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Modified nucleotide cofactors and the interaction of the RNAome with the proteome

修饰的核苷酸辅因子以及 RNAome 与蛋白质组的相互作用

基本信息

批准号：
8178871
负责人：
GARY B RUVKUN
金额：
$ 35万
依托单位：
MASSACHUSETTS GENERAL HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-09-30 至 2015-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8178871
关键词：
Affect Amino Acids Animals Bar Codes Base Pairing Binding Biology Caenorhabditis elegans Charge Chemicals Cloning Code Coenzyme A Comparative Genomic Analysis Complement Complementary RNA Coupled DNA Data Defect Disulfides Eukaryotic Cell Fostering Genome Histone H3 Libraries Ligation Link Maps Mass Spectrum Analysis Mediating Methionine MicroRNAs Modeling Modification Nucleic Acid Binding Nucleic Acid Hybridization Nucleic Acids Nucleotides Oligonucleotides Pathway interactions Peptide Fragments Peptide Hydrolases Peptides Plants Positioning Attribute Protein Analysis Protein Biochemistry Proteins Proteome Proteomics RNA RNA Binding RNA Interference Research Series Small Interfering RNA Small RNA Surveys Techniques Temperature Tertiary Protein Structure Tetrahymena alpha Tubulin cofactor comparative inorganic phosphate link protein melting nuclease sulfation

项目摘要

DESCRIPTION (provided by applicant): We have preliminary evidence that small RNAs of the size of siRNA and miRNAs are coupled to modified nucleotides such as coenzyme A and bis-MGD. These modifications would allow small RNAs to covalently engage with the proteome to constitute nucleic acid bar codes on these proteins. We propose to use powerful nucleic acid hybridization and selection techniques to purify the proteins covalently linked to RNA and to use mass spectroscopy to identify those linked proteins. Using RNAi of these proteins we will survey for defects in small RNA function and thus an involvement of these proteins in the functions of small RNAs. Because all of the modifications are predicted to occur at the 5' end of small RNAs, most of these modified RNAs will have been systematically missed by surveys most of which demands 5' OH or 5' phosphates on the RNAs for cloning. PUBLIC HEALTH RELEVANCE: Our preliminary data points to the involvement of modified nucleotides in small RNA function. The implications of this research are enormous: if such small RNAs are the active components of RNAi and miRNA function, as it pertains to the engagement with the proteome, then the field of small RNAs may have systematically missed a major world of small RNAs. In the same way that by not looking at 22 nt for biology, by not being able to detect small RNAs with 5' modifications, the small RNA field may have another dark matter situation. We will remedy this.

描述（由申请人提供）：我们有初步证据表明 siRNA 和 miRNA 大小的小 RNA 与修饰的核苷酸（例如辅酶 A 和 bis-MGD）偶联。这些修饰将使小 RNA 与蛋白质组共价结合，在这些蛋白质上构成核酸条形码。我们建议使用强大的核酸杂交和选择技术来纯化与 RNA 共价连接的蛋白质，并使用质谱来鉴定这些连接的蛋白质。使用这些蛋白质的 RNAi，我们将调查小 RNA 功能的缺陷，从而调查这些蛋白质是否参与小 RNA 的功能。由于预计所有修饰都发生在小 RNA 的 5' 末端，因此大多数需要在 RNA 上添加 5' OH 或 5' 磷酸盐进行克隆的调查将系统性地漏掉大多数这些修饰的 RNA。公共健康相关性：我们的初步数据表明修饰核苷酸参与小 RNA 功能。这项研究的意义是巨大的：如果这样的小RNA是RNAi和miRNA功能的活性成分，因为它涉及与蛋白质组的结合，那么小RNA领域可能会系统地错过小RNA的主要世界。就像生物学上不关注 22 nt，无法检测到 5' 修饰的小 RNA 一样，小 RNA 领域可能会出现另一种暗物质情况。我们会解决这个问题。