Chemical Optoepigenetic Regulation of Chromatin-Mediated Neuroplasticity

描述（由申请人提供）：越来越多的证据表明，表观遗传机制在人类健康和疾病的各个方面发挥着关键作用，包括神经退行性疾病和神经精神疾病。这种作用包括从神经发生到突触发生的基本细胞过程，对开发治疗和理想地预防疾病病理生理学的新疗法具有重要意义。然而，尽管我们观察表观基因组和转录组的能力有了巨大的进步，但由于大多数现有的药理学探针的多效性和缺乏合适的遗传工具，我们扰乱表观基因组并以精确的时间控制和空间分辨率操纵转录程序的能力仍然受到严重限制。为了克服这些局限性，并能够在神经回路中靶向特定的细胞类型，我们提出了一种综合的多学科方法--从合成化学到神经生物学--将创新的、可扩展的“化学光表观遗传”技术与人类和小鼠神经元的表观基因组和转录组分析相结合。我们的神经调节策略利用具有快速热弛豫动力学的光开关化合物，其具有与其表观遗传靶点的缓慢结合动力学。使用I类组蛋白去乙酰化酶（HDAC）家族，包含染色质修饰复合物，我们的工作已经证明是染色质介导的神经可塑性的关键调节剂，以推进这种方法的测试，拟议项目的具体目标是：1）合成、表征和优化物理性质，能够光依赖性抑制含有不同I类HDAC同种型的神经元染色质修饰复合物的脱乙酰酶活性的光表观遗传探针的生物化学效力和选择性; 2）在不同HDAC复合物的精确时间操作后，确定培养的人干细胞衍生的神经元中的表观基因组和转录组变化;以及3）使用新的光表观遗传学探针来时间操作空间限定的小鼠神经元的表观基因组，以增强突触发生并调节海马回路功能。总的来说，通过在HDAC活性的时空控制方面提供显著的改进，结合在产生同种型和复合物选择性HDAC抑制剂方面的进展，我们预计我们的方法将限制目前可用的小分子工具的多效性作用。通过选择性操纵神经回路特定区域的表观基因组，我们预期能够显著提高我们对表观遗传状态的特定时间调节如何影响神经可塑性的理解，并能够描述神经回路内定义的神经元亚型中表观遗传机制的贡献。重要的是，我们开发化学光表观遗传探针的方法广泛适用于操纵表观遗传调控机制，并且可以扩展以组装用于组合光表观遗传研究的分子工具包。这些工具在神经表观遗传学领域具有广泛的适用性，并有助于推进开发针对神经可塑性的改进疗法的努力。

DESCRIPTION (provided by applicant): Growing evidence points to a critical role for epigenetic mechanisms in diverse aspects of human health and disease, including neurodegenerative and neuropsychiatric disorders. This role includes fundamental cellular processes ranging from neurogenesis to synaptogenesis, with significant implications for the development of novel therapeutics to treat and ideally prevent disease pathophysiology. However, despite dramatic advances in our ability to observe the epigenome and transcriptome, our ability to perturb the epigenome and manipulate transcriptional programs with precise temporal control and spatial resolution remains severely limited due to the pleiotropic effects of most existing pharmacological probes and the lack of suitable genetic tools. To overcome these limitations and enable targeting specific cell types within neurocircuits, we propose an integrated, multidisciplinary approach-spanning synthetic chemistry to neurobiology- combining innovative, and scalable 'chemical optoepigenetic' technologies together with epigenome and transcriptome analysis in human and mouse neurons. Our strategy for neuromodulation exploits photoswitchable compounds with fast thermal relaxation kinetics that possess slow-binding kinetics with their epigenetic targets. Using the family of class I histone deacetylase (HDAC)-containing chromatin-modifying complexes, which our work has demonstrated as key regulators of chromatin-mediated neuroplasticity, to advance the testing of this methodology, the specific aims of the proposed project are to: 1) synthesize, characterize and optimize the physical properties, biochemical potency and selectivity of optoepigenetic probes capable of light-dependent inhibition of the deacetylase activity of neuronal chromatin-modifying complexes containing different class I HDAC isoforms; 2) determine the epigenome and transcriptome changes in cultured human stem cell-derived neurons after precise temporal manipulation of different HDAC complexes; and 3) use the novel optoepigenetic probes to temporally manipulate the epigenome of spatially defined mouse neurons to enhance synaptogenesis and modulate hippocampal circuit function. Overall, by providing significant improvements in spatiotemporal control of HDAC activity in combination with advances in the generation of isoform and complex-selective HDAC inhibitors, we anticipate our approach will limit the pleiotropic effects of currently available small molecule tools. Through selective manipulation of the epigenome in specific regions of neurocircuits, we anticipate being able to significantly improve our understanding of how specific temporal regulation of epigenetic states affects neuroplasticity and to be able to delineate the contribution of epigenetic mechanisms in defined neuronal subtypes within neurocircuits. Importantly, our approach developing chemical optoepigenetic probes is broadly applicable to manipulating epigenetic regulatory mechanisms and could be scaled to enable the assembly of a molecular tool kit for combinatorial optoepigenetic studies. Such tools could have wide applicability in the field of neuroepigenetics and help advance efforts to develop improved therapeutics targeting neuroplasticity.