MOLECULAR BIOLOGY OF CL CHANNEL IN CYSTIC FIBROSIS

现在已知囊性纤维化是由隐性遗传引起的。 上皮细胞内阴离子转运调节的缺陷。 在这些上皮细胞中，氯化物被运输穿过细胞。 通过可由环状AMP打开的通道的膜 依赖蛋白激酶。最近，有研究表明，在囊性癌中 纤维化时，该激酶不能打开该通道，即使 膜上存在通道。 哥伦比亚长老会囊性纤维化中心由 正在合作的多学科调查小组 对这种疾病的分子缺陷进行鉴定。 我们已经纯化了上皮氯离子通道，并使用 在这个过程中，我们将克隆通道的基因并对其进行测序。 已经开发了表达离子转运的方法 青蛙卵母细胞中的蛋白质，这将有助于克隆 氯离子通道。开发了另一种表达系统，该系统 依靠DNA介导的基因转移，这将使克隆成为可能 甲状腺上皮细胞中碘和氯转运蛋白的研究 细胞。该方法还将用于识别结构和 这些基因的调节性突变体。对氯化物的监管 在人类中将研究多种蛋白质运动的通道 囊性癌患者来源的上皮细胞和In细胞 纤维化症。氯离子通道将在平面上重建 脂双层及其运动对其电生理的影响 将对其特点进行研究。 该中心将促进所有相关各方的合作 关于囊性遗传性缺陷的鉴定小组 纤维化症。

Cystic fibrosis is now known to be due to a recessive genetic defect in the regulation of anion transport in epithelial cells. In these epithelia, chloride is transported across the cell membrane through a channel that can be opened by cyclic AMP dependent protein kinase. Recently, it was shown that in cystic fibrosis, the kinase fails to open the channel, even though the channel is present in the membrane. The Columbia-Presbyterian Cystic Fibrosis Center is composed of a multidisciplinary group of investigators who are collaborating towards identification of the molecular defect in this disease. We have already purified the epithelial chloride channel and using this procedure we will clone and sequence the gene for the channel. Methods have been developed for expression of ion transport proteins in frog oocytes which will facilitate the cloning of the chloride channel. Another expression system was developed which relies on DNA-mediated gene transfer, that will allow the cloning of iodide and chloride transport proteins from thyroid epithelial cells. This method will also be used to identify structural and regulatory mutants of these genes. Regulation of the chloride channel by a variety of protein kineses will be studied in human epithelial cells and in cells derived from patients with cystic fibrosis. The chloride channel will be reconstituted in planar lipid bilayer and the effect of kineses on its electrophysiological characteristics will be studied. The Center will promote the collaboration of all the involved groups towards the identification of the genetic defect in cystic fibrosis.