The Role of TSC Genes During Brain Development

描述（由申请人提供）：本提案的目标是：1)确定TSC基因用于控制神经祖细胞产生神经元和神经胶质的机制，2)确定这些过程中的缺陷如何导致结节性硬化症（TSC）的皮质畸形。TSC是由TSC1或TSC2基因突变引起的，是癫痫和自闭症最常见的遗传原因。这些特征很可能是由于几乎所有患者都存在的脑皮质畸形（结节）。先前的研究表明，在具有异常胶质细胞、畸形神经元和表达神经元和胶质标记物的“巨大”细胞的块状细胞中存在严重的层状破坏。这些发现表明，TSC基因在特定神经元群体的产生以及神经祖细胞从神经元到胶质细胞的转换过程中起着关键作用。通常控制这一过程的机制尚不清楚，但似乎与细胞周期退出和G1的长度有关。细胞周期蛋白、细胞周期蛋白依赖性激酶（cdk）和cdk抑制剂如p27kip1 （p27）介导细胞周期增殖、细胞周期总长度和G1持续时间。值得注意的是，虽然Tsc1或tsc2缺陷成纤维细胞的p27水平和活性降低，但Tsc1或tsc2缺陷神经祖细胞的类似改变尚未报道。我们的具体目标是：1)确定Tsd基因在神经祖细胞产生神经元和胶质细胞的时间上的作用，2)确定7sc1缺陷神经祖细胞的G1持续时间和增殖，以及3)确定p27在7sc7缺陷神经祖细胞中的表达、亚细胞定位和功能。我们将通过体内和体外研究7sc1缺陷的神经祖细胞来实现这些目标。这些神经祖细胞分化的能力将使用谱系特异性标记来确定。此外，我们将使用S期示踪剂来测量细胞周期长度、G1持续时间以及tsc1缺失神经祖细胞小鼠中活跃增殖的细胞比例。最后，将确定p27的表达、亚细胞定位和功能。候选人将利用K08奖，通过与他的导师、范德比尔特大学神经科学研究界的互动，以及与国内外发展神经生物学领域的领导者的积极参与，获得发展神经生物学方面的专业知识。总的来说，这个奖项应该使他成为一名独立的内科科学家，他将成功地竞争未来的NIH校外资助。相关性：结节性硬化症（TSC）是一种遗传性疾病，其表现包括癫痫、脑肿瘤、自闭症和发育迟缓。本研究旨在了解异常神经祖细胞在TSC中的作用。这些发现可能对患有TSC的个体以及患有癫痫和自闭症的非TSC患者具有广泛的治疗意义。

DESCRIPTION (PROVIDED BY APPLICANT): The goals of this proposal are to 1) define mechanisms used by the TSC genes to control the generation of neurons and then glia from neural progenitor cells and 2) determine how defects in these processes result in cortical malformations in Tuberous Sclerosis Complex (TSC). TSC is caused by mutation of either the TSC1 or TSC2 genes and is the most common genetic cause of epilepsy and autism. These features are very likely due to cortical brain malformations (tubers) that are found in almost all patients. Previous work has demonstrated severe laminar disruptions within tubers with abnormal glia, dysmorphic neurons, and "giant" cells expressing neuronal as well as glia markers. These findings suggest that the TSC genes play a critical role during the generation of specific neuronal populations as well as the switch from neuronal to glia production by neural progenitor cells. Mechanisms that normally control this process are not well understood but appear linked to cell cycle exit and the length of G1. Proliferation, total cell cycle length, and G1 duration are mediated by cyclins, cyclin-dependent kinases (cdk), and cdk inhibitors such as p27kip1 (p27). Notably, while Tsc1 or Tsc2-deficient fibroblasts have decreased levels and activity of p27, similar alterations in Tsc1 or Tsc2-deficient neural progenitor cells have not been reported. Our Specific Aims are: 1) Determine the role of the Tsd gene on the timing of neuronal and glia production from neural progenitor cells, 2) determine G1 duration and proliferation in 7sc1-deficient neural progenitor cells, and 3) determine p27 expression, subcellular localization, and function in 7sc7-deficient neural progenitor cells. We will achieve these aims by studying 7sc1-deficient neural progenitor cells both in vivo and in vitro. The ability of these neural progenitor cells to differentiate will be determined using lineage specific markers. In addition, we will use S phase tracers to measure cell cycle length, G1 duration, and the proportion of cells that are actively proliferating in mice with Tsc1-deficient neural progenitor cells. Finally, p27 expression, subcellular localization, and function will be determined. The candidate will utilize this K08 Award to gain expertise in developmental neurobiology though interactions with his mentor, the Neuroscience research community at Vanderbilt University, and active involvement with national and international leaders in Developmental Neurobiology. Overall, this award should position him to become an independent physician-scientist who will successfully compete for future extramural NIH funding. Relevance: Tuberous Sclerosis Complex (TSC) is a genetic disease whose manifestations include seizure disorders, brain tumors, autism and developmental delay. This proposal seeks to understand the role of abnormal neural progenitor cells to TSC. These findings will likely have broad therapeutic implications for individuals with TSC as well as non-TSC patients with seizure disorders and autism.