Na, K-ATPase Regulation by Glucocorticoids in Lung

糖皮质激素对肺中 Na、K-ATP 酶的调节

• 批准号: 6833440
• 负责人: CHRIS H WENDT
• 金额: $ 33.41万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-12-15 至 2007-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6833440
• 关键词: DNA; SDS polyacrylamide gel electrophoresis; cell line; chromatin immunoprecipitation; epithelium; gel mobility shift assay; genetic transcription; glucocorticoids; heat shock proteins; laboratory rat; lung; mass spectrometry; molecular chaperones; nucleic acid sequence; polymerase chain reaction; site directed mutagenesis; sodium potassium exchanging ATPase; tissue /cell culture; transcription factor; western blottings

DESCRIPTION (provided by applicant): To allow for normal gas exchange at birth, the ion transport functions of the lung must change dramatically as the fetus transits into postnatal life. Late in gestation, as the fetus approaches birth, the lung changes from a chloride secreting to a sodium-absorbing organ. This results in the net resorption of salt and water from the alveolar airspace and prepares the fetus for the successful transition to the postnatal period in which the alveolus is dry and normal gas exchange takes place. This resorptive process occurs through the active transport of sodium via the basolateral Na, K-ATPase (sodium pump) and the passive transport of sodium, primarily through the apical amiloride-sensitive sodium channel. During late gestation, glucocorticoids influence the expression of the Na, K-ATPase gene in a manner that is dependent on the gestational age at the time of glucocorticoid administration, and the duration and dose of glucocorticoid therapy. The major effect of glucocorticoids is on the beta1 subunit, demonstrating a concomitant increase in gene expression, protein levels and activity as measured by wet to dry weights. Glucocorticoids regulate the Na, K-ATPase both transcriptionally and translationally depending on cell type. We found that glucocorticoids increase Na, K-ATPase gene expression via transcriptional regulation in fetal and adult lung epithelial cells, however, the regulatory elements and specific transcription factors involved in this upregulation remain unknown. Although the Na, K-ATPase beta1 promoter contains partial glucocorticoid receptor elements (GRE), few of these elements are classical GRE. In addition, upregulation by glucocorticoids occurs over hours, suggesting regulation may be through a unique delayed primary response that involves secondary proteins. The main objective of this grant is to extend our previous studies of pump transcriptional regulation by glucocorticoids and explore in vitro mapping of DNA regulatory elements and transcription factor identification that are critical for Na, K-ATPase beta1 regulation comparing fetal to adult lung epithelial cells. I have chosen to study the Na, K-ATPase beta1, since: 1) glucocorticoids increase beta1 protein levels concomitant to increased mRNA levels; 2) the Na, K-ATPase beta1 subunit is the rate limiting subunit in lung epithelial cells; and 3) the Na, K-ATPase beta1 subunit is necessary for the assembly of alpha-beta heterodimers and plasma membrane targeting.

描述（由申请人提供）：为了在出生时进行正常的气体交换，随着胎儿过渡到出生后，肺的离子转运功能必须发生显著变化。 在妊娠后期，随着胎儿接近出生，肺从一个分泌氯化物的器官转变为一个吸收钠的器官。 这导致盐和水从肺泡气隙的净再吸收，并为胎儿成功过渡到出生后时期做好准备，在出生后时期，肺泡干燥并进行正常的气体交换。 该吸收过程通过基底外侧Na，K-ATP酶（钠泵）主动转运钠和主要通过顶端阿米洛利敏感钠通道被动转运钠而发生。 在妊娠晚期，糖皮质激素影响Na，K-ATP酶基因的表达，其方式取决于糖皮质激素给药时的胎龄以及糖皮质激素治疗的持续时间和剂量。 糖皮质激素的主要作用是对β 1亚基，表明基因表达、蛋白质水平和活性（通过湿重至干重测量）同时增加。 糖皮质激素调节Na，K-ATP酶的转录和转录依赖于细胞类型。 我们发现糖皮质激素通过转录调控增加胎儿和成人肺上皮细胞Na，K-ATP酶基因的表达，然而，参与这种上调的调控元件和特异性转录因子尚不清楚。 虽然Na，K-ATP酶β 1启动子含有部分糖皮质激素受体元件（GRE），但这些元件中很少有经典的GRE。 此外，糖皮质激素的上调发生在数小时内，这表明调节可能是通过一个独特的延迟初级反应，涉及二级蛋白。 这项资助的主要目的是扩展我们以前的研究泵转录调节糖皮质激素和探索在体外映射的DNA调控元件和转录因子的鉴定，这是至关重要的Na，K-ATP酶β 1调节比较胎儿和成人肺上皮细胞。 我选择研究Na，K-ATP酶β 1，因为：1）糖皮质激素增加β 1蛋白水平，同时增加mRNA水平; 2）Na，K-ATP酶β 1亚基是肺上皮细胞中的限速亚基; 3）Na，K-ATP酶β 1亚基是α-β异二聚体组装和质膜靶向所必需的。