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A model of neuronal cell development and differentiation: Mouse rod photoreceptor

神经元细胞发育和分化模型：小鼠视杆光感受器

基本信息

批准号：
7845395
负责人：
JUDITH Mosinger OGILVIE
金额：
$ 22.13万
依托单位：
SAINT LOUIS UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-07-10 至 2013-09-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7845395
关键词：
Adolescent Adult Animal Model Apical Apoptosis Blindness Cell Cycle Cell Differentiation process Cells Child Childhood Cilia Defect Degenerative Disorder Development Disease Electron Microscopy Epithelial Cells Eukaryotic Cell Failure Family Genes Goals Health Immunohistochemistry Inherited Investigation Label Lead Maintenance Membrane Microarray Analysis Modeling Molecular Monomeric GTP-Binding Proteins Mus Neonatal Neurons Organ Culture Techniques Pathway interactions Pattern Photoreceptors Proteins RNA RNA Interference Regulation Research Retina Retinal Retinitis Pigmentosa Role Scientist Sensory Signal Pathway Source Synapses Techniques Testing Therapeutic Intervention Time Vertebrate Photoreceptors Vision Visual Western Blotting Wild Type Mouse Work base cell type developmental neurobiology early onset gain of function improved in vivo innovation insight interest loss of function mouse model mutant neuron development overexpression phosphoric diester hydrolase photoreceptor degeneration polarized cell postnatal precursor cell programs public health relevance rab GTP-Binding Proteins research study retinal rods synaptogenesis trafficking vector

项目摘要

DESCRIPTION (provided by applicant): Photoreceptors are highly specialized ciliated epithelial cells that are easily accessible and undergo cell differentiation postnatally. The long-term goal of our research program is to understand the molecular signaling pathways underlying vertebrate photoreceptor development and cell differentiation, with particular focus on how disruption of these pathways leads to early onset vision loss. The objective of this proposal is to determine the role of the protein PRA1 in rod photoreceptor cell differentiation. During development, precursor photoreceptor cells exit the cell cycle and undergo a transformation to produce highly polarized cells with unique sensory specializations. Although morphological differentiation of photoreceptors is well characterized, less is known about the underlying molecular mechanisms. The rd1 mouse retina, characterized by early-onset, rapid degeneration of rod photoreceptors, was the first identified animal model of retinitis pigmentosa. Interestingly, rd1 rods show developmental defects, including failure to undergo normal synaptogenesis and outer segment differentiation, significantly before degenerative changes appear, presenting a model for investigations of photoreceptor development as well as a congenital neuronal disease. In preliminary studies, we have performed microarray analysis of wild type mouse retina compared to rd1 mutant retina at P2-8, prior to onset of photoreceptor apoptosis. Only two genes were consistently and significantly downregulated in the rd1 mouse at all time points: PDE6b, the mutant gene, and Prenylated rab acceptor 1 (PRA1). Small GTPases in the Rab family are known to be critical for vesicular trafficking involved in maintenance of both the outer segment and the synapse in the mature photoreceptor. PRA1 regulates delivery of Rab GTPases to membranes in eukaryotic cells, although its precise function and mechanism of action are poorly understood. We hypothesize that PRA1 regulation of vesicular trafficking is necessary and sufficient for synaptic and outer segment differentiation in rod photoreceptors. We propose to test our hypothesis with the following specific aims: (1) Determine the expression pattern of PRA1 in the adult and developing wild type and rd1 mouse retinas using Western blot and immunohistochemistry, and (2) determine whether PRA1 is necessary and sufficient for synaptic and outer segment differentiation in rod photoreceptors using RNA based loss-of-function and gain-of-function techniques. The proposed work is innovative in being the first to identify the significance of PRA1 in an animal model of neuronal development and disease. Upon completion of this work, we expect to have determined whether PRA1 expression can restore normal rod photoreceptor cell differentiation in the rd1 mouse retina. These results are ultimately expected to have a significant impact on the field of developmental neurobiology and to provide insights into mechanisms underlying aberrant differentiation, leading to improvement in childhood visual health. PUBLIC HEALTH RELEVANCE: The proposed experiments will elucidate mechanisms involved in development and differentiation of a highly specialized neuronal epithelial cell type: the rod photoreceptor cell. The rd1 mouse model of juvenile retinitis pigmentosa will be used to gain insights into the source of developmental aberrations. Greater understanding of these mechanisms could ultimately lead to targeted therapeutic interventions for juvenile retinitis pigmentosa and improved childhood visual health.

描述（由申请人提供）：光感受器是高度特化的纤毛上皮细胞，易于接近并在出生后经历细胞分化。我们研究计划的长期目标是了解脊椎动物感光细胞发育和细胞分化的分子信号通路，特别关注这些通路的破坏如何导致早发性视力丧失。本提案的目的是确定蛋白质PRA 1在视杆细胞分化中的作用。在发育过程中，前体感光细胞退出细胞周期并经历转化以产生具有独特感觉特化的高度极化细胞。虽然光感受器的形态分化特征很好，但对潜在的分子机制知之甚少。rd 1小鼠视网膜的特点是早期发病，视杆细胞迅速变性，是第一个确定的视网膜色素变性的动物模型。有趣的是，rd 1杆显示发育缺陷，包括未能进行正常的突触和外节分化，显着的退行性变化出现之前，提出了一个模型的感光细胞发育以及先天性神经元疾病的调查。在初步研究中，我们已经进行了野生型小鼠视网膜的微阵列分析相比，rd 1突变体视网膜在P2-8，感光细胞凋亡的发病之前。在所有时间点，只有两个基因在rd 1小鼠中一致且显著下调：PDE 6 b（突变基因）和Prenylated rab受体1（PRA 1）。已知Rab家族中的小GTP酶对于参与成熟光感受器中的外节和突触的维持的囊泡运输是关键的。PRA 1调节Rab GTP酶向真核细胞膜的递送，尽管其确切功能和作用机制知之甚少。我们推测，PRA 1调节囊泡运输是必要的和足够的突触和外段分化杆光感受器。我们提出用以下具体目标来检验我们的假设：（1）使用Western印迹和免疫组织化学来确定PRA 1在成年和发育中的野生型和rd 1小鼠视网膜中的表达模式，以及（2）使用基于RNA的功能丧失和功能获得技术来确定PRA 1是否是视杆细胞中突触和外节分化所必需和充分的。这项工作的创新之处在于首次确定了PRA 1在神经元发育和疾病动物模型中的重要性。在完成这项工作后，我们希望确定PRA 1表达是否可以恢复rd 1小鼠视网膜中正常的视杆细胞分化。这些结果最终有望对发育神经生物学领域产生重大影响，并为异常分化的机制提供见解，从而改善儿童视觉健康。公共卫生相关性：拟议的实验将阐明一个高度专业化的神经元上皮细胞类型的发展和分化的机制：杆感光细胞。青少年视网膜色素变性的rd 1小鼠模型将用于深入了解发育畸变的来源。对这些机制的更深入了解可能最终导致针对青少年视网膜色素变性的有针对性的治疗干预，并改善儿童的视觉健康。