Regulation of CFTR gene expression

CFTR 基因表达的调控

• 批准号: 6640838
• 负责人: Martin John Walsh
• 金额: $ 29.66万
• 依托单位: ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-07-05 至 2005-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6640838
• 关键词: DNA footprinting; acetylation; acyltransferase; amidohydrolases; cAMP response element binding protein; cell cycle; cell differentiation; cell line; chloride channels; chromatin; cyclic AMP; cystic fibrosis; deoxyribonuclease I; enzyme mechanism; genetic promoter element; genetic transcription; histones; immunoprecipitation; molecular pathology; protein protein interaction

Cystic fibrosis (CF), an autosomal recessive disorder, is caused by a dysfunctional cystic fibrosis transmembrane conductance regulator (CFTR) gene product. The CFTR gene (CFTR) is regulated by promoter sequences that give rise to low levels of tissue-specific gene transcription. The overall objective of this proposal is to determine the mechanisms that direct transcription of CFTR to learn how CFTR transcription is related to the pathophysiology of CF. The acetylation of core histones within the nucleosome is associated with the formation of "open" chromatin, necessary for transcriptional activation. Furthermore, histone deacetylation is associated with repressive, or "closed", chromatin structure. The alteration in chromatin is critical for the control of gene transcription in vivo. We have demonstrated that CFTR transcription is directly associated with histone acetyltransferase (HAT) and histone deacetylase (HDAC) activity, but it is unclear how histone acetylation regulates CFTR transcription. We propose that histone acetylation direct the transcription of CFTR. Currently, the mechanisms that direct histone acetylation to a specific gene and that regulate gene transcription are not well understood. First, we wish to test the hypotheses that regulation of CFTR transcription in vivo is mediated through histone acetylation. Based on our previous work we will examine whether transcription factors ATF1 and CBF/NF-Y and the histone acetyltransferase coactivator proteins, p300/CREB-binding protein (p300/CBP) and p300/CBP-associated factor (PCAF) mediate histone acetylation of CFTR. We will determine the function of histone acetylation in the regulation of CFTR transcription. CFTR is a target of histone acetylation when the CFTR promoter contains an inverted CCAAT (Y-box) element in vivo. Both CFTR transcript initiation and cAMP-mediated CFTR transcription regulation require the Y-box element, suggesting that this element, by interacting with specific transcription factors, may be essential to the modification of chromatin in CFTR. Second, we will test the hypothesis that CCAAT displacement protein (CDP) recruits HDACs to CFTR in vivo and is regulated by acetylation. CDP, a critical trans-acting regulator of CFTR transcriptional repression, is directly associated with a co-repressor complex composed of specific HDAC activity. CDP interacts with the co-activator p300/CBP. Furthermore, CDP is regulated by acetyltransferase (FAT) activities of p300/CBP and PCAF. We plan to test the role of CDP as a substrate for acetyltransferase and deacetylase function and to determine whether the acetylation of CDP effect transcription of CFTR.

囊性纤维化是一种常染色体隐性遗传病，由囊性纤维化跨膜传导调节因子基因产物功能障碍引起。CFTR基因受启动子序列的调控，从而导致低水平的组织特异性基因转录。这个建议的总体目标是确定引导CFTR转录的机制，以了解CFTR转录是如何与CF的病理生理相关的。核小体内核心组蛋白的乙酰化与“开放”染色质的形成有关，这是转录激活所必需的。此外，组蛋白去乙酰化与抑制或“封闭”染色质结构有关。染色质的改变是体内基因转录调控的关键。我们已经证明了组蛋白乙酰转移酶(HAT)和组蛋白脱乙酰基酶(HDAC)活性与CFTR转录直接相关，但组蛋白乙酰化如何调控CFTR转录尚不清楚。我们认为组蛋白乙酰化指导了cftr的转录。目前，将组蛋白乙酰化到特定基因并调节基因转录的机制尚不清楚。首先，我们希望验证体内CFTR转录调控是通过组蛋白乙酰化介导的假设。在我们以前工作的基础上，我们将检测转录因子atf1和CBF/NF-Y以及组蛋白乙酰转移酶共激活蛋白p300/CREB结合蛋白(p300/CBP)和p300/CBP相关因子(PCAF)是否介导组蛋白乙酰化。我们将确定组蛋白乙酰化在cftr转录调控中的作用。在体内，当cftr启动子含有倒置的CCAAT(Y-box)元件时，cftr是组蛋白乙酰化的靶点。无论是cftr转录起始还是cAMP介导的cftr转录调控都需要Y-box元件，提示该元件通过与特定的转录因子相互作用，可能对cftr中染色质的修饰起重要作用。其次，我们将验证CCAAT置换蛋白(CDP)在体内将HDAC招募到CFTR并受乙酰化调控的假设。CDP是cftr转录抑制的关键反式作用调节因子，它直接与由特定的HDAC活性组成的辅助抑制物复合体有关。CDP与共激活子p300/CBP相互作用。此外，CDP还受p300/CBP和PCAF的乙酰转移酶(FAT)活性的调节。我们计划测试CDP作为乙酰转移酶和脱乙酰酶功能底物的作用，并确定CDP的乙酰化是否影响CFTR的转录。