Evolution of Enhancer Function in the Immunoglobulin Heavy Chain Gene

免疫球蛋白重链基因增强子功能的进化

• 批准号: 9807531
• 负责人: Gregory Warr
• 金额: $ 33.4万
• 依托单位: Medical University of South Carolina
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-09-15 至 2002-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9807531&HistoricalAwards=false
• 关键词: Evolution; Enhancer; Function; Immunoglobulin; Heavy

9807531 Warr Transcriptional control is now recognized to be important for all aspects of immunoglobulin (Ig) gene expression: that is, enhancer function is important not just for driving functional expression of the gene, but it is also essential for the processes of V (D) J recombination, heavy chain class switching, and somatic hypermutation of the Ig genes to occur. The channel catfish provides a well-developed and unique model to study the function of Ig genes at the phylogenetic level of teleost fish: the immunobiology of this species has been extensively studied and well documented, and the Ig genes (for mu, delta, and L chains) have all been cloned. The overall objective of the project is to understand the function of the IgH enhancer of the channel catfish, which has been shown in prior studies to occupy an unusual position in the locus, and to possess an unusually diffuse structure. The following questions will be addressed in this project. 1) Which motifs in the catfish enhancer are important in driving transcription? 2) What is the role played by Oct1 in driving expression from the promoter-associated octamer found in all VH genes (including those of the catfish)? Is a co-activator such as Bob 1 involved in the function of Oct1 and Oct2 in the catfish? 3) What is the nature of the factors (other than octamer transcription factors) that may drive transcription from the catfish enhancer? The approaches to be employed include site-directed mutagenesis to delete individual or multiple motifs (such as octamer, muE3, muE5) from the enhancer to determine their contribution to function, and the cloning of transcription factors (such as Oct1) and the co-activator Bob-1 (a co-activator of Oct transcription factors) from the catfish, in order to define their roles in driving transcription of the Ig genes at this level of evolution. Antibodies have two polypeptide chains, a heavy chain and a light chain. The expression of heavy chain genes occurs largely at the level of transcript ion, the process by which information in DNA is copied into mRNA. Transcription is regulated by sequences in the DNA, called enhancers, which interact with protein transcription factors. This project is investigating the enhancers and transcription factors that regulate the expression of antibody heavy chains in catfish.

转录控制现在被认为对免疫球蛋白（Ig）基因表达的各个方面都很重要：也就是说，增强子功能不仅对驱动基因的功能性表达很重要，而且对V (D) J重组、重链类转换和Ig基因的体细胞超突变的发生也很重要。通道鲶鱼为研究硬骨鱼系统发育水平上Ig基因的功能提供了一个成熟而独特的模型：该物种的免疫生物学已被广泛研究和充分记录，并且Ig基因（mu链，delta链和L链）都已被克隆。该项目的总体目标是了解通道鲶鱼的IgH增强子的功能，在先前的研究中已经表明，它在基因座中占据了一个不寻常的位置，并具有一个不寻常的弥散结构。以下问题将在这个项目中解决。1)鲶鱼增强子中哪些基序在驱动转录中是重要的？2) Oct1在驱动所有VH基因（包括鲶鱼基因）中发现的启动子相关八聚体的表达中起什么作用？像Bob 1这样的辅助激活因子是否参与了鲶鱼Oct1和Oct2的功能？3)可能驱动鲶鱼增强子转录的因子（八聚体转录因子除外）的性质是什么？所采用的方法包括位点定向诱变，从增强子中删除单个或多个基序（如octamer， muE3, muE5）以确定它们对功能的贡献，以及从鲶鱼中克隆转录因子（如Oct1）和共激活因子Bob-1 （Oct转录因子的共激活因子），以确定它们在这种进化水平上驱动Ig基因转录的作用。抗体有两条多肽链，一条重链和一条轻链。重链基因的表达主要发生在转录离子水平，即DNA中的信息被复制到mRNA的过程。转录受DNA中称为增强子的序列调控，增强子与蛋白质转录因子相互作用。本项目旨在研究鲶鱼抗体重链表达的增强子和转录因子。