Comparative genomic and molecular genetics approaches to retinal stem cells

视网膜干细胞的比较基因组和分子遗传学方法

• 批准号: 7677879
• 负责人: Li Cai
• 金额: $ 22.51万
• 依托单位: RUTGERS, THE STATE UNIV OF N.J.
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-09-01 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7677879
• 关键词: Binding; Biological Process; Cell Proliferation; Cell physiology; Cells; Cellular Morphology; Chromatin Structure; Complex; Computer Simulation; DNA; DNA Sequence; Databases; Development; Disease; Electroporation; Elements; Embryo; Enhancers; Epigenetic Process; Functional RNA; Gene Expression; Generations; Genes; Genetic Transcription; Genome; Genomics; Goals; Green Fluorescent Proteins; Individual; Injury; Location; Methods; Molecular Genetics; Nerve Degeneration; Neuraxis; Neuroglia; Neurons; Nucleic Acid Regulatory Sequences; Open Reading Frames; Pattern; Population; Proteins; Replacement Therapy; Reporter; Reporter Genes; Resources; Retina; Retinal; Retinal Degeneration; Spinal cord injury; Stem Cell Research; System; Techniques; Testing; Therapeutic; Therapeutic Intervention; Tissues; Trans-Activators; Transcriptional Regulation; Transplant-Related Disorder; base; cell type; combinatorial; comparative; design; genome-wide; in vivo; nerve stem cell; novel; plasmid DNA; public health relevance; retinal progenitor cell; stem cell population

DESCRIPTION (provided by applicant): The long term goal of this proposal is to develop therapeutic interventions in the generation of neurons from neural stem cells (NSC) for replacement therapies of disease and injury in the central nervous system (CNS). Neural stem cells give rise to all of the different cell types in the CNS. Understanding of the complex transcriptional networks that control the process of cell fate determination in NSC is crucial in applications for therapeutic purpose. The generation of diversity of neurons and glial cells is achieved through cell proliferation, cell fate determination and differentiation of embryonic NSC populations into progressively more specialized cell types that make up the CNS. The uniqueness of individual neurons and glial cells is determined by combinatorial patterns of gene expression; and gene expression is largely controlled at the level of DNA sequence (cis-regulatory element, genetic), as well as by chromatin structure (epigenetic). One of the key components in transcription regulation is the enhancer, a non-coding DNA sequence that is often evolutionarily conserved. Upon binding of trans-acting factors, enhancers determine tissue or cell type-specific expression of particular genes. We will choose genes that are crucial to NSC cell fate determination from genome wide gene expression studies and analyze the non-coding DNA regions for their regulatory functions (e.g. as an enhancer) in NSC gene expression during cell fate determination. We will predict (in silico) and functionally characterize (in vivo) the predicted putative enhancers in chick retinal stem cells. We refer to this kind of enhancer as NSC enhancers. Our focus will be on NSC enhancers that are important for the development of the retina. The two specific aims are: 1) to predict evolutionarily conserved NSC enhancers; and 2) to verify and characterize putative NSC enhancers. The successful completion of the proposed study will help to identify transcriptional control networks that are crucial for cell fate determination of neural stem cells. In addition, novel retina-specific enhancers identified from this study can be used to identify novel protein factors that are previously unknown for their function in controlling neural cell fate determination. Our findings will ultimately provide an integrated transcription network that controls NSC cell fate determination in the retina (can also be applied to other developmental systems in general). Such an understanding of the transcriptional networks is fundamental to the development of potential treatments or therapeutic transplants for diseases ranging from retinal degeneration to spinal cord injury. PUBLIC HEALTH RELEVANCE: The goal of this proposal is to search for DNA sequence elements that exist in the non-protein coding regions of the genome that regulate the generation of nerve cells from neural stem cells. The successful completion of the proposed studies will help the development of potential treatments or therapeutic transplants for diseases ranging from neural degeneration to spinal cord injury.

描述(申请人提供)：这项建议的长期目标是开发从神经干细胞(NSC)生成神经元的治疗干预措施，用于中枢神经系统(CNS)疾病和损伤的替代疗法。神经干细胞在中枢神经系统中产生了所有不同类型的细胞。了解神经干细胞中控制细胞命运决定过程的复杂转录网络在治疗应用中是至关重要的。神经元和神经胶质细胞多样性的产生是通过细胞增殖、细胞命运决定和胚胎神经干细胞群体分化为组成中枢神经系统的逐渐更专业的细胞类型来实现的。单个神经元和神经胶质细胞的独特性是由基因表达的组合模式决定的；基因表达在很大程度上受DNA序列(顺式调控元件，遗传)和染色质结构(表观遗传)的控制。转录调控的关键成分之一是增强子，这是一种通常在进化上保守的非编码DNA序列。当反式作用因子结合时，增强子确定特定基因的组织或细胞类型特异性表达。我们将从全基因组的基因表达研究中选择对NSC细胞命运决定至关重要的基因，并在细胞命运决定过程中分析非编码DNA区域在NSC基因表达中的调节功能(例如作为增强子)。我们将(在硅胶中)预测并(在体内)对预测的鸡视网膜干细胞中可能的增强剂进行功能表征。我们将这种增强剂称为NSC增强剂。我们的重点将放在对视网膜发育至关重要的NSC增强剂上。两个特定的目标是：1)预测进化上保守的NSC增强子；2)验证和鉴定可能的NSC增强子。这项拟议研究的成功完成将有助于识别对神经干细胞的细胞命运决定至关重要的转录控制网络。此外，从这项研究中发现的新型视网膜特异性增强剂可用于识别以前未知其在控制神经细胞命运决定中的功能的新的蛋白质因子。我们的发现最终将提供一个整合的转录网络，控制视网膜中NSC细胞的命运决定(通常也可以应用于其他发育系统)。这种对转录网络的了解对于开发从视网膜变性到脊髓损伤等疾病的潜在治疗或治疗性移植是至关重要的。 与公共卫生相关：这项提议的目标是寻找存在于基因组非蛋白质编码区的DNA序列元件，这些元件调节神经干细胞生成神经细胞。拟议研究的成功完成将有助于开发潜在的治疗方法或治疗性移植，以治疗从神经退化到脊髓损伤的各种疾病。

项目成果

Analysis of retinal cell development in chick embryo by immunohistochemistry and in ovo electroporation techniques.

通过免疫组织化学和卵内电穿孔技术分析鸡胚视网膜细胞发育。

• DOI: 10.1186/1471-213x-10-8
• 发表时间: 2010-01-20
• 期刊: BMC DEVELOPMENTAL BIOLOGY
• 作者: Doh, Sung Tae;Hao, Hailing;Loh, Stephanie C.;Patel, Tapan;Tawil, Haim Y.;Chen, David K.;Pashkova, Anna;Shen, Andy;Wang, Huimin;Cai, Li
• 通讯作者: Cai, Li

In ovo electroporation in embryonic chick retina.

胚胎鸡视网膜的卵内电穿孔。

• DOI: 10.3791/3792
• 发表时间: 2012
• 期刊: Journal of visualized experiments : JoVE
• 作者: Islam,MohammedM;Doh,SungTae;Cai,Li
• 通讯作者: Cai,Li