Transcriptional regulation over neurogenesis of cortical output neuron segmental identity and diversity

皮质输出神经元节段同一性和多样性的神经发生的转录调控

• 批准号: 10638147
• 负责人: Vibhu Vinodchandra Sahni
• 金额: $ 46.38万
• 依托单位: WINIFRED MASTERSON BURKE MED RES INST
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-03-01 至 2028-02-29
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10638147
• 关键词: AT Rich Sequence; ATAC-seq; Adult; Affect; Age; Amyotrophic Lateral Sclerosis; Axon; Binding Proteins; Birth; Brain Stem; Cells; Cerebral Palsy; Corticospinal Tracts; Data; Development; Disease model; Embryo; Embryonic Development; Foundations; Gene Delivery; Genes; Genetic Transcription; Goals; Health Care Costs; Human; In Vitro; Individual; Injections; Injury; Investigation; Knock-in; Knock-in Mouse; Knockout Mice; Label; Life; Link; Mediating; Molecular; Molecular Target; Motor Neuron Disease; Motor Neurons; Motor output; Movement; Mus; Natural regeneration; Neocortex; Nervous System; Neurons; Output; Paralysed; Pathway interactions; Play; Population; Process; Public Health; Quality of life; Recovery; Recovery of Function; Regulation; Reporter; Research; Resolution; Role; Specific qualifier value; Specificity; Spinal; Spinal Cord; Spinal cord injury; Stroke; Testing; Time; Transcriptional Regulation; Transgenic Organisms; Traumatic Brain Injury; Vertebral column; Work; conditional knockout; cost; directed differentiation; experimental study; functional plasticity; gain of function; high-throughput drug screening; in utero; in vivo; loss of function; loved ones; motor control; motor impairment; motor recovery; neocortical; neurogenesis; neuropeptide Y; novel; overexpression; personalized approach; post stroke; postmitotic; postnatal; regenerative; repaired; single-cell RNA sequencing; skills; spasticity; stem cell differentiation; stroke recovery; transcription factor; transcriptomics; translational neuroscience

Subcerebral projection neurons (SCPN) reside in the neocortex, and extend axons to subcerebral targets in the brainstem and spinal cord. Corticospinal neurons (CSN), a subclass of SCPN, project to the spinal cord and their axons form the corticospinal tract (CST), a critical circuit for voluntary motor control. In addition, projections from the neocortex to brainstem targets function in parallel with CSN to exert motor control. Degeneration of these projection neurons in amyotrophic lateral sclerosis (ALS), along with degeneration of spinal motor neurons, causes spasticity and paralysis. In humans, damage to the CST after spinal cord injury is a principal cause of loss of voluntary motor control. Further, integrity of corticospinal connectivity is centrally linked to recovery from stroke and cerebral palsy. There have been multiple investigations detailing the role of cortico-brainstem vs. corticospinal projections in both motor control, as well as their distinct contributions to functional recovery in these distinct causes of paralysis. However, it remains unclear when and how these distinct projections are established during development. Understanding the molecular basis of this specification and differentiation during development therefore holds significant promise in establishing approaches that are tailored to enhancing plasticity of these related, yet distinct circuits. A necessary first step toward this ultimate goal is to identify the molecular mechanisms directing SCPN axons to brainstem (cortico-brainstem neurons) versus spinal cord (CSN). Ongoing work in our lab has identified that such projections are initially specified during the process of axon extension during development. We have identified that cortico-brainstem and corticospinal neurons express distinct genes and can be molecularly distinguished in mice by birth. Further our data suggest that the transcriptional regulator Satb2 acts, in part, to specify cortico-brainstem neurons. This proposal investigates the hypothesis that transcriptional regulation controls the development of cortico-brainstem vs. corticospinal projections by late embryonic development in mice. Building on this foundation, we will first identify the time point in development when these projections are specified using knock-in Cre reporter mice (Aim 1). This will be tested using intracerebral injections of AAV-reporters at distinct developmental times in utero with adult analysis of axonal projections. In Aim 2, we will investigate the transcriptional targets of Satb2 in SCPN at later developmental times by profiling all SCPN in Satb2 WT and conditional KO mice, as well as by Satb2 overexpression, at single cell resolution. Finally, in Aim 3 using subpopulation-specific transgenic Cre lines, we will investigate SCPN axon targeting in both Satb2 loss- and gain-of-function. In addition, we will investigate whether misexpression of Satb2 target genes can alter SCPN targeting to the brainstem vs. spinal targets. Together, our work will discern in-depth, the mechanisms of when and how transcriptional regulation controls SCPN segmental “identity” thereby providing a mechanistic framework for subsequent identification of molecules controlling segmentally appropriate SCPN connectivity with subcerebral targets.

大脑下投射神经元（SCPN）位于新皮层中，并将轴突延伸到大脑下靶区， 脑干和脊髓皮质脊髓神经元（CSN）是SPN的一个亚类，投射到脊髓及其周围 轴突形成皮质脊髓束（CST），这是自主运动控制的关键回路。此外，预计 脑干靶向的新皮层与CSN并行地发挥作用以施加运动控制。这些退化 肌萎缩侧索硬化症（ALS）中的投射神经元，沿着脊髓运动神经元的变性， 导致痉挛和瘫痪在人类中，脊髓损伤后对CST的损伤是脊髓损伤的主要原因。 失去自主运动控制。此外，皮质脊髓连接的完整性与从脊髓损伤中恢复密切相关。 中风和脑瘫。已经有多项研究详细说明了皮质脑干与 皮质脊髓投射在运动控制中，以及它们对功能恢复的独特贡献， 这些不同的瘫痪原因。然而，目前尚不清楚这些不同的预测何时以及如何发生。 在发展过程中建立。了解本规范和差异的分子基础 因此，在制定适合于增强 这些相关但不同的回路的可塑性。实现这一最终目标的必要的第一步是确定 将SCPN轴突导向脑干（皮质-脑干神经元）与脊髓的分子机制 （CSN）。我们实验室正在进行的工作已经确定，这种预测最初是在 轴突在发育过程中延伸。我们已经确定皮质脑干和皮质脊髓神经元表达 不同的基因，可以通过出生在小鼠中进行分子区分。进一步我们的数据表明， 转录调节子Satb 2部分地起作用以指定皮质-脑干神经元。该提案调查了 转录调控控制皮质-脑干与皮质-脊髓发育的假说 小鼠胚胎发育后期的预测。在此基础上，我们将首先确定时间点 当使用敲入Cre报告小鼠指定这些投射时，在发育中（Aim 1）。这将受到考验 使用在子宫内不同发育时间脑内注射AAV报告基因， 轴突投射在目标2中，我们将在稍后的研究中研究SCPN中Satb 2的转录靶点。 通过分析Satb 2 WT和条件性KO小鼠中的所有SCPN以及Satb 2 过表达，在单细胞分辨率。最后，在目标3中，使用亚群特异性转基因Cre系，我们 将研究SCPN轴突靶向Satb 2功能丧失和获得。此外，我们将调查 Satb 2靶基因的错误表达是否可以改变SCPN靶向脑干与脊髓靶点。 总之，我们的工作将深入了解转录调控何时以及如何控制的机制。 SCPN节段“同一性”，从而为随后的分子鉴定提供了一个机制框架 控制节段性适当的SCPN与大脑下目标的连接。