Spatio-temporal control of FOXO3

描述(申请人提供)：在线虫中的研究定义了一种信号通路，控制寿命和对压力的反应。胰岛素生长因子受体(Daf-2)或PI3‘K(AGE-1)功能等位基因的缺失会导致寿命延长2-3倍，并对紫外线照射、氧化损伤和高温等应激因素具有强大的抵抗力。所有这些有益的作用都是由转录因子Daf-16介导的。虽然Daf-16被广泛表达，但其长寿效应是由特定组织中的Daf-16活性介导的。这表明Daf-16参与了组织之间的对话，确保了对环境变化和代谢功能的协调反应。哺乳动物有四个Daf-16同源基因，FOX03是系统发育上最接近的--基于序列和功能。在多种神经退行性变模型中，促进FOXO_3的核定位具有很强的神经保护作用。了解FOXO_3在体内的作用需要对其核定位进行时空控制。在这 工具开发计划我们将制造FOX03与雌激素受体的配体结合区域融合的转基因小鼠(一种对他莫昔芬敏感的版本，称为“ERT”)。当插入到CAGGS-FLOXED-STOP载体中时，我们将能够以细胞类型特异性的方式表达FOXO_3，并通过给药他莫昔芬来驱动它。在这项建议中，我们将对这些小鼠的转基因泄漏、组织特异性表达、非靶标效应和他莫昔芬诱导FOX03靶基因表达的特性进行表征。这一提议的成果是一种多功能的小鼠，它允许时空控制FOX03基因的转录。 公共卫生相关性：在模型生物和培养皿中进行的观察表明，一种名为FOX03的特定蛋白质具有深刻的保护作用，可以抵御导致肌萎缩侧索硬化症(ALS)的侮辱，ALS也被称为Lou Gehrig病。在这个“工具开发”的提议中，我将制造一只小鼠，它将使我们能够研究FOXO_3在小鼠疾病模型中的时空作用。

DESCRIPTION (provided by applicant): Work in C. elegans has defined a signaling pathway that controls longevity and response to stress. Loss of function alleles of the insulin-growth factor receptor (Daf-2) or PI3'K (age-1) lead to 2-3x longer life and powerful resistance to stressors such as UV irradiation, oxidative insult and heat. All of these beneficial actions are mediated by the transcription factor, Daf-16. While Daf-16 is widely expressed, the pro-longevity effects are mediated by Daf-16 activity in specific tissues. This indicates that Daf-16 participate in a conversation among tissues that ensures coordinated responses to environmental changes and metabolic function. Mammals have four Daf-16 orthologs and FOXO3 is the phylogenetically the closest - based on sequence and function. Promotion of nuclear localization of FOXO3 is strongly neuroprotective in multiple models of neurodegeneration. Understanding the role of FOXO3 in vivo will require spatio-temporal control of its nuclear localization. In this tool-development proposal we will make transgenic mice in which FOXO3 is fused to the ligand binding domain of the estrogen receptor (a tamoxifen-sensitive version called "ERT"). When inserted into the CAGGS-floxed-STOP vector we will be able to express FOXO3 in a cell-type specific manner and drive it nuclear by administration of tamoxifen. In this proposal we will characterize these mice for transgene leakiness, tissue specific expression, off target effects and tamoxifen inducibility of FOXO3 target gene expression. The deliverable of this proposal is a versatile mouse that permits spatio-temporal control of FOXO3 gene transcription. PUBLIC HEALTH RELEVANCE: Observations made in model organisms and petri dishes indicate that a specific protein called FOXO3 has profound protective properties against insults that cause Amyotrophic Lateral Sclerosis (ALS), also known as Lou Gehrig's Disease. In this "tool- development" proposal I will make a mouse that will allow us to study the spatio- temporal role of FOXO3 in mouse models of disease.