Role of mTORC1 dependent translation in neurological deficits of TSC

mTORC1 依赖性翻译在 TSC 神经功能缺陷中的作用

基本信息

批准号：
10424796
负责人：
Matthew Binder
金额：
$ 6.76万
依托单位：
YALE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-07-01 至 2024-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10424796
关键词：
Address Anatomy Attenuated Axon Behavior Behavioral Binding Proteins Bioinformatics Brain CRISPR/Cas technology Caregivers Clinical Complex Corpus striatum structure Development Disease Electrophysiology (science)Electroporation Epilepsy Etiology Eukaryotic Initiation Factors FRAP1 gene Genes High Prevalence Hyperactivity Hypertrophy Incidence Individual Knowledge Laboratories Lead Medial Microscopy Modeling Molecular Mus Mutation Neurologic Neurologic Deficit Neurons Pathologic Patients Phenotype Population Prefrontal Cortex Property Ras homolog enriched in brain Research Role SDZ RAD Seizures Signal Pathway Signal Transduction Social Behavior Social Interaction Synapses TSC1 gene TSC2 gene Testing Therapeutic Transgenic Model Translations Tuberous Sclerosis Ultrasonics Western Blotting autism spectrum disorder autistic axon growth behavioral phenotyping comorbidity design developmental disease effective therapy hippocampal pyramidal neuron in utero inhibitor insight mouse model novel prevent relating to nervous system small hairpin RNA social synaptic function trait transcriptome sequencing translatome vocalization

项目摘要

ABSTRACT Tuberous sclerosis complex (TSC) is a developmental disorder caused by a mutation in the TSC1 or TSC2 gene. Mutations in TSC genes result in hyperactivation of the mechanistic target of rapamycin complex 1 (mTORC1), a signaling pathway vital for development. Individuals with TSC present with a high incidence of neurological conditions, among the most prevalent of which are epilepsy and autism spectrum disorder (ASD). While mTORC1 inhibitors can effectively attenuate seizures, they do not improve TSC’s autistic comorbidity. Thus, there is a critical need to better understand the etiology of autism traits to identify novel treatments. Our laboratoryhasdevelopedamousemodelofTSCin which constitutively active Rheb is expressed in developing cortical neurons of the medial prefrontal cortex (mPFC), a region that is dysregulated in TSC and implicated in ASD. This model allows for a more precise activation of mTORC1 than transgenic models and is thus ideally suited to parse out complex etiology. Using this model, our lab found alterations in local mPFC connectivity that resemble the pathological changes in TSC’s autistic comorbidity. Moreover, when aberrant cap dependent translation was normalized via the activation of a translational repressor, the observed alterations were prevented. While connectivity has been examined within the mPFC in a hyperactive mTORC1 state, the accompanying behavioral phenotype has not been assessed nor have mTORC1’s effects on specific subcortical projections from the mPFC. To this end, the striatum is a compelling target, as it is heavily interconnected with the mPFC and is integral to socio-communicative behaviors (core deficits in ASD). However, axonal connectivity in TSC is understudied and mPFC-striatal projections have not been assessed. Furthermore, while normalizing translation can mitigate cellular alterations in hyperactive mTORC1 conditions, the specific downstream molecular alterations that account for these changes are unknown, as are their efficacy to attenuate behavioral deficits. Therefore, the specific aims are to: 1). Determine whether hyperactive mTORC1 in mPFC-striatal projections contributes to socio-communicative deficits. 2). Determine whether increasing mTORC1 activity during early mPFC development leads to alterations in cortico-striatal projections resulting in an excitation-inhibition imbalance in the striatum. 3). Determine whether altered cap-dependent translation and downstream molecules contribute to alterations in axonal connectivity and socio-communicative deficits. To address these aims, in utero electroporation (IUE) will be used to increase mTORC1 in pyramidal neurons of the mPFC. Vocalization and social interaction tests will assess the phenotype, whereas lightsheet microscopy, axonal tracing, and electrophysiology will assess mPFC-striatal connectivity. Potential candidate molecules downstream of translation that control axonal growth will be identified via Trap, and shRNA will be designed to assess the effects of normalizing the identified genes on socio-communicative deficits and alterations in mPFC-striatal connectivity.

抽象的结节性硬化症 (TSC) 是一种由 TSC1 或 TSC2 突变引起的发育障碍 TSC 基因突变导致雷帕霉素复合物 1 的机制靶点过度激活。 (mTORC1) 是一种对发育至关重要的信号通路，患有 TSC 的个体的发病率很高。神经系统疾病，其中最常见的是癫痫和自闭症谱系障碍（ASD）。虽然 mTORC1 抑制剂可以有效减轻癫痫发作，但它们不能改善 TSC 的自闭症合并症。因此，迫切需要更好地了解自闭症特征的病因学，以确定新的治疗方法。实验室开发了 TSCin 小鼠模型，其组成型活性 Rheb 在发育中的皮质中表达内侧前额叶皮层 (mPFC) 的神经元，该区域在 TSC 中失调并与自闭症谱系障碍有关。该模型比转基因模型能够更精确地激活 mTORC1，因此非常适合使用这个模型，我们的实验室发现了类似于局部 mPFC 连接的改变。此外，当帽依赖性翻译异常时，TSC 的自闭症合并症的病理变化。通过翻译抑制子的激活而正常化，观察到的改变被阻止。在 mPFC 内，mPFC 内的连接性已在 mTORC1 过度活跃状态下进行了检查，伴随着行为尚未评估表型，也没有评估 mTORC1 对特定皮层下投射的影响为此，纹状体是一个引人注目的目标，因为它与 mPFC 密切相关，并且然而，TSC 的轴突连接是社会交流行为的组成部分（自闭症谱系障碍的核心缺陷）。此外，在标准化翻译时，尚未对其进行充分研究和 mPFC 纹状体预测评估。可以减轻 mTORC1 过度活跃条件下的细胞改变，特定的下游分子导致这些变化的改变以及它们减轻行为缺陷的功效尚不清楚。因此，具体目标是： 1) 确定 mPFC 纹状体投射中 mTORC1 是否过度活跃。 2).确定早期是否增加mTORC1活性 mPFC 的发育导致皮质纹状体投射的改变，从而导致兴奋抑制确定纹状体的不平衡3)。有助于改变轴突连接和社会沟通缺陷以在子宫内实现这些目标。电穿孔 (IUE) 将用于增加 mPFC 锥体神经元中的 mTORC1。社会互动测试将评估表型，而光片显微镜、轴突追踪和电生理学将评估 mPFC-纹状体下游的潜在候选分子。控制轴突生长的翻译将通过 Trap 进行识别，shRNA 将被设计来评估效果使已识别的社会沟通缺陷和 mPFC-纹状体连接改变的基因正常化。