Elucidating the gene regulatory networks that drive neural regeneration

阐明驱动神经再生的基因调控网络

• 批准号: 10541717
• 负责人: Jared A Tangeman
• 金额: $ 4.07万
• 依托单位: MIAMI UNIVERSITY OXFORD
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-07-01 至 2023-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10541717
• 关键词: ASCL1 gene; ATAC-seq; Acute; Address; Adult; Amphibia; Animal Model; Automobile Driving; Behavior; Binding; Binding Sites; Bioinformatics; Biological Assay; Cell Differentiation process; Cell Nucleus; Cells; Characteristics; Chickens; Chromatin; Competence; Data; Data Analyses; Data Set; Development; Development Plans; Embryo; Environment; Excision; FGF2 gene; Foundations; Future; Gene Activation; Gene Expression; Gene Expression Profiling; Gene Expression Regulation; Genes; Genetic Transcription; Genomics; Goals; Grant; Homeobox; Human; Injury; Intervention; Modality; Modeling; Moderate Activity; Multiomic Data; Natural regeneration; Nerve Regeneration; Neural Retina; Neuraxis; Neurons; Organism; Outcome; Output; Phase; Pigmentation physiologic function; Population; Positioning Attribute; Postdoctoral Fellow; Principal Investigator; Regenerative capacity; Regenerative response; Regulator Genes; Replacement Therapy; Research; Research Training; Resolution; Retina; Role; Route; Small Nuclear RNA; Source; Structure of retinal pigment epithelium; System; Techniques; Time; Training and Infrastructure; Vertebrates; Work; Writing; Zebrafish; career; career development; cell type; computer infrastructure; data handling; effective therapy; epigenomics; experience; gene regulatory network; genome-wide; in silico; innovation; insight; mouse model; multiple omics; neurogenesis; neuron regeneration; novel; novel strategies; outreach; pedagogical content; programs; regenerative; relating to nervous system; retinal damage; retinal neuron; skills; transcription factor; transcriptome sequencing; transcriptomics; treatment response

Project Summary / Abstract There are no effective treatments to replace damaged retinal neurons, reflecting a fundamental inability for humans to mount robust regenerative responses within the central nervous system. To address this, it will be critical to understand the diversity of gene regulatory mechanisms that can impede or drive neuron regeneration across vertebrate contexts. Embryonic amniotes have a transitory ability to regenerate retinal neurons from cells of the retinal pigment epithelium (RPE) if supplied with exogenous FGF2 at the time of retinal injury. This mechanism of regeneration can be readily induced at embryonic day 4 (E4) of chicken development, but RPE neural competence is lost by embryonic day 5 (E5). The overarching objective of the proposed research is to profile changes in gene regulation that dispossess RPE cells of their neural competency as they differentiate. Specific aim 1 will interrogate transcription factor regulatory activity within RPE cells across the E4 / E5 developmental window by integrating gene expression analysis, chromatin accessibility profiling, and transcription factor binding assays. Preliminary bulk and single nuclei RNA-seq datasets revealed the acute activation of neural retina transcription factor profiles at both E4 and E5, such as PAX6, ASCL1, and VSX2. In contrast, genes associated with RPE maturity, such as OTX2 and pigmentation genes, were elevated in the E5 RPE independently of retinectomy and FGF2 treatment. Similarly, chromatin accessibility suggested wider dysregulation of OTX2 and related homeobox transcription factor binding sites. During the 1-year F99 phase, OTX2 binding activity will be profiled in intact and FGF2-treated RPE cells at E4 and E5 stages. Additionally, single nuclei RNA-sequencing will capture the heterogeneous transcriptional states of RPE cells during differentiation and FGF2 treatment response at E4 and E5. These results will be integrated to into a model that describes how changes in the RPE gene regulatory landscape culminates in a loss of neural competency. In specific aim 2, the gene regulatory networks present in adult vertebrate models of central nervous system regeneration will be interrogated to inspire novel routes for the induction of mammalian regeneration. This K00 phase will focus on the development of key research skills, including multi-omics data analysis approaches, techniques for spatial transcriptomics and single cell epigenomics, and cross-species genomics / transcriptomics. Up to 4 years will be spent on the K00 phase in an environment directly supportive of these applications. Specific professional development objectives will be concurrently pursued, such as pedagogical development, diversity outreach initiatives, and grant writing. Together, these aims encompass a career development plan that will lead to formation of an independent research program and result in impactful research focused on expanding human central nervous system regenerative capacity.

项目总结/摘要 目前还没有有效的治疗方法来取代受损的视网膜神经元，这反映了视网膜神经元的根本性无能。 人类在中枢神经系统内建立强大的再生反应。为了解决这个问题， 这对于理解能够阻碍或驱动神经元再生的基因调控机制的多样性至关重要 跨脊椎动物的环境。胚胎视网膜神经元具有从细胞再生视网膜神经元的短暂能力 如果在视网膜损伤时提供外源性FGF 2，则视网膜色素上皮（RPE）的生长受到抑制。这 在鸡胚发育的第4天（E4），再生机制可以容易地诱导，但RPE 神经能力在胚胎第5天（E5）丧失。拟议研究的总体目标是 基因调控谱的变化，剥夺了RPE细胞的神经能力，因为他们分化。 特异性目的1将询问跨越E4 / E5的RPE细胞内的转录因子调节活性。 通过整合基因表达分析，染色质可及性分析， 转录因子结合测定。初步的批量和单核RNA-seq数据集揭示了急性 在E4和E5时激活神经视网膜转录因子谱，如PAX 6、ASCL 1和VSX 2。在 相反，与RPE成熟相关的基因，如OTX 2和色素沉着基因，在E5中升高， RPE独立于视网膜切除术和FGF 2治疗。同样，染色质可及性表明更广泛的 OTX 2和相关同源框转录因子结合位点的失调。在为期一年的F99阶段， 将在E4和E5阶段的完整和FGF 2处理的RPE细胞中分析0 TX 2结合活性。此外，本发明还 单核RNA测序将捕获RPE细胞的异质性转录状态， 在E4和E5时的分化和FGF 2治疗反应。这些结果将被整合到一个模型中， 描述了RPE基因调控景观的变化如何导致神经能力的丧失。在 具体目标2、成年脊椎动物中枢神经系统模型中存在的基因调控网络 再生将被询问，以激发新的途径诱导哺乳动物再生。这个K 00 阶段将侧重于发展关键的研究技能，包括多组学数据分析方法， 空间转录组学、单细胞表观基因组学和跨物种基因组学技术 转录组学在直接支持这些的环境中，K 00阶段将花费长达4年的时间 应用.具体的专业发展目标将同时进行，如教学 发展，多样性外展倡议，和赠款写作。总之，这些目标包括一个职业生涯 发展计划，这将导致形成一个独立的研究计划，并导致有影响力的研究 专注于扩大人类中枢神经系统的再生能力。