Subplate-dependent mechanisms of cortical circuit assembly

项目摘要/摘要 大脑皮层的功能需要在发育过程中正确组装电路连接性。子板 位于灰质和白质交界处的神经元在组装大脑皮层方面起着至关重要的作用。 回路，特别是丘脑皮质轴突的引导和感觉图的形成。亚板神经元有 也牵涉到老茧体部的形成。三种非细胞自主机制通过哪个子板 神经元支持轴突的寻路：1)先锋，其中亚板神经元延伸第一个 轴突和为皮质轴突铺设的路径；2)共同分束，其中亚板紧密地下降轴突 与上升轴突相互作用(“握手”)，引导相互连接；3)细胞外基质，其中 亚板下神经元为新生白质中轴突的生长提供了底物。亚极板消融研究 显示下丘脑神经元是不可或缺的。然而，亚板下神经元介导的遗传基础 轴突的引导在很大程度上是未知的，部分原因是胚胎时缺乏接触亚板下神经元的遗传途径。 电路发展的各个阶段。在最近的一项研究中(Doyle等人，PNAS，2021年)，我们报告了一种从基因上 目标是下丘脑的神经元。使用这种方法来询问亚板下神经元基因的必要性和充分性。 在介导的电路组装中，我们发现染色质重构体ARID1A是连接所必需的 下丘脑神经元的功能。人类ARID1A突变是Coffin-Siris综合征的原因之一，一种发育性 以胼胝体发育不全为特征的疾病。我们发现皮质ARID1a基因缺失会导致膝盖骨发育不全和 丘脑皮质轴突错位使人联想到亚板消融。这些错误的接线表型与 下丘脑神经元转录特性的中断，以及下丘脑神经元连接功能的缺陷， 包括下丘脑-丘脑皮质轴突“握手”和细胞外基质。因此，在ARID1A中，我们确定了一个 亚板神经元依赖性轴突引导功能的多功能调节器--一项开启的关键发现 下丘脑神经元的分子和机制研究之门。在使用相同基因的初步研究中 策略，我们进一步确定了转录因子中亚板介导的电路连接的另一个调控因子 索克斯5号。人类SOX5基因突变导致Lamb-Shaffer综合征，这是一种神经发育障碍 通过智力残疾。在这里，我们将利用ARID1A和SOX5提供的特殊机会 研究下丘脑的神经元。我们将利用我们在分子遗传学、电路神经生物学、基因组学和 染色质生物学，以确定ARID1A和Sox5的转录和基因组靶点(目标1和2)和 了解亚板下神经元依赖轴突引导的候选基因的功能(目标3)。一把钥匙 我们的工作表明，亚板神经元的缺陷可能是对神经的贡献被低估了 神经发育障碍中的电路错误，包括与染色质失调有关的疾病。 因此，成功完成拟议的下丘脑神经元研究将阐明基本机制 以及在发育性大脑疾病中下丘脑功能障碍的潜在后果。

PROJECT SUMMARY/ABSTRACT Cerebral cortex function requires correct assembly of circuit connectivities during development. Subplate neurons, strategically positioned at the gray and white matter interface, play essential roles in assembling cortical circuits, notably in guidance of thalamocortical axons and formation of sensory maps. Subplate neurons have also been implicated in corpus callosum formation. Three non-cell autonomous mechanisms by which subplate neurons support axon pathfinding have been proposed: 1) pioneering, wherein subplate neurons extend the first axons and lay a path for cortical axons to follow; 2) co-fasciculation, wherein subplate descending axons closely interact (“handshake”) with ascending axons to guide reciprocal connectivity; 3) extracellular matrix, wherein subplate neurons provide a substrate for axon growth in nascent white matter. Subplate ablation studies have shown subplate neurons to be indispensable. However, the genetic underpinnings of subplate neuron-mediated axon guidance are largely unknown, in part due to a lack of genetic access to subplate neurons at embryonic stages of circuit development. In a recent study (Doyle et al., PNAS, 2021), we reported a strategy to genetically target subplate neurons. Using this approach to interrogate gene necessity and sufficiency in subplate neuron- mediated circuit assembly, we discovered that the chromatin remodeler Arid1a is essential for the wiring functions of subplate neurons. Human ARID1A mutations are a cause of Coffin-Siris syndrome, a developmental disorder characterized by callosal agenesis. We found that cortical Arid1a deletion led to callosal agenesis and thalamocortical axon misrouting reminiscent of subplate ablation. These miswiring phenotypes coincided with disruptions in the transcriptional identity of subplate neurons, and deficits in subplate neuron wiring functions, including subplate-thalamocortical axon “handshake” and extracellular matrix. Thus, in Arid1a, we identified a multifunctional regulator of subplate neuron-dependent axon guidance functions – a key discovery that opens doors to molecular and mechanistic studies on subplate neurons. In preliminary studies using the same genetic strategy, we further identified an additional regulator of subplate-mediated circuit wiring in the transcription factor Sox5. Mutations in human SOX5 cause Lamb-Shaffer syndrome, a neurodevelopmental disorder characterized by intellectual disability. Here, we will leverage the exceptional opportunities that Arid1a and Sox5 provide to study subplate neurons. We will use our expertise in molecular genetics, circuit neurobiology, genomics, and chromatin biology, to identify the transcriptomic and genomic targets of Arid1a and Sox5 (Aims 1 and 2) and gain a functional understanding of candidate genes in subplate neuron-dependent axon guidance (Aim 3). A key implication of our work is that deficits in subplate neurons may be an underappreciated contributor to neural circuit miswiring in neurodevelopmental disorders, including those associated with chromatin dysregulation. Successful completion of the proposed study on subplate neurons will thus illuminate fundamental mechanisms of circuit development, and the potential consequences of subplate dysfunction in developmental brain disorders.