Characterization of the promoter region of the cystic fibrosis transmembrane conductance regulator gene.
Characterization of the promoter region of the cystic fibrosis transmembrane conductance regulator gene.
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DOI：
10.1016/s0021-9258(18)54252-6
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发表时间：
1991-12
期刊：
影响因子：
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通讯作者：
J. Chou;R. Rozmahel;L. Tsui
中科院分区：
文献类型：
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作者：
J. Chou;R. Rozmahel;L. Tsui
为了鉴定囊性纤维化跨膜传导调节因子（CFTR）基因的转录调节元件，将位于5 '上游区域的DNA片段与细菌氯霉素乙酰转移酶（CAT）报告基因融合，并转染到各种细胞系中以测试启动子活性。这些研究结果表明，至少有两个积极的和一个消极的顺式作用元件参与CFTR转录起始。其中之一是由转录起始位点上游约226个碱基的5'缺失构建体界定的近端正元件。这种最小启动子序列（-226至+98）单独似乎足以指导细胞特异性CAT表达。另一方面，紧邻-227上游的序列似乎含有负调控元件;将该序列与近端元件（例如含有序列-345至+98的构建体）包含在一起使得CFTR启动子失活。这种负调控元件也可以抑制异源启动子的活性。此外，DNA转染研究表明，在所检测的CFTR启动子区域外存在另一个正调控元件，因为该区域（例如-658至+98）在CAT测定中不能发挥功能，可以通过病毒增强子元件的存在来克服。
To identify the transcription regulatory elements of the cystic fibrosis transmembrane conductance regulator (CFTR) gene, DNA fragments located in the 5‘-upstream region were fused with the bacterial chloramphenicol acetyltransferase (CAT) reporter gene and transfected into various cell lines to test for promoter activity. The results of these studies suggested that there were at least two positive and one negative cisacting elements involved in CFTR transcription initiation. One of them was a proximal, positive element delimited by the 5‘ deletion constructs -226 base parts upstream of the transcription start site. This minimal promoter sequence (-226 to +98) alone seemed to be sufficient to direct cell-specific CAT expression. The sequences immediately upstream of -227, on the other hand, appeared to contain a negative regulatory element; inclusion of this sequence with the proximal element (e.g. a construct containing sequences -345 to +98) rendered the CFTR promoter inactive. This negative regulatory element could also suppress the activity of a heterologous promoter. In addition, the DNA transfection study suggested the existence of another positive regulatory element outside the CFTR promoter region examined, as the inability of this region (e.g. -658 to +98) to function in a CAT assay could be overcome by the presence of a viral enhancer element.