Long noncoding RNA regulation of neural stem cells

神经干细胞的长非编码RNA调控

• 批准号: 9105277
• 负责人: DANIEL A LIM
• 金额: $ 34.67万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-02-15 至 2021-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9105277
• 关键词: Address; Adult; Alternative Splicing; Alzheimer' s Disease; Bioinformatics; Biological; Biology; Brain; Brain Neoplasms; Cell Lineage; Cells; Code; Collaborations; Complex; Data; Development; Developmental Delay Disorders; Embryo; Genetic Models; Goals; Human; Human Genome; Immunoprecipitation; In Vitro; Knockout Mice; Knowledge; Malignant Neoplasms; Mass Spectrum Analysis; Mental disorders; Messenger RNA; Molecular; Mus; Names; Neurodegenerative Disorders; Neuronal Differentiation; Neurons; Nuclear; Nucleotides; Outcome; Phenocopy; Phenotype; Population; Process; Production; Protein Microchips; Proteins; RNA; RNA Processing; RNA Splicing; RNA-Protein Interaction; Regulation; Ribonucleoproteins; Role; Schizophrenia; Testing; Transcript; Transgenic Organisms; Untranslated RNA; Ventricular; Western Blotting; Work; base; daughter cell; genetic approach; genome-wide analysis; human disease; in vivo; insight; knock-down; mammalian genome; nerve stem cell; nervous system disorder; neurodevelopment; neurogenesis; novel; postnatal; progenitor; programs; public health relevance; relating to nervous system; stem cell population; subventricular zone; therapeutic target; therapy development; transcriptome

 DESCRIPTION (provided by applicant): Long noncoding RNAs (lncRNAs) - transcripts longer than 200 nucleotides with little evidence of protein coding potential - have been implicated in a wide range of human neurological disorders including cancer, developmental delay, schizophrenia and Alzheimer's disease. While the mammalian genome has been discovered to transcribe many thousands of lncRNAs, very few lncRNAs have been characterized in terms of in vivo function and molecular mechanism. In a genome-wide analysis of lncRNAs in adult ventricular- subventricular zone (V-SVZ) neurogenesis, we identified a novel lncRNA transcript named Pinky (Pnky). We have recently demonstrated that Pnky regulates the production of neurons from NSCs of the embryonic and postnatal brain. Pnky is a neural-specific, nuclear lncRNA transcript. In the V-SVZ neurogenic lineage, Pnky is expressed in NSCs and becomes downregulated during neuronal differentiation. In postnatal V-SVZ NSCs, Pnky knockdown potentiates neuronal lineage commitment and expands the transit-amplifying cell population, increasing neuron production several-fold. Pnky is evolutionarily conserved and expressed in NSCs of the developing human brain. In the embryonic mouse cortex, Pnky knockdown increases neuronal differentiation and depletes the NSC population. Mass spectrometry, Western blot, and RNA immunoprecipitation analysis indicates that Pnky physically interacts with PTBP1, a known regulator of neurogenesis, brain tumors, direct cell reprogramming, and RNA splicing. In NSCs, Pnky and PTBP1 regulate the expression and alternative splicing of a core set of transcripts that relates to the cellular phenotype. We have since generated a Pnky conditional knockout (Pnky-cKO) mouse, and this genetic model of Pnky-deficiency phenocopied Pnky knockdown both in vitro and in vivo. The overall goal of the proposed work is to understand the in vivo function and mechanism of Pnky. Aim 1 is to determine the role of Pnky in adult V-SVZ neurogenesis by studying Pnky-deficiency and Pnky transgenic expression in vivo. Preliminary Data, our expertise in V-SVZ biology, and the use of multiple, complementary approaches for manipulating Pnky expression support the feasibility of Aim 1. Aim 2 is to determine the mechanism(s) by which Pnky regulates neurogenesis. Whether Pnky and PTBP1 functionally interact will be investigated with the analysis of biological phenotypes, transcriptome changes, and RNA-protein interactions. The discovery of additional factors that interact with Pnky will provide the basis for investigating other potential lncRNA mechanisms. In addition to Preliminary Data, collaborations with Dr. Aaron Diaz (bioinformatics), Dr. Nevan Krogan (mass spectrometry), Dr. Seth Blackshaw (protein microarrays), and Dr. Hiten Madhani (RNA splicing, RNA-protein interactions) support the feasibility of Aim 2. Such knowledge of lncRNA developmental and mechanistic function will provide critical insight into how lncRNAs can underlie neurological disease and may inform the development of lncRNAs as therapeutic targets.

 描述（由适用提供）：长期未编码的RNA（LNCRNA） - 在200种蛋白质编码潜力的证据的成绩单超过200个核动脉底，在广泛的人类神经系统疾病中暗示了包括癌症，发育迟缓，精神分裂症和阿尔茨海默氏病。尽管已经发现哺乳动物的基因组可以抄录数千个LNCRNA，但很少有LNCRNA以体内功能和分子机制来表征。在成人心室 - 脑室区域（V-SVZ）神经发生中LNCRNA的全基因组分析中，我们确定了一个名为Pinky（PNKY）的新型LNCRNA转录本。我们最近证明，PNKY调节来自胚胎和产后大脑NSC的神经元的产生。 PNKY是一种神经特异性的核LNCRNA转录本。在V-SVZ神经源谱系中，PNKY在NSC中表达，并在神经元分化过程中被下调。在产后V-SVZ NSC中，PNKY敲低潜在的神经谱系承诺并扩大了扩大过境的细胞群体，从而增加了神经元的产生多数倍。 PNKY在进化上是保守的，并在发展中大脑的NSC中表达。在胚胎小鼠皮质中，PNKY敲低增加了神经元的分化并耗尽了NSC种群。质谱，蛋白质印迹和RNA免疫沉淀分析表明，PNKY与PTBP1物理相互作用，PTBP1是神经发生，脑肿瘤，直接细胞重编程和RNA剪接的已知调节剂。在NSC中，PNKY和PTBP1调节与细胞表型有关的核心转录本的表达和替代剪接。从那以后，我们产生了PNKY条件敲除（PNKY-CKO）小鼠，并且这种PNKY缺乏态性苯并pnky pnky的遗传模型在体外和体内均可敲除。拟议工作的总体目标是了解PNKY的体内功能和机制。目的1是通过研究体内的PNKY缺乏症和PNKY转基因表达来确定PNKY在成人V-SVZ神经发生中的作用。初步数据，我们在V-SVZ生物学方面的专业知识以及使用多种完全的操纵PNKY表达的方法支持AIM 1的可行性。AIM2是确定PNKY调节神经发生的机制。 PNKY和PTBP1在功能上相互作用是否将通过分析生物学表型，转录组变化和RNA-蛋白质相互作用进行研究。与PNKY相互作用的其他因素的发现将为研究其他潜在的LNCRNA机制提供基础。除了初步数据，还与Aaron Diaz博士（生物信息学），Nevan Krogan博士（质谱法），Seth Blackshaw博士，Seth Blackshaw博士（蛋白质微阵列）和Hiten Madhani博士和Hiten Madhani博士（RNA拼接，RNA，RNA，RNA，RNA - 蛋白质相互作用）支持批判性的目标2。神经疾病的基础，可以告知LNCRNA作为治疗靶点的发展。