Genomewide discovery & analysis of alternative promoters

全基因组发现

基本信息

批准号：
7371108
负责人：
RAMANA V DAVULURI
金额：
$ 2.07万
依托单位：
OHIO STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2006
资助国家：
美国
起止时间：
2006-04-27 至 2008-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7371108
关键词：
Antibodies Applications Grants Automobile Driving Biological Assay Chromatin Code Computing Methodologies DNA Data Databases Development Environment Euchromatin Exons Future Gene Expression Gene Expression Regulation Genes Genetic Transcription Genome Genomics Heterochromatin Histones Human Human Genome Laboratories Length Luciferases Lysine Maps Measures Mining Mus Nucleic Acid Regulatory Sequences Orthologous Gene Pattern Play Polymerase Chain Reaction Promoter Regions Proteins Research Research Personnel Rodent Role Route Sampling Specific qualifier value Staging Testing Tissue Sample Tissues Transcription Initiation Transcription Initiation Site cell type chromatin immunoprecipitation computerized tools data mining functional genomics genome sequencing improved mammalian genome mouse genome novel programs promoter prototype tool transcription factor

项目摘要

Promoters located at the 5'-ends of genes play a critical role in regulating transcriptional initiation. Emerging evidence suggests that a significant fraction of -30,000 human genes likely contain alternative promoters, which produce more elaborate regulation of gene expression in different tissues, cell-types and/or developmental stages. Despite vast information available for the human genome sequences, a comprehensive approach for identifying and characterizing alternative promoters of gene loci is still lacking. One effective approach is to analyze orthologous sequences in both mouse and human genomes. It has not been thoroughly explored as to what extent the 5'-end regulatory regions of a locus show sequence similarity between human and rodents. We hypothesize that the basal (core) promoter regions necessary for gene transcription are conserved between human and mouse. Computational approaches will be used to mine both human and mouse genomes to identify functional orthologous sequences. The derived information will be added into a prototype database for mammalian promoters, called MPromDb, developed by us. Next, these computationally derived sequences will be experimentally verified. We further hypothesize that a functional genomic sequence is in a euchromatic environment that presents an open chromatin configuration, allowing for the access of transcription factors for driving gene expression. Instead of taking the usual route to analyze gene expression, we will determine the chromatin status of a test genomic sequence as a measure for a functional promoter. The chromatin immunoprecipitation (ChIP) microarray, or the so-called ChlP-on-chip assay, previously developed in our laboratory will be extended to simultaneously assess the euchromatin status of ~1,500 putative promoters of 250 pairs of human and mouse orthologous genes. The combination of computational, statistical and highthroughput experimental approaches proposed in this grant application will help better characterize the gene regulatory regions in the human genome, one of the grand challenges of future genome research. Specifically we will: (1) Develop computational tools for annotating experimentally known alternative promoters and first exons. (2) Conduct ChlP-on-chip and luciferase assays to verify computationally annotated alternative promoter sequences of human and mouse orthologous genes, and (3) Develop computational methods to detect alternative promoters and first exons in the human and mouse genomes.

位于基因 5' 端的启动子在调节转录起始中发挥着关键作用。新兴有证据表明 -30,000 个人类基因中的很大一部分可能含有替代启动子，其在不同组织、细胞类型和/或中产生更精细的基因表达调节发展阶段。尽管人类基因组序列有大量信息，仍然缺乏识别和表征基因位点替代启动子的综合方法。一种有效的方法是分析小鼠和人类基因组中的直系同源序列。它没有已深入探索基因座 5' 端调控区在多大程度上显示出序列相似性人类和啮齿动物之间。我们假设基因必需的基础（核心）启动子区域人类和小鼠之间的转录是保守的。计算方法将用于挖掘人类和小鼠基因组以确定功能性直系同源序列。得出的信息将被添加到我们开发的哺乳动物启动子原型数据库中，称为MPromDb。下一个，这些计算得出的序列将得到实验验证。我们进一步假设功能基因组序列处于常染色质环境中，呈现开放染色质配置，允许访问转录因子来驱动基因表达。而不是采取分析基因表达的常用途径，我们将确定测试基因组的染色质状态序列作为功能启动子的测量。染色质免疫沉淀 (ChIP) 微阵列，或我们实验室之前开发的所谓 ChlP-on-chip 检测将扩展到同时进行评估 250 对人类和小鼠直系同源基因的约 1,500 个推定启动子的常染色质状态基因。提出了计算、统计和高通量实验方法的结合在这项拨款申请中，将有助于更好地表征人类基因组中的基因调控区域，其中一个未来基因组研究的巨大挑战。具体来说，我们将：（1）开发计算工具注释实验已知的替代启动子和第一个外显子。 (2) 进行片上 ChLP 和荧光素酶测定验证人类和小鼠的计算注释替代启动子序列直系同源基因，以及 (3) 开发计算方法来检测替代启动子和第一个外显子在人类和小鼠基因组中。