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不同方面发挥着关键作用，包括神经退行性和神经精神障碍。这种作用包括从神经发生到突触形成的基本细胞过程，对于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很大的进步，但由于大多数现有药物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操纵转录程序的能力仍然受到严重限制。为了克服这些限制，并能够针对神经电路中的特定细胞类型，我们提出了一种集成的多学科方法-跨越合成化学到神经生物学-结合创新和可扩展的化学光表观遗传学技术，以及对人类和小鼠神经元的表观基因组和转录组分析。我们的神经调节策略利用了具有快速热松弛动力学的光开关化合物，这些化合物与其表观遗传靶标具有缓慢的结合动力学。利用我们的工作证明的组蛋白脱乙酰酶(HDAC)类染色质修饰复合体家族作为染色质介导的神经可塑性的关键调节因子来推进这一方法学的测试，本项目的具体目标是：1)合成、表征和优化能够光依赖地抑制含有不同I类HDAC亚型的神经元染色质修饰复合体的脱乙酰酶活性的光表观遗传探针的物理性质、生化效力和选择性；2)测定不同HDAC复合体在时间上的精确操作后培养的人干细胞来源的神经元的表观基因组和转录组的变化；3)利用新型的光-表观遗传探针对空间定位的小鼠神经元的表观基因组进行时间操作，以促进突触发生和调节海马区的回路功能。总体而言，通过在HDAC活性的时空控制方面提供显著的改进，以及在生成异构型和络合物选择性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。通过选择性地操纵神经回路特定区域的表观基因组，我们有望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中特定神经元亚型的表观遗传机制的贡献。重要的是，我们开发化学光表观遗传学探针的方法广泛适用于操纵表观遗传调控机制，并可以扩大规模，以组装用于组合光表观遗传学研究的分子工具包。这些工具可能在神经表观遗传学领域具有广泛的适用性，并有助于推动针对神经可塑性的改进疗法的开发工作。

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.