The Role of Alternative Splicing Factor Sfrs10 in Neural Development

选择性剪接因子 Sfrs10 在神经发育中的作用

• 批准号: 7788630
• 负责人: RAHUL N KANADIA
• 金额: $ 9万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-02-01 至 2010-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7788630
• 关键词: Affect; Alternative Splicing; Alzheimer' s Disease; Antibodies; Architecture; Autistic Disorder; Bioinformatics; Biological Assay; Biological Models; Birth Order; Brain; Cell Separation; Cells; Central Nervous System Part; Code; Complementary DNA; Complex; DNA; Data; Defect; Development; Disease; Drosophila genus; Electroporation; Embryo; Event; Exclusion; Exons; Fluorescence; Genes; Goals; House mice; Human; Human Genome; In Vitro; Infection; Knockout Mice; Lead; Link; Mammals; Masks; Messenger RNA; Motor Neurons; Mus; Mutation; Myotonic Dystrophy; Neural Retina; Neuraxis; Neuroglia; Neurons; Nonsense Codon; Northern Blotting; Pathogenesis; Pathway interactions; Pattern; Play; Process; Protein Isoforms; Proteins; Proteome; RNA; RNA Interference; RNA Splicing; Reporting; Research; Research Proposals; Resolution; Retina; Retinal; Reverse Transcriptase Polymerase Chain Reaction; Role; Set protein; Spinal Muscular Atrophy; Structure; Techniques; Technology; Tissues; Tretinoin; Ultrasonography; Vertebrates; Virus; Work; base; cell type; design; gain of function; ganglion cell; glutamate receptor type B; human fetus tissue; in vivo; injured; insight; interest; loss of function; nervous system development; neurodevelopment; numb protein; postnatal; public health relevance; relating to nervous system; research study; retinal progenitor cell; sex determination; tool

DESCRIPTION (provided by applicant): Through alternative splicing a single gene can generate functionally diverse set of proteins. Indeed, AS is invoked to reconcile the difference between the relatively few (~24,000) protein coding genes in the human genome to its vast proteome. Interestingly, of all the mammal tissues, the central nervous system (CNS) has the highest degree of AS. Thus, it is generally accepted that AS plays a crucial role in neural development, but there is a paucity of information on this issue. Given the complexity of CNS development combined with that of AS, we have chosen to employ the neural retina as our model system. The retina is derived from the CNS, is a relatively simple tissue, has a well defined laminar structure, it has six neural cell types and one glia, birth order of each cell type is known, and importantly it is the most accessible part of the CNS. The overall goal of this research proposal is to investigated the contribution of AS in neural development by studying an AS factor, Sfrs10 in the mouse retina. Previous studies in Drosophila have shown that Sfrs10 is required during sex determination and others have shown that Sfrs10 regulates the AS some neural genes. Based on this and our preliminary work the hypothesis underlying this proposal is that Sfrs10 is essential for neural cell fate determination and differentiation. There are two specific aims and the first specific aim will accomplish the following goals. We will characterize the expression pattern and AS of Sfrs10 during embryonic and postnatal retinal development. In addition, loss (RNAi) and gain of function will be performed by either in vivo DNA electroporation of a postnatal day 0 mouse retina or by in vitro electroporation of embryonic retinal explant cultures. We will employ ultrasound guided delivery technology to deliver viruses with either a gain or a loss of function construct into E10.5 embryo. A significant effort will be dedicated towards finding the targets of Sfrs10. Moreover, a conditional knockout mouse will be generated to validate the aforementioned experiments and provide a tool for the second specific aim. The second specific aim will investigate the AS status of the targets of Sfrs10 at the single cell resolution. The premise here is that tissues consists of several cell types and if each cell type splices an exon differently, then the use of entire tissue for analysis might mask this interesting fact. PUBLIC HEALTH RELEVANCE: Understanding the role of alternative splicing in development is essential to our understanding the underlying mechanism that leads to diseases. There are several diseases such as Alzheimer's disease, myotonic dystrophy, spinal muscular atrophy and autism that are linked to defects in alternative splicing of specific genes or are caused by mutations in genes that regulate the process of alternative splicing.

描述(申请人提供)：通过选择性剪接，单个基因可以产生一组功能不同的蛋白质。事实上，AS被用来协调人类基因组中相对较少的(约24,000)个蛋白质编码基因与其庞大的蛋白质组之间的差异。有趣的是，在所有哺乳动物组织中，中枢神经系统(CNS)的AS程度最高。因此，人们普遍认为AS在神经发育中起着至关重要的作用，但关于这一问题的信息很少。考虑到CNS开发的复杂性和 因此，我们选择使用神经视网膜作为我们的模型系统。视网膜起源于中枢神经系统，是一种相对简单的组织，具有明确的板层结构，它有六种神经细胞类型和一种神经胶质细胞，每种细胞类型的出生顺序是已知的，重要的是它是中枢神经系统最容易接近的部分。这项研究计划的总体目标是通过研究小鼠视网膜中的AS因子Sfrs10来研究AS在神经发育中的作用。以前对果蝇的研究表明，性别决定过程中需要Sfrs10，其他研究表明，Sfrs10作为一些神经基因来调节。基于这一点和我们的初步工作，这一提议背后的假设是，Sfrs10对神经细胞命运的决定和分化是必不可少的。有两个具体目标，第一个具体目标将实现以下目标。我们将描述Sfrs10在胚胎和出生后视网膜发育过程中的表达模式和AS。此外，功能的丧失(RNAi)和功能的获得将通过出生后第0天的小鼠视网膜的体内DNA电穿孔或胚胎视网膜外植体培养的体外电穿孔来执行。我们将使用超声引导传递技术将功能构建获得或丧失的病毒输送到E10.5胚胎中。将作出重大努力，寻找瑞士法郎10的目标。此外，还将生成一个有条件的基因敲除小鼠来验证上述实验，并为第二个特定目的提供工具。第二个具体目标将调查AS的状况 在单胞分辨率下的目标为10瑞士法郎。这里的前提是，组织由几种细胞类型组成，如果每种细胞类型拼接的外显子不同，那么使用整个组织进行分析可能会掩盖这一有趣的事实。 公共卫生相关性：了解选择性剪接在发育中的作用对于我们理解导致疾病的潜在机制至关重要。有几种疾病，如阿尔茨海默病、强直性肌营养不良症、脊髓性肌萎缩症和自闭症，与特定基因选择性剪接缺陷有关，或由调节选择性剪接过程的基因突变引起。

项目成果

Expression analysis of an evolutionarily conserved alternative splicing factor, Sfrs10, in age-related macular degeneration.

• DOI: 10.1371/journal.pone.0075964
• 发表时间: 2013
• 期刊: PloS one
• 影响因子: 3.7
• 作者: Karunakaran DK;Banday AR;Wu Q;Kanadia R
• 通讯作者: Kanadia R

The expression analysis of Sfrs10 and Celf4 during mouse retinal development.

Sfrs10和Celf4在小鼠视网膜发育过程中的表达分析。

• DOI: 10.1016/j.gep.2013.07.009
• 发表时间: 2013
• 期刊: Gene expression patterns : GEP
• 作者: Karunakaran,DeviKrishnaPriya;Congdon,Sean;Guerrette,Thomas;Banday,AbdulRouf;Lemoine,Christopher;Chhaya,Nisarg;Kanadia,Rahul
• 通讯作者: Kanadia,Rahul