CYSTIC FIBROSIS TRANSMEMBRANE CONDUCTANCE REGULATOR

这个子项目是利用由NIH/NCRR资助的中心拨款提供的资源的许多研究子项目之一。子项目和调查员(PI)可能从另一个NIH来源获得了主要资金，因此可能会出现在其他CRISE条目中。列出的机构是针对中心的，而不一定是针对调查员的机构。囊性纤维化是一种常染色体隐性遗传病，由囊性纤维化跨膜电导调节蛋白(CFTR)突变引起，CFTR是多种器官系统上皮细胞中的主要氯离子通道。在慢性阻塞性肺疾病患者中，氯离子跨上皮细胞膜转运的严重减少导致阻塞性肺疾病、慢性鼻窦炎、胰腺功能不全、肠梗阻和男性不育。虽然基因型-表型相关性表明，临床表现中的一些可变性是由于CFTR型的特定突变，但在相同突变的患者中存在极端的变异性，这表明CFTR型以外的因素可能对疾病严重程度起作用。通过上游信号事件调节正常CFTR功能的基因是CF的合理候选修饰物。CAMP受体的上游调控依赖于cAMP，最近的研究表明，cAMP依赖的蛋白激酶A通过磷酸化cAMP介导的氯离子转运。在上皮细胞中，cAMP水平受细胞外激动剂通过G蛋白偶联跨膜受体的影响。因此，CFTR的活性可以受到G蛋白偶联信号过程的影响，G蛋白偶联信号过程激活腺酰环化酶并产生cAMP。我们有兴趣研究这一上游激活途径中的基因在调节CFTR的功能，进而调节疾病严重程度中的作用。为此，我们建议研究与这一途径相关的基因发生严重改变的患者的CFTR功能。奥尔布赖特遗传性骨营养不良症(Aho)患者已被证明对β-肾上腺素能刺激的反应降低了cAMP水平。在过去的十年中，这被证明是由于GNAS1的突变所致，GNAS1是编码异源三聚体G蛋白(Gs-α)的刺激性阿尔法亚单位的基因。GNAS1基因突变的患者具有特征的生理表型，包括身材矮小、指趾短、肥胖、圆形相和皮下骨化。这些患者中的一部分也有激素抵抗，经典的定义是肾脏对甲状旁腺激素的反应降低，但许多患者也被证明对通过Gs-α发出信号的其他激素有抵抗。这种激素抵抗的确切机制尚不清楚，因为同一个家族中携带相同GNAS1突变的患者在激素抵抗方面可能有不同的表型。我们有关于一名AHO患者的初步数据，他对鼻黏膜上皮和皮肤中的β-激动剂的CFTR反应降低。这表明改变的Gs-α功能和降低的CFTR功能之间存在联系。为了更好地了解cAMP依赖通路在调节CFTR功能中的作用，我们建议对AHO患者的CFTR功能进行表征。通过检测上皮组织中GNAS1 RNA水平和cAMP水平，我们将确定cAMP依赖途径的预期通量水平。然后，将在这些相同的组织中评估CFTR的功能。这项研究的结果将确定cAMP依赖的通路在CFTR功能中的作用，这可能指导未来CF患者的治疗选择。它还可能解决AHO患者激素抵抗的机制，因为这将是第一个直接在体内表征GNAS1突变对其一个细胞靶点的影响的研究。来自家庭成员的结果将有助于AHO的基因型-表型相关性，并澄清来自家族变异的功能缺陷。

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. Cystic Fibrosis is (CF) is an autosomal recessive disorder caused by mutations in the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR), the primary chloride (Cl-) channel in epithelial cells of numerous organ systems. In CF patients, a severe reduction in Cl- transport across epithelial cell membranes results in obstructive pulmonary disease, chronic sinusitis, pancreatic insufficiency, intestinal obstruction, and male infertility. While genotype-phenotype correlations have shown that some of the variablility in clinical presentation is due to specific mutations in CFTR, extreme variability exists among patients with identical mutations suggesting factors other than CFTR genotype may contribute to disease severity. Genes regulating normal CFTR function via upstream signaling events are rational candidate modifiers of CF. Upstream regulation of CFTR is cAMP dependent, and recent studies have shown CFTR mediated Cl- transport is modulated through phosphorylation by cyclic AMP (cAMP)-dependent protein kinase A. In epithelial cells cAMP levels are influenced by extra-cellular agonists via G-protein coupled transmembrane receptors. Thus, CFTR activity can be affected by G-protein coupled signaling processes that activate adenylyl cyclase and which generate cAMP. We are interested in investigating the role of genes in this upstream activation pathway in modulating the function of CFTR, and in turn disease severity. To this end, we propose to study CFTR function in patients with severe alterations in genes involved in this pathway. Patients with Albright's Hereditary Osteodystrophy (AHO) have been shown to have decreased cAMP levels in response to beta-adrenergic stimulation. Over the last decade this has been shown to be due to mutations in GNAS1, the gene that encodes the stimulatory alpha subunit of heterotrimeric G proteins (Gs-alpha). Patients with mutations in GNAS1 have a characteristic physical phenotype including short stature, brachydactyly, obesity, rounded facies, and subcutaneous ossifications. A subset of these patients also have hormone resistance, classically defined as decreased renal response to parathyroid hormone, but many patients have also been documented to have resistance to other hormones that signal via Gs-alpha. The exact mechanism for this hormone resistance is unclear, as patients within the same family and carrying the same mutation in GNAS1 can have variable phenotypes with regard to hormone resistance. We have preliminary data on one patient with AHO who has a decreased CFTR response to beta-agonists in the nasal epithelia and the skin. This suggests a link between altered Gs-alpha function and decreased CFTR function. To better understand the role of the cAMP-dependent pathway in modulating CFTR function, we propose to characterize CFTR function in patients with AHO. By examining GNAS1 RNA levels and cAMP levels in epithelial tissues, we will define the expected level of flux through the cAMP dependent pathway. CFTR function will then be assessed in these same tissues. The results of this study will define the role of the cAMP-dependent pathway in CFTR function, which may guide future therapeutic options for patients with CF. It may also address the mechanism of hormone resistance in AHO patients since it will be the first study directly characterizing, in vivo, the effects of GNAS1 mutation on one of its cellular targets. Results from family members will aid in genotype-phenotype correlations in AHO and clarify functional defects from familial variation.