Genomewide discovery & analysis of alternative promoters

全基因组发现

• 批准号: 7033451
• 负责人: RAMANA V DAVULURI
• 金额: $ 32.5万
• 依托单位: OHIO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-04-27 至 2010-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7033451
• 关键词: chromatin; clinical research; genes; genome; human

DESCRIPTION (provided by applicant): Promoters located at the 5'-ends of genes play a critical role in regulating transcriptional initiation. Emerging evidence suggests that a significant fraction of approximately 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 approximately 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.

描述（由申请人提供）：位于基因5 '端的启动子在调节转录起始中起关键作用。新出现的证据表明，大约30，000个人类基因中的很大一部分可能含有替代启动子，这些启动子在不同组织，细胞类型和/或发育阶段中产生更精细的基因表达调控。尽管人类基因组序列有大量的信息，但仍然缺乏用于鉴定和表征基因位点的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正向同源序列。尚未彻底探索基因座的5 '端调控区在人类和啮齿动物之间显示出多大程度的序列相似性。我们假设基因转录所需的基础（核心）启动子区域在人类和小鼠之间是保守的。计算方法将用于挖掘人类和小鼠基因组，以识别功能性直系同源序列。衍生的信息将被添加到一个原型数据库的哺乳动物启动子，称为MPromDb，由我们开发。接下来，将对这些计算得出的序列进行实验验证。我们进一步假设功能性基因组序列处于常染色质环境中，该环境呈现开放的染色质构型，允许转录因子进入以驱动基因表达。而不是采取通常的路线来分析基因表达，我们将确定一个测试基因组序列的染色质状态作为一个功能性启动子的措施。染色质免疫沉淀（ChIP）微阵列，或所谓的ChIP芯片上的测定，以前在我们的实验室开发的将扩展到同时评估约1,500个假定的启动子的常染色质状态的250对人类和小鼠的orthopathic基因。计算，统计和高通量实验方法的组合，提出了在这个补助金申请将有助于更好地表征基因调控区域在人类基因组中，未来基因组研究的重大挑战之一。具体而言，我们将：（1）开发计算工具，用于注释实验已知的替代启动子和第一外显子。(2)进行ChIP芯片和荧光素酶测定以验证人类和小鼠直链淀粉基因的计算注释的替代启动子序列，以及（3）开发检测人类和小鼠基因组中的替代启动子和第一外显子的计算方法。