Novel Functions of OLIG2 in Regulating Human Interneuron Production in Health and Disease

兴奋性/抑制性平衡失调(E/I失衡)是认知功能障碍的潜在原因之一 唐氏综合征(DS)，一种以人类21号染色体三倍体为特征的神经发育疾病 (HSA21)。DS的E/I失衡在很大程度上是由于GABA能中间神经元的过度产生所致。至 研究DS的智能障碍机制，必须更好地了解中间神经元是如何产生的 神经前体细胞(NPC)在发育过程中受到调控。OLIG基因，包括OLIG1和OLIG2， 被映射到HSA21上，并在DS中三重。对小鼠模型的研究表明，在胚胎期间 在发育过程中，寡核苷酸1和2都在神经节隆起中大量表达，这是一种位于脑内的结构 在胚胎端脑的腹侧，大多数皮质中间神经元就是从那里诞生的。此外，奥利格 基因对中间神经元的产生起着关键的调节作用。值得注意的是，OLIG基因的表达在 人类与啮齿动物形成神经节隆起。我们发现，人类的分化诱导了 多能干细胞(HiPSCs)到前脑腹侧的神经前体细胞概括了以前在人脑中的发现。 OLIG2在神经节隆起的一群人的NPC中表达的组织。相比之下， OLIG1在这些NPC中表达很少，并且与OLIG2具有互补的表达模式。向上 到目前为止，人OLIG基因在人GABA能神经元发育中的作用主要是 未知。利用DS患者来源的HiPSCs，我们进一步证实了OLIG2在DS患者中过表达 HiPSC来源的腹侧前脑神经前体细胞。因此，我们假设OLIG基因的异常表达， 尤其是人DS腹侧前脑神经干细胞中的OLIG2，决定了GABA能神经元的过度产生 这会导致E/I失衡，并导致DS的智力残疾。 为了检验这一假设，我提出了三个具体目标。目的1：确定OLIG2在调节中的作用 人的GABA能神经元由正常的HiPSC产生。我们将使用OLIG2基因敲除生成的HiPSC 利用CRISPR/Cas9技术研究OLIG2在人中间神经元发育中的作用。目标2：实现 确定OLIG2是否是DS患者GABA能神经元过度产生的原因基因。通过使用 对照和DS hiPSCs，以及具有归一化OLIG2基因剂量的DS hiPSCs，我们将确定 OLIG2的过度表达是否导致DS中GABA能神经元的过度产生。目标3：通过使用 新的人源化神经元嵌合小鼠模型，我们将进一步考察神经元间的规范 正常和双侧HiPSC来源的腹侧前脑神经前体细胞及其整合和在E/I失衡中的作用 在完整的神经回路中的活体。这项拟议的研究结果将为我们提供对该功能的新见解 OLIG2在调节人GABA能神经元产生中的作用及其对开发潜能的新启示 调控OLIG2基因表达在DS治疗中的应用

Disruption of excitatory/inhibitory balance (E/I imbalance) is one of the underlying causes of cognitive deficit of Down syndrome (DS), a neurodevelopmental disease characterized by triplication of human chromosome 21 (HSA21). This E/I imbalance in DS is largely resulted from overproduction of GABAergic interneurons. To study the mechanisms of intellectual disability of DS, we must better understand how interneuron production from neural progenitor cells (NPCs) is regulated during development. OLIG genes, including OLIG1 and OLIG2, are mapped to HSA21 and triplicated in DS. Studies in mouse models demonstrate that during embryonic development, both Olig1 and 2 are abundantly expressed in the ganglionic eminence, a brain structure located in the ventral embryonic telencephalon and from where most cortical interneurons are born. Moreover, Olig genes critically regulate interneuron production. Notably, expression of OLIG genes is starkly different in the human versus rodent developing ganglionic eminence. We found that differentiation of human induced pluripotent stem cell (hiPSCs) to ventral forebrain NPCs recapitulated the previous findings in human brain tissue that OLIG2 was expressed in a subpopulation of human NPCs in the ganglionic eminence. In contrast, OLIG1 was expressed in very few of these NPCs and had a complimentary expression pattern with OLIG2. Up to now, the functions of human OLIG genes in the development of human GABAergic neuron is largely unknown. Using DS patient-derived hiPSCs, we further identified that OLIG2 was overexpressed in the DS hiPSC-derived ventral forebrain NPCs. Therefore, we hypothesize that abnormal expression of OLIG genes, particularly OLIG2, in human DS ventral forebrain NPCs determines the overproduction of GABAergic neurons from these progenitors, which leads to E/I imbalance and significantly contributes to intellectual disability of DS. To test the hypothesis, I proposed three specific aims. Aim 1: to determine the role of OLIG2 in regulating human GABAergic neuron production from normal hiPSCs. We will employ OLIG2 knockout hiPSCs generated by using CRISPR/Cas9 technology to study the role of OLIG2 in human interneuron development. Aim 2: to determine whether OLIG2 is a causal gene of the overproduction of GABAergic neurons in DS. By using control and DS hiPSCs, as well as the DS hiPSCs with normalized OLIG2 gene dosage, we will determine whether overexpression of OLIG2 causes the overproduction of GABAergic neurons in DS. Aim 3: by using a novel humanized neuronal chimeric mouse model, we will further examine interneuron specification of the normal and DS hiPSC-derived ventral forebrain NPCs and their integration and contribution to E/I imbalance in vivo within intact neural circuits. Findings from this proposed study will provide novel insights into the function of OLIG2 in regulating human GABAergic neuron production and shed new light on developing potential therapeutic applications for DS by regulating the expression of OLIG2 gene.