Progenitor Regulation Underlying Cortical Interneuron Specification

皮层中间神经元规范下的祖细胞调控 抑制性GABA能中间神经元调节复杂的皮质环路活动。大多数皮质中间神经元 起源于表达转录因子Nkx2.1的内侧神经节隆起(MGE)前体细胞 MGE背侧的腹侧MGE和NKX6.2。而兴奋性主神经元的产生 广泛研究，了解中间神经元的发生，特别是MGE的行为和调节 祖先，仍然非常有限。拟议的项目解决了这一悬而未决的知识差距。 尽管成熟的皮质中间神经元是高度多样化的，但最初的决定是采用神经胶质细胞还是 表达神经元命运的生长抑素(SOM+)或小白蛋白(PV+)在MGE中作为祖细胞。 基于来自史和罗斯实验室的强有力的初步数据，我们假设一个紧密的空间和 对MGE祖细胞行为和基因表达特性的时间调控对于 中间神经元的产生流向大脑皮层。该项目包括两个主要目标： 目的1探讨祖细胞极性决定子、分区缺陷之间的功能相互作用 3(PARD3)和G1期活性细胞周期蛋白Cyclins D1(CD1)与D2(CD2)在调节MGE细胞中的作用 除法，检验2个假设：1。MGE前驱分裂模式和动力学在空间和时间上 调节以产生适当的PV+和SOM+中间神经元数量。逆转录病毒和MADM技术 计算模型将用于确定MGE的空间(背侧与腹侧)和时间动力学 放射状胶质前体细胞(RGP)和中间前体细胞(IPC)的分裂模式和中间神经元输出。1B. PARD3和CD1与CD2的差异相互作用协调MGE前体分裂模式和 动态调节适当的PV+与SOM+命运。研究追踪PARD3和PARD3之间的不同相互作用 CD2与CD1在控制MGE RGPS和IPC的分裂模式、动力学和皮质神经元间输出方面的比较。 目的2探索调控MGE RGPS时空行为的核心分子程序 和IPC使用基于10倍基因组学的野生型、CD2-/-、CD1-/-、PARD3 CKO和 Pard3cKO；CD2-/-双突变体。假设：MGE的输出取决于定义CD1和CD1的基因表达 CD2依赖的祖细胞库及其分裂模式和动力学。表达网络将通过以下方式进行验证 组织和分子操作，包括关键驱动因素候选的CRISPR/Cas9基因组编辑 其表达在CD1或CD2和PARD3突变型MGE中发生改变。 该项目将解决关于皮质间神经元发生和 为MGE前体的空间和时间调节提供了一个基本框架，阐明了它们的 分裂模式和动力学，它们的中间神经元输出，以及协调MGE的基本核心程序 祖细胞分裂调节和中间神经元规范。

PROGENITOR REGULATION UNDERLYING CORTICAL INTERNEURON SPECIFICATION Inhibitory GABAergic interneurons modulate complex cortical circuit operation. Most cortical interneurons originate from medial ganglionic eminence (MGE) progenitors that express the transcription factors, NKX2.1 in the ventral MGE and NKX6.2 in the dorsal MGE. While excitatory principal neuron production has been extensively studied, understanding of interneuron genesis, particularly the behavior and regulation of MGE progenitors, remains very limited. The proposed project addresses this outstanding knowledge gap. Although mature cortical interneurons are highly diverse, primary decisions to adopt a glial or somatostatin (SOM+) or parvalbumin (PV+) expressing neuronal fate are made as progenitors in the MGE. Based on strong preliminary data from the Shi and Ross laboratories, we hypothesize that a tight spatial and temporal regulation of the behavior and gene expression properties of MGE progenitors is essential for proper production of interneurons destined for the cortex. The project encompasses two major goals: Aim 1 probes the functional interactions between progenitor cell polarity determinant, partition defective 3 (PARD3), and G1-phase active cell cycle proteins, cyclins D1 (cD1) vs. D2 (cD2), in regulating MGE cell division, testing 2 hypotheses: 1a. MGE progenitor division modes and dynamics are spatially and temporally regulated to generate proper PV+ and SOM+ interneuron numbers. Retroviral and MADM technologies with computational modeling will be used to determine spatial (dorsal vs. ventral) and temporal dynamics of MGE radial glial progenitor (RGP) and intermediate progenitor cell (IPC) division modes and interneuron output. 1b. Differential interactions between PARD3 and cD1 vs. cD2 coordinate MGE progenitor division mode and dynamics to regulate proper PV+ vs. SOM+ fates. Studies pursue differential interactions between PARD3 and cD2 vs. cD1 in governing division mode, dynamics, and cortical interneuron output of MGE RGPs and IPCs. Aim 2 explores the core molecular program regulating the spatial and temporal behavior of MGE RGPs and IPCs using 10X Genomics-based single cell RNA sequencing of wildtype, cD2–/–, cD1–/–, Pard3 cKO and Pard3cKO;cD2–/– double mutants. Hypothesis: MGE output depends on gene expression defining cD1 and cD2 dependent progenitor pools, their division mode and dynamics. Expression networks will be validated by histological and molecular manipulation, including CRISPR/Cas9 genome editing of candidates for key drivers of MGE progenitor specification and whose expression is altered in cD1 or cD2 and Pard3 mutant MGE. The project will address a substantial knowledge gap regarding cortical interneuron genesis and provides a fundamental framework for the spatial and temporal regulation of MGE progenitors, elucidating their division mode and dynamics, their interneuron output, and the underlying core program coordinating MGE progenitor division regulation and interneuron specification.