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In Vivo Characterization of Major ENCODE-Predicted Classes of Noncoding Elements

主要编码预测非编码元素类别的体内表征

基本信息

批准号：
10241190
负责人：
Len Alexander Pennacchio
金额：
$ 135.88万
依托单位：
UNIVERSITY OF CALIF-LAWRENC BERKELEY LAB
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：
2017-02-01 至 2022-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10241190
关键词：
Alleles Biochemical Biological Biological Assay Biology Boundary Elements CRISPR/Cas technology Categories Chromatin Clustered Regularly Interspaced Short Palindromic Repeats Collection Communities Complex DNA DNA Methylation Data Data Set Development Disease Elements Embryonic Development Encyclopedias Engineering Enhancers Excision Gene Expression Gene Transfer Techniques Genes Genetic Enhancer Element Genetic Transcription Genomics Goals Gold Health Human Human Genome Human body In Vitro Individual Knock-in Knock-in Mouse Knock-out Knockout Mice Laboratories Lead Link Methods Modeling Molecular Molecular Profiling Mus National Human Genome Research Institute Organ Organism Phase Phenotype Process Property Regulatory Element Reporter Research Personnel Resources Scheme Technology Testing Time Tissues Transgenic Mice Transgenic Organisms Untranslated RNA Variant Work base epigenomics experimental study genome editing genome-wide genomic data human disease in vivo insight mouse genome programs protective allele risk variant synergism tool transcriptome transcriptome sequencing

项目摘要

PROJECT SUMMARY We propose to establish a Center for In Vivo Characterization of ENCODE Elements (CIViC) as part of ENCODE Phase 4. Understanding the function of the 98% of the human genome that is noncoding remains one of the most pressing challenges in genomics. The ENCODE Program has enabled major progress toward obtaining genome-wide molecular signatures associated with the human and mouse genome. During ENCODE3 our group contributed to the mapping of enhancer-associated marks, DNA methylation, and transcriptomes from multiple mouse tissues across closely spaced time points of embryogenesis, resulting in >750 datasets defining the in vivo epigenomic landscape during mammalian development. Our group has also characterized over 3,000 candidate enhancer sequences in transgenic mouse assays, including more than 400 through our participation in ENCODE2 and ENCODE3. Despite this progress, enhancers are only one of many noncoding molecular functions that have been inferred from ENCODE data. Other major proposed categories of noncoding sequences identified through ENCODE and other publicly available data sets include DNA elements with predicted functions, such as “super-enhancers” (very large enhancers with possibly distinct functions) or chromatin domain boundary elements. They also include sequence classes of unknown function primarily defined by specific assays, such as differentially methylated regions (DMRs). The functional impact of these different classes of noncoding sequences on organismal biology and human health remains minimally explored, representing a major limitation of the ENCODE encyclopedia. The Center for In Vivo Characterization of ENCODE Elements will use CRISPR/Cas9 genome editing to systematically explore the biological significance of several classes of noncoding function based on ENCODE3 data. Leveraging the streamlined set of mouse engineering tools available in our laboratory, we will: 1. Perform integrative analysis of ENCODE3 and complementary data sets to identify and prioritize representative sequences from 3 different classes of noncoding elements (enhancers and super-enhancers, boundary elements, DMRs); 2. Systematically delete a total of 48 representative sequences in mice and perform RNA-seq and gross organismal phenotyping to understand the in vivo consequences of these deletions; 3. Continue to make transgenic enhancer characterization capabilities available to ENCODE investigators to validate and calibrate enhancer prediction methods. We will also use transgenics and CRISPR knock-in editing to test human disease-associated alleles of ENCODE-predicted enhancer elements. All efforts will be closely coordinated with other ENCODE4 functional characterization groups to focus on common sets of elements to be characterized using the full ENCODE-wide arsenal of in vitro and in vivo characterization methods. Our results will provide an understanding of the in vivo significance of different classes of noncoding elements and thereby substantially increase the value of the ENCODE encyclopedia.

项目总结