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Genome wide identification and functional analysis of chromatin regulatory RNAs

染色质调节 RNA 的全基因组鉴定和功能分析

基本信息

批准号：
10062511
负责人：
William James Greenleaf
金额：
$ 61.6万
依托单位：
STANFORD UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-12-01 至 2022-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10062511
关键词：
Binding Proteins Binding Sites Biology Cell Maintenance Cell physiology Cells Chimera organism Chromatin Chromosomal RNA Chromosome Mapping Chromosome Structures Chromosomes Clustered Regularly Interspaced Short Palindromic Repeats Code DNA DNA Sequence Data Defect Development Dosage Compensation (Genetics)Drosophila genus Drosophila melanogaster Female Functional disorder Gene Expression Genes Genetic Transcription Genome Genomic Imprinting Genomic Segment Human Human Chromosomes Human Development Human Genome Infrastructure Libraries Link Maps Messenger RNA Metabolic Methods Mouse Cell Line Nuclear Output Phenotype Play Process Proteins RNA RNA Binding Regulation Resources Role Route Salvelinus Sampling Site Stress Structure System Technology Testing Time Transcriptional Regulation Untranslated RNA Validation X Chromosome base cell type comparative developmental disease ds-DNA experimental study fly gene repression genome-wide genome-wide analysis genomic locus histone modification human disease human tissue knock-down male next generation sequencing novel novel strategies protein expression response single cell analysis stressor transcriptome sequencing tumor progression

项目摘要

In addition to the transcription of protein coding genes in the genome, a large amount of transcription encodes RNA molecules that do not generate mRNA. These noncoding RNAs play important roles in the cell that include regulating dosage compensation, controlling genomic imprinting and regulating transcription. However, human cells transcribe thousands of noncoding RNAs and we have only ascribed functions to a small number. One of the main challenges to understanding the functions of noncoding RNAs is that technologies to rapidly identify and characterize noncoding RNAs are lacking. In this proposal, we develop a novel method that makes it possible to identify, in any cell type, all of the noncoding RNAs that interact with chromosomes and at the same time map the sites where those RNAs bind chromatin. Our approach involves directly linking noncoding RNAs to the underlying DNA by generating a covalent chimera between a chromosome bound RNA and DNA. Using next generation sequencing, we can identify the RNAs in the cell that are likely to regulate chromosome structure or function and define their sites of action on the chromosome. In our first Aim we use Drosophila cells to develop this approach, taking advantage of the fact that established chromosomal RNAs, roX1 and roX2, are known to coat the X chromosome to accomplish dosage compensation in the fly. We then broaden this approach in Aim 2 and identify the RNAs that bind chromatin throughout the human genome and develop a new analytical infrastructure to classify and functionally assign these RNAs. In Aim 3 develop perturbation experiments to test the functions of noncoding RNAs and RNA motifs for their impact on local chromosome accessibility, histone modification state and transcriptional output. We apply a system to redirect noncoding RNAs to new genomic regions to test their functional impact on chromosomes and to regulate different genomic regions through RNA dependent control. By defining the landscape of chromatin associated RNAs in humans and the sites that they regulate in the cell our proposal how these RNAs function as well as the impacts of defects in RNA dependent control that result in cellular dysfunction.

除了基因组中蛋白质编码基因的转录外，还有大量的