Identifying mTOR Dependent Periods During Brain Development

识别大脑发育过程中 mTOR 依赖期

• 批准号: 10240722
• 负责人: KEVIN C ESS
• 金额: $ 45.16万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-01 至 2025-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10240722
• 关键词: Acute; Address; Affect; Aftercare; Age; Archives; Behavioral; Benign; Biological Models; Brain; Brain Neoplasms; Cardiac; Cell Cycle; Cell Line; Cell Lineage; Cell Survival; Cell model; Cell physiology; Cells; Cerebrum; Clinical; Complex; Cortical Malformation; Coupled; Cryopreservation; Custom; Cytometry; Data; Data Set; Development; Disease; Disease Progression; Drug Exposure; Electrophysiology (science); Embryonic Development; Engineering; Epilepsy; FRAP1 gene; Flow Cytometry; Future; Genes; Hamartoma; Human; Image; In Vitro; Individual; Institution; Intellectual functioning disability; Kidney; Kinetics; Lesion; Life; Lung; Mammals; Measurement; Measures; Mediating; Mental disorders; Mutation; Neuroglia; Neurologic Symptoms; Neurons; Organoids; Outcome; Partial Epilepsies; Pathogenesis; Pathway interactions; Patients; Pattern; Pharmaceutical Preparations; Pharmacology; Phenotype; Phosphoproteins; Phosphotransferases; Process; Protein-Serine-Threonine Kinases; Proteins; Protocols documentation; Publishing; RNA; Regulation; Resected; Series; Signal Pathway; Signal Transduction; Symptoms; Systems Biology; TSC1 gene; TSC1/2 gene; TSC2 gene; Testing; Therapeutic Intervention; Time; Tissues; Tuberous sclerosis protein complex; analysis pipeline; autism spectrum disorder; biobank; body system; cell growth; cell type; cellular imaging; cohort; computational pipelines; design; excitatory neuron; induced pluripotent stem cell; inhibitory neuron; mTOR Signaling Pathway; mTORopathies; multiplexed imaging; mutant; mutational status; nerve stem cell; neurodevelopment; postnatal; progenitor; relating to nervous system; response; single-cell RNA sequencing; stem cells; tissue archive; tumor

Tuberous Sclerosis Complex (TSC) is a multi-organ disorder caused by mutations in the TSC1 or TSC2 genes. TSC is a challenging disease to approach as there are many involved organ systems, which have distinct profiles of symptom onset, disease progression, and in some cases even stability or regression of the benign tumors known as hamartomas. These multiple examples of distinct time courses in each organ system strongly suggest that the TSC1/TSC2 genes control cell signaling pathways that are tissue specific and developmentally regulated, resulting in lesions that present at different times in the lifetime of the patient. These pathways certainly include mTOR kinase signaling, but critical upstream and downstream regulators of this developmentally regulated process in specific tissues remain poorly understood. The neurological manifestations of TSC are typically severe at very early ages and include epilepsy, intellectual disability, autism, and behavioral/psychiatric disorders. Recent findings in human cell-derived model systems suggest that neural development is disrupted in TSC, and that proper regulation of mTOR signaling is especially important in human brain in comparison to other mammals. However, the cellular mechanisms connecting TSC1/2 mutation and the phenotypic outcomes of this mutation are not well understood. This project will use patient-derived cells and single-cell measurements of protein and RNA to measure altered signaling pathways in various cell types of the human brain and also address how these abnormalities impact specific developmental stages. Examined stages will span early neural progenitor cells to more mature neurons found in the postnatal brain. Human induced pluripotent stem cells (iPSCs) from patients carrying TSC2 mutations will be used to generate lineage-committed progenitors and differentiated neurons and glia. We will also use freshly resected human tubers as well as previously resected human tubers that have been fixed and stored, and will employ custom-designed computational pipelines to compare the developmental trajectories of TSC2-mutant cells to matched controls and larger published datasets. Using cutting edge cell imaging and analysis protocols, we will test the overarching hypothesis that tubers from patients with TSC and stem cell derivative neural cells and tissues have mTOR-dependent and mTOR- independent signaling abnormalities that are lineage- and temporally-restricted. Finally, we will quantitatively compare signaling dynamics in specific developmental stages and lineages between TSC2 mutant cells and cells derived from a second “mTORopathy” with overlapping but non-identical clinical features, to dissect the function of different components of this pathway in neural development and pathogenesis and reveal compensatory signaling after treatment of cells carrying TSC2 or DEPDC5 mutations.

结节硬化症复合物（TSC）是由TSC1或TSC2突变引起的多器官疾病 基因。 TSC是一种挑战疾病，因为有许多涉及的器官系统，这些系统 具有症状发作，疾病进展的明显特征，在某些情况下甚至稳定或 良性肿瘤的回归被称为Hamartomas。这些不同时间的多个例子 每个器官系统中的课程强烈建议TSC1/TSC2基因控制细胞信号传导 特定组织并受到调节的途径，导致病变 在患者一生中的不同时间出现。这些途径当然包括mtor 激酶信号传导，但关键的上游和下游调节器 特定组织中的调节过程仍然很少理解。神经系统表现 TSC通常很早就很严重，包括癫痫，知地残疾，自闭症， 和行为/精神疾病。人类细胞来源模型系统的最新发现表明 TSC中神经发育受到了破坏，MTOR信号的适当调节是 与其他哺乳动物相比，在人脑中尤其重要。但是，细胞 连接TSC1/2突变和该突变的表型结果的机制不是很好 理解。该项目将使用患者衍生的细胞和蛋白质的单细胞测量和 RNA测量人脑各种细胞类型的信号传导途径的改变，也解决 这些异常如何影响特定的发展阶段。检查的阶段将早早 神经祖细胞到产后大脑中发现的更成熟的神经元。人类诱导 携带TSC2突变患者的多能干细胞（IPSC）将用于产生 谱系的祖细胞以及分化的神经元和神经胶质。我们还将新鲜使用 切除的人类块茎以及以前已切除的人类块茎已被固定和储存， 并将采用定制设计的计算管道来比较发展轨迹 TSC2突变细胞与匹配的控件和较大已发布的数据集。使用尖端细胞 成像和分析方案，我们将测试总体假设，即患者的块茎 TSC和干细胞衍生神经细胞和组织具有mTOR依赖性，并且 谱系和暂时限制的MTOR-独立信号异常。最后，我们 将定量比较特定发育阶段和谱系的信号传导动力学 在TSC2突变细胞和源自第二个“ mtoropathy”的细胞之间，并重叠，但 非相同的临床特征，以剖析不同组成部分的功能 神经发育和发病机理中的途径，并揭示 处理携带TSC2或DEPDC5突变的细胞后的补偿信号传导。