，包括我们的研究，提供了强有力的证据表明神经元衰老可能是中枢神经系统神经元内在轴突生长能力丧失的一个关键收敛过程。事实上，许多调节轴突再生的基因也是衰老的标志基因（基因组不稳定、端粒磨损、表观遗传改变和营养感应等）。首先，最近的研究和我们的初步结果表明，c-Myc 和 p53 这两个众所周知的基因参与 DNA 修复和基因组不稳定抗衰老，支持视神经再生。其次，我们的初步研究表明端粒酶逆转转录酶（TERT）对于体内感觉轴突再生是必需的。第三，衰老往往与组蛋白 3 在赖氨酸 27 (H3K27) 处的甲基化减少，H3K4 的甲基化增加，从而导致异染色质的量。作为支持，H3K27 去甲基化酶 UTX 的水平在衰老过程中增加，并且在c中敲除UTX。秀丽隐杆线虫可以延年益寿。我们未发表的研究表明，敲除 UTX 和其 RGC 中的目标基因 Magi3 极大地促进了视神经再生。四、胰岛素和IGF-1 信号传导（IIS）途径是营养感应的关键调节因子，是最保守的衰老控制途径进化中的途径。 IGF-1 和许多 IIS 下游目标，例如 Pten/PI3K、Akt 和 mTor，都是视神经再生的重要调节因子。我们发表的研究和最近的一项研究表明 Sirt1 和LKB1，两种重要的营养传感器，具有调节感觉轴突和脊髓再生的功能，分别。 Foxo3 是 Akt 信号传导的另一个关键靶标，最近被证明可以促进血管细胞生长通过 Sirt1 进行再生。最后，最近的研究结果表明，细胞重编程过程可以逆转延缓衰老并恢复细胞活力。重要的是，对几个重编程因子的操纵，例如 KLF4 和 Lin28 已被证明可以促进视神经再生。因此，我们假设老龄化监管可以操纵基因/通路，通过成熟中枢神经系统的复兴来促进视神经再生神经元。在目标 1 中，我们将确定在 RGC 中操纵 miR-138/Sirt1、TERT 和 Foxo3 是否可以促进视神经再生。在目标 2 中，我们将首先确定这些衰老基因是否与肌球蛋白 II 结合敲除或增强的神经活动将对再生产生协同作用。然后我们将使用RNA-seq 对纯化的 RGC 进行 ATAC-seq，以探索这些衰老基因如何调节视神经再生。在目标 3 中，通过对不同发育、成熟和衰老阶段的纯化 RGC 进行 RNA-seq 和 ATAC-seq，我们将首先使用先进的综合生物信息学分析来识别最重要的候选衰老基因及其相关的转录因子，两者都可以协调 RGC 的衰老。然后我们将执行功能筛选实验以确定它们在调节轴突生长和视神经再生中的作用。