MOLECULAR GENETICS OF CYSTIC FIBROSIS

囊性纤维化（CF）是最常见的严重常染色体隐性遗传病 在高加索人群中。在北美，大约每2500人中就有1人 活产受CF影响，其特征在于慢性 阻塞性肺疾病，胰腺功能不全， 外分泌腺的电解质、液体和大分子分泌。的 基本的生化缺陷是未知的。CF基因的鉴定， 提供了一个机会，了解缺陷和病理生理学 在分子水平上的疾病，这将使发展 理性治疗根据DNA序列分析， 产物（CFTR）被预测为具有2 ATP结合域。基因分析表明，大约70%的 CF染色体遭受3个碱基对缺失，这对应于单个 在CFTR的位置508处的氨基酸缺失。五个具体目标是 在本申请中提出：（1）CF基因中的另外的突变将 为了绘制CFTR的功能结构域而进行鉴定;简单 将为每种突变制定检测程序; 已经在剩余的30%的CF中发现了突变， 染色体(2)突变和“表位标签”将被引入到 选择CFTR的区域，以映射其功能结构域和拓扑结构， 与哺乳动物细胞质膜的关系。(3)的效果 一些天然突变的CFTR的生物合成， 特别是已经检测到的无义和移码突变， 将研究CF中是否存在真正的“无效”突变。（四） 为了了解控制CF基因表达的因素， 负责基础启动子活性的序列元件， CFTR的组织特异性将通过常规技术确定 使用报告基因构建体（CAT）;转基因小鼠携带 CF基因启动子和报告基因（E. coli lacZ）也将 研究CFTR的发育调控。(5)DNA序列 在初步研究中发现并显示， 与人类基因组中CFTR相关的基因将被分离和表征， 关于它们的染色体位置、表达模式和它们的 与CFTR的结构和功能关系。后一种信息 可能为CF基因的进化功能提供重要的见解 和其他类似CFTR的基因。这些研究与2项平行进行， 其他单独资助的方法（使用酵母和小鼠模型）， 在阐明基本缺陷方面， 在CF。

Cystic fibrosis (CF) is the most common severe autosomal recessive disorder in the Caucasian population. In North America, approximately 1 in 2,500 live-births is affected with CF, which is characterized by chronic obstructive lung disease, pancreatic insufficiency, abnormalities of electrolyte, fluid and macromolecule secretion of exocrine glands. The basic biochemical defect is unknown. The identification of the CF gene has provided an opportunity to understand the defect and the pathophysiology of the disease at the molecular level, which will allow the development of rational therapy. On the basis of DNA sequence analysis, the CF gene product (CFTR) is predicted to be a transmembrane protein with 2 ATP-binding domains. Genetic analysis shows that approximately 70% of the CF chromosomes suffer a 3 base pair deletion which corresponds to a single amino acid deletion at position 508 of CFTR. Five specific aims are proposed in this application: (1) Additional mutations in the CF gene will be identified in order to map the functional domains of CFTR; simple detection procedures will be developed for each mutation; a number of mutations have already been identified for the remaining 30% of CF chromosomes. (2) Mutations and "epitope tags" will be introduced into selected regions of CFTR to map its functional domains and topology in relation to the plasma membrane in mammalian cells. (3) The effect on the biosynthesis of CFTR of some of the naturally occurring mutations, especially the nonsense and frameshift mutations that have been detected, will be studied to investigate if true "null" mutations exist in CF. (4) In order to understand the factors governing CF gene expression, the sequence elements responsible for basal promoter activity and tissue-specificity for CFTR will be determined by conventional techniques with the use of reporter gene constructs (CAT); transgenic mice carrying the CF gene promoter and a reporter gene (E. coli lacZ) will also be constructed to study developmental regulation of CFTR. (5) DNA sequences that have been detected and shown in preliminary studies, to be closely related to CFTR in the human genome will be isolated and characterized, with respect to their chromosomal location, expression pattern, and their structural and functional relationships with CFTR. The latter information may provide important insight into the evolution function of the CF gene and other CFTR-like genes. These studies, conducted in parallel with 2 other separately funded approaches (use of yeast and mouse models), constitute a comprehensive research program in elucidating the basic defect in CF.