Global mapping of altered neural circuits in a mouse model of DDX3X mutations

项目摘要 自闭症谱系障碍（Autism Spectrum Disorder，ASD）是一种常见的神经发育障碍， 等位基因，但没有有效的药物治疗。X连锁基因DDX 3X的突变是高风险的 ASD的因素。受影响的个体主要是女性，因此研究DDX 3X可能会提供对性别的见解 大脑发育和功能的差异。DDX 3X编码对mRNA至关重要的DEAD盒RNA解旋酶 新陈代谢. DDX 3X广泛表达，其在大脑中的功能刚刚开始显现： 调节皮质神经发生、后脑发育和突触发生。然而，我们不知道 DDX 3X突变的电路水平决定因素。迫切需要填补这些空白，因为在我们这样做之前， 解释ASD的复杂性和开发有效的治疗方法仍然遥不可及。为了解决这个 未满足的需求，在我们的实验室中产生了具有DDX 3X突变的构建体和表面有效性的小鼠 （Ddx 3x +/-小鼠）。长期目标是了解ASD的细胞和电路生物学，并确定新的 治疗干预的目标。总的目标是捕捉突变的神经机制， ASD风险基因DDX 3X与多模式和整体分析。核心假设是Ddx 3x调节 支持复杂行为的离皮质神经回路的分子特性、连通性和活性。的 基本原理是，一旦我们确定了可靠的神经基质，就可以开发基于机制的治疗方法。 并进行了临床前测试。该假设将通过追求三个具体目标进行测试：1）识别皮质 群体和Ddx 3x +/-小鼠中受影响的分子特征; 2）用 在Ddx 3x +/-小鼠中改变的连接性和/或活性;和，3）解剖和操纵离皮质电路驱动 Ddx 3x +/-小鼠的异常行为。在目标1下，受Ddx 3x影响的主要分子和细胞系综 将使用单细胞转录组学和3D细胞作图来鉴定突变。在目标2下，基于活动 被Ddx 3x突变破坏的神经基质将使用3D映射立即早期- 行为后的基因表达根据目标3，将使用化学遗传学方法操纵电路。的 该提案是创新的，因为它使用尖端方法来绘制定义神经元的3D景观。 群体和通过新的ASD小鼠模型的整个大脑的全脑活动。也是创新 因为DDX 3X是最近发现的ASD的高置信风险基因，其在塑造大脑中的作用 电路仍然完全未知。该应用程序意义重大，因为它将促进我们对 ASD的复杂性，达到全脑，电路级的分辨率，同时推动发展一个强大的 平台，以探索跨模型和发展阶段的趋同。这些结果预计将有 这是一个积极的影响，因为它们将为ASD的新型治疗干预铺平道路。

PROJECT SUMMARY Autism spectrum disorder (ASD) is a prevalent neurodevelopmental disorder with dozens of highly penetrant risk alleles and yet no effective pharmacological treatment. Mutations in the X-linked gene DDX3X are a high-risk factor for ASD. Affected individuals are predominantly females, so studying DDX3X might offer insights into sex differences in brain development and function. DDX3X encodes a DEAD-box RNA helicase critical for mRNA metabolism. DDX3X is broadly expressed, and its functions in the brain are just beginning to emerge: Ddx3x regulates cortical neurogenesis, hindbrain development, and synaptogenesis. However, we do not know the circuit-level determinants of DDX3X mutations. There is a critical need to fill these gaps because, until we do so, deciphering the complexity of ASD and developing effective therapeutics remain out of reach. To address this unmet need, a mouse with construct and face validity for DDX3X mutations was generated in our laboratory (Ddx3x+/- mice). The long-term goal is to understand the cellular and circuitry biology of ASD and identify new targets for therapeutic intervention. The overall objective is to capture the neural mechanisms of mutations in the ASD risk gene DDX3X with multimodal and holistic profiling. The central hypothesis is that Ddx3x regulates the molecular identity, connectivity, and activity of corticofugal circuits subserving complex behaviors. The rationale is that, once we identify reliable neural substrates, mechanism-based therapeutics can be developed and tested pre-clinically. The hypothesis will be tested by pursuing three Specific Aims: 1) Identify the cortical populations and the molecular signatures affected in Ddx3x+/- mice; 2) Map brain-wide neural ensembles with altered connectivity and/or activity in Ddx3x+/- mice; and, 3) Dissect and manipulate corticofugal circuits driving abnormal behavior in Ddx3x+/- mice. Under Aim 1, the major molecular and cellular ensembles affected by Ddx3x mutations will be identified using single-cell transcriptomics and 3D cellular mapping. Under Aim 2, activity-based neural substrates that are disrupted by Ddx3x mutations will be dissected using 3D mapping of immediate-early genes expression after behavior. Under Aim 3, circuits will be manipulated with chemogenetics approaches. The proposal is innovative because it uses cutting-edge methods to map the 3D landscape of defined neuronal populations and whole-brain activity through the entire brain of a novel ASD mouse model. It is also innovative because DDX3X is a high-confidence risk gene for ASD just recently discovered, and its role on shaping brain circuits is still completely unknown. The application is significant because it will advance our understanding of ASD complexity by reaching whole-brain, circuitry-level resolution, while propelling the development of a robust platform to probe convergences across models and developmental stages. These results are expected to have a positive impact because they will pave the way for novel therapeutic interventions for ASD.