IN VIVO PHOSPHORYLATION SITES IN CYSTIC FIBROSIS TRANSMEMB CONDUCTANCE REGULATO

囊性纤维化跨膜电导调节中的体内磷酸化位点

• 批准号: 7355045
• 负责人: DAVID C GADSBY
• 金额: $ 0.37万
• 依托单位: ROCKEFELLER UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-03-01 至 2007-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7355045
• 关键词: VIVO; PHOSPHORYLATION; SITES; CYSTIC; FIBROSIS

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Inherited mutations in the chloride channel known as the cystic fibrosis transmembrane conductance regulator (CFTR) lead to the symptoms of cystic fibrosis. To become open, the channel must be phosphorylated by cAMP-dependent protein kinase at sites within its regulatory domain. Previous mass spectrometric experiments have identified numerous serine residues in the regulatory domain that are phosphorylated in vitro, and have allowed determination of the relative ease of in vitro phosphorylation at the various sites. The current work examines human CFTR expressed in Xenopus oocytes to determine the in vivo phosphorylation pattern. It was observed that Ser-768 of CFTR was essentially quantitatively phosphorylated in vivo. Other work in the Gadsby lab has shown that mutation of this residue to alanine, thus preventing its phosphorylation, results in a constitutively active ion channel. Thus phosphorylation of Ser-768 appears to be a negative regulator of channel opening. Other sites of in vivo serine phosphorylation were characterized, and examination of the in vivo phosphorylation state of CFTR containing the S768A mutation gave no evidence of a change in phosphorylation pattern at the other sites within the regulatory domain.

该子项目是利用NIH/NCRR资助的中心赠款提供的资源的许多研究子项目之一。子弹和调查员（PI）可能已经从其他NIH来源获得了主要资金，因此可以在其他清晰的条目中代表。列出的机构适用于该中心，这不一定是调查员的机构。被称为囊性纤维化跨膜电导调节剂（CFTR）的氯化物通道中的遗传突变导致囊性纤维化的症状。为了开放，必须在其调节域内的位点依赖CAMP依赖性蛋白激酶磷酸化通道。先前的质谱实验已经确定了在调节结构域中的许多丝氨酸残基，它们在体外被磷酸化，并允许确定各个部位的体外磷酸化的相对易于易度。当前的工作研究了在爪蟾卵母细胞中表达的人CFTR，以确定体内磷酸化模式。据观察，CFTR的Ser-768基本上是在体内定量磷酸化的。 Gadsby实验室中的其他工作表明，该残基对丙氨酸的突变，从而阻止其磷酸化，从而导致组成性活性离子通道。因此，Ser-768的磷酸化似乎是通道开口的负调节剂。还表征了体内丝氨酸磷酸化的其他部位，并检查了含有S768A突变的CFTR的体内磷酸化状态，没有证据表明调节域内其他部位的磷酸化模式发生了变化。